Exam 2: Biochemistry Flashcards

Question 1

Q

Cell

Compartmentalization

Answer

A

Topologically equivalent spaces:

Nucleus & Cytosol

Perinuclear cistern, ER cisterna, Golgi cisterna, Lysosomes, Transport vesicles & Endosomes

Movement between topologically inequivalent spaces requires translocators.

Question 2

Q

Cellular Transport

Mechanisms

Answer

A

Gated transport
Transmembrane transport
Vesicular transport

Question 3

Q

Gated Transport

Answer

A

Large openings act as selective gates.

Between topographically equivalent spaces ⇒ does not cross a membrane.

Ex. nucleus ↔︎ cytosol

Question 4

Q

Transmembrane Transport

Answer

A

Between topologically inequivalent spaces ⇒ crosses a membrane
Uses translocators
- dependent on targeting signals
- protein moved in a denatured form
Ex.
- import of nascent peptides into RER
- import from cytosol into mitochondria and peroxisomes

Question 5

Q

Vesicular Transport

Answer

A

Between topologically equivalent spaces when each is membrane bound
Transport vesicles carry proteins and membranes
Anterograde → “forward”
- ER to Golgi
Retrograde → “backward”
- Golgi → ER
- Endosomes → Golgi
Ex.
- ER → Golgi
- Golgi → lysosomes / plasma membrane
- Endocytosis & Exocytosis

Question 6

Q

Vesicle Structure

Answer

A

Inner layer formed from adaptor proteins links outer layer (cage) to the membrane
Cage proteins cover cytosolic surface forming a coat
- Assembly requires energy & GTP binding proteins
- Functions:
  - collect specific membrane and soluble cargo
  - direction formation of vesicles
- Removed before vesicles fuse
- Differs depending on destination and direction of movement

Question 7

Q

COPII Coating

Answer

A

Coats anterograde transport vesicles from ER → Golgi.

Question 8

Q

COPI Coating

Answer

A

Coats retrograde transport vesicles from the Golgi → ER.

Question 9

Q

Clathrin Coating

Answer

A

Transport vesicles from Golgi → endosomal compartments / plasma membrane.

Transport vesicles from plasma membrane → endosomes.

Question 10

Q

Vesicle Movement

Answer

A

Budding and targeting uses movement along cytoskeletal tracks:

Microtubules using kinesin and dynein

Actin filaments using Myosin II or Myosin V

Motor proteins recruited by Rab proteins.

Question 11

Q

Vesicle Targeting

Answer

A

Docking and fusion mediated by SNARE proteins.

SNARE proteins on transport vesicle bind complementary SNARE proteins on the target membrane.

Forces two membranes close together so lipid bilayers can fuse.

Question 12

Q

Genetic Code

Definition

Answer

A

The sequence relationship between the bases in the gene or mRNA and the amino acid in the protein.

3 consecutive nucleotides on mRNA ⇒ codon

Question 13

Q

Genetic Code

Characteristics

Answer

A

64 possible combinations of the 4 bases
- 61 code for AA
- 3 code for stop codons
Codons are contiguous and do not overlap
Degenerate ⇒ multiple codons can code for a single AA
Unambiguous ⇒ each codon codes for one AA

Question 14

Q

Wobble Hypothesis

Answer

A

The pairing between the codon and anticodon adheres to the usual base-pairing rules at the first two bases but is less strict for the third.

First two bases predominate in tRNA selection
Some tRNAs can bind to more than one codon that codes for the same AA

Question 15

Q

Start Codon

Answer

A

AUG ⇒ Methionine

N-terminal methionine formylated in prokaryotes ⇒ fMet

Question 16

Q

Stop Codons

Answer

A

UAA ⇒ U are awful

UAG ⇒ U are gross

UGA ⇒ U go away

Question 17

Q

Rules of

Protein Production

Answer

A

Anticodon of tRNA pairs with codon of mRNA in anti-parallel fashion
mRNA read in 5’ ⇒ 3’ direction
Codons read sequentially by charged tRNAs
Proceeds from N-terminus ⇒ C-terminus

Question 18

Q

Protein Synthesis

Stages

Answer

A

Activation of amino acid
Chain initiation
Chain elongation
Chain termination
Co/post translational processing

Question 19

Q

tRNA Structure

Answer

A

Acceptor end links the 5’ and 3’ ends forming clover structure.

3’-OH terminal CCA has AA attached to acceptor site.

Anticodon triplet base pairs with mRNA codon.

Question 20

Q

Amino Acid

Activation

Answer

A

“Charging the tRNA”

Catalyzed by aminoacyl-tRNA synthetase.

20-different aminoacyl tRNA synthetases recognizes ONE amino acid and ALL its cognate tRNAs
Traps energy from hydrolysis of ATP → AMP + PP_i in AA~AMP complex
- Two high energy bonds from ATP required
Forms high energy bond between tRNA and AA used later to link AA to polypeptide chain
Proofing mechanism to ensure correct AA attached

Question 21

Q

Ribosome Structure

Answer

A

Each subunit contains 3 critical sites:

A site ⇒ accepts the incoming aminoacylated tRNA

P site ⇒ holds tRNA and has ribosomal peptidyl transferase to form peptide bond

E site ⇒ temporarily holds deacylated tRNA until it exits the ribosome

Question 22

Q

Small Ribosomal Subunit

Functions

Answer

A

Formation of the initiation complex
Decodes the genetic information i.e. reads mRNA
Binds both the 5’ end of mRNA and the tRNA-amino acid complex at the loading site
Controls the fidelity of codon-anticodon pairing

Question 23

Q

Large Ribosomal Subunit

Functions

Answer

A

Contains ribosomal peptidyl transferase activity that joins the AA to the polypeptide chain
Contains translocation domain
Contains tunnel where nascent peptide threaded

Question 24

Q

Translation Initiation

Question 25

Q

eIF2

Regulation

Answer

A

Activated by guanine nucleotide exchange factor.

Replaces GDP with GTP.

Inactivated by a GTPase.

Under conditions of stress, several kinases phosphorylate eIF2.

Phosphorylated eIF2 cannot bind gunine nucleotide exchange factor.

Low [ternary complex] ⇒ increased activation of amino synthesis genes.

Question 26

Q

mTOR

Answer

A

Mechanistic Target of Rapamycin

Serine/threonine kinase.

Major growth factor and nutrient sensor in the cell.

Able to modulate translational activity.

Activated via phosphorylation by insulin or IGF-1 via protein kinase B (Akt) pathway.

Also activated by nutrients, especially leucine.

Question 27

Q

eIF4 Regulation

Answer

A

Resources are plentiful
- mTOR activated by protein kinase B (Akt) pathway
  - Initiated by Insulin or IGF-1, leucine
- Activated mTOR phosphorylates 4E-BP1 (eIF4 binding protein)
- Phosphorylated 4E-BP1 releases eIF4E which can join the eIF4 complex
- Translation proceeds
Resources are scarce
- ↑ [AMP] ⇒ AMP kinase activation
- AMPK inhibits mTOR
- 4E-BP1 remains unphosphorylated
- eIF4E remains sequestered by 4E-BP1
- Translation is blocked

Question 28

Q

Translation Elongation

Answer

A

Depends on formation of peptide bond.

Aided by G-protein elongation factors (EFs).

Requires 2 ATP (charge tRNA) and 2 GTP (EFs) per cycle.

4-step repetitive process:

Charged tRNA brought into A site
- Aminoacyl-tRNA recruited by eEF1-GTP
- eEF1-GTP hydrolyzed by GTPase to eEF1-GDP and recycled.
Amino acid attached to nascent polypeptide chain located in the P site by ribosomal peptidyl transferase
- Ribozyme component of large ribosomal subunit
Ribosome advances 3 nucleotides towards 3’ end of mRNA ⇒ translocation
- Promoted by GTP-dependent eEF2
  - Target for diphtheria toxin
Empty tRNA moved to the E-site

Question 29

Q

Translation Termination

Answer

A

Release factors (RFs) recognize the stop codon and bind to the A site mimicking a tRNA
Peptidyl transferase hydrolyzes the completed peptide chain from the final tRNA in the P site
Hydrolysis of two GTP ⇒ GDP
RFs use energy from GTP hydrolysis to induce conformation change in ribosome causing release of nascent polypeptide through exit tunnel
mRNA released
Large and small ribosomal subunits dissociate

Question 30

Q

Protein Processing

Overview

Answer

A

Co-translational vs Post-translational

Can occur in the cytoplasm or other cellular organelles.

Protein folding
Covalent modifications
- Attachment of sugars, phosphate groups, or lipids
- Linked by disulfide bridge
Cleavage
Multiple subunits linked into quaternary structure

Question 31

Q

Protein Folding

Answer

A

Determined mostly by primary sequence of the peptide.

Primarily involves non-covalent interactions between AA side chains:

Hydrogen bonding

Van der Waals interactions

Electrostatic interactions

Native-like secondary structures assemble into domains ⇒ molten globules.

Aided by chaperones.

Question 32

Q

Chaperones

Answer

A

Binds to “sticky” hydrophobic patches on nascent polypeptide chains.
Prevents non-productive folding pathways or aggregation.
Helps protein to fold by stabilizing intermediate conformations.
Helps with creation of complexes.
Stabilizes proteins as they move through intracellular organelles.
Ex. Heat shock proteins

Question 33

Q

Disulfide Bond

Formation

Answer

A

Formed between the thiol groups of cysteine residues as proteins fold in the RER.

Catalyzed by protein disulfide isomerase.

Intramolecular disulfide bonds:

Within a single polypeptide chain
Usually stabilize the tertiary structure

Intermolecular disulfide bonds:

Between seperate polypeptide chains
Usually stabilize quarternary structure

Question 34

Q

Unfolded Protein Response (URP)

Mechanism

Answer

A

UPR normally inhibited when chaperone supply adequate.

“Extra” chaperones bind to sensing receptors in ER lumen.

When [misfolded proteins] ↑ or too many proteins made, “extra” chaperones dissociate from sensing receptors for use in the cell.

Sensing receptors become activated.

Unfolded protein response begins:

↓ protein synthesis
↑ chaperone production ⇒ ↑ capacity of ER to fold proteins
Continued problem w/ refolding ⇒ aggregation/accumulation of abnormal proteins ⇒ destruction of unfolded proteins
Failure to resolve high levels of unfolded proteins ⇒ apoptosis

Question 35

Q

Proteolytic Cleavage

Answer

A

Important step in the maturation processing of many proteins.

Proteases cleave by hydrolysis reaction.

Removal of N-terminus signal sequence
- Cleaved by signal peptide peptidase
- Associated wih the translocon
Removal of N-terminus initiator methionine
- Allows for addition of acetyl group or fatty acid chains
Cleavage of precursor proteins
- Activates zymogens

Question 36

Q

Insulin Processing

Answer

A

On RER: Insulin synthesized as preproinsulin
In ER: met_i and signal sequence removed converting to proinsulin
- Proinsulin folds, stabilized by two interchain disulfide bonds
In Golgi, proinsulin packaged into vesicles
In vesicles: internal sequence removed (C peptide or connecting sequence)
Regulated secretion of insulin and C peptide

Question 37

Q

Trypsin

Question 38

Q

Functions of Glycosylation

Answer

A

↑ solubility to prevent agglutination
Protects against proteolysis
Helps folding and oligomerization
Role in cellular sorting
Recognition and antigenicity

Question 39

Q

N-linked Glycosylation

Answer

A

Carb attached to N-atom in side chain of asparagine.

Proteins mostly remain in ER or go to Golgi for export.

asn-X-thr/ser motif
14 sugar residue transferred from dolichol phosphate ⇒ NH2 on Asn en bloc by protein-oligosaccharyl transferase
- On RER and associated w/ protein translocator
Sugars added/removed as protein moves ER ⇒ Golgi
Two categories w/ same 5 sugar core:
- Complex oligosaccharides
  - diverse sugars
- High-mannose oligosaccharides
  - mostly mannose

Question 40

Q

O-linked Glycosylation

Answer

A

Carb attached to O-atom in side chain of serine or threonine.

Much less common.

Added in the Golgi
GalNAc is the first sugar
Added one at a time by glycosyltransferases
Only a few residues
Length varies

Question 41

Q

Cytosolic Glycosylation

Answer

A

Proteins made by free ribosomes.
Simpler modifications.
Sugar moieties are not premade.
Transferred as a precursor oligosaccharide.

Question 42

Q

Phosphorylation

Answer

A

Reversible post-translational +/- of phosphoryl groups.

Kinases ⇒ add

Phosphatases ⇒ remove

Serine/Threonine Kinases

Tyrosine Kinases

Question 43

Q

Protein Degradation

Answer

A

Regulate 1/2 life
- Papidly degraded proteins usu. regulatory
  - Transcription factors, signaling molecules, cytokines
Defective/damaged cells removed
Ubiquitin-proteasome and lysosomal pathways

Question 44

Q

Ubiquitin-Proteasome Pathway

Answer

A

Cytosolic pathway

Selective
ATP dependent
Quantitatively more significant
Polyubiquitinated protein targeted to proteasome
- Ubiquitin ⇒ 76 AA polypeptide

Question 45

Q

Ubiquitination Steps

Answer

A

E1 hydrolyzes ATP and adenylates ubiquitin
Adenylated ubiquitin transferred to E2 via thioester bond
E3 (Ubiqitin ligases) recognizes appropriate proteins and transfers ubiquitin
≥ 4 Ubiquitin added ⇒ proteasome degradation

Question 46

Q

N-end rule

Answer

A

“stabilizing” AA on N-terminal like Met ↑ 1/2 life

“destabilizing” AA on N-terminal like Arg ↓1/2 life

Question 47

Q

PEST Sequences

Answer

A

1+ internal PEST sequences target proteins for rapid degradation

Question 48

Q

Lysosomal Pathway

of

Protein Degradation

Answer

A

Lysosomes have non-specific proteases called cathepsins
- Acid hydrolases
Degrades mostly ingested proteins
Role in cellular turnover

Question 49

Q

Cystic Fibrosis

Answer

A

∆F508 deletion
Misfolding ⇒ ubiquitination ⇒ proteasomal degradation

Question 50

Q

Prokaryotic

Protein Synthesis

Answer

A

Simpler with fewer factors
Ribosomal subunits smaller
Uses ATP to charge tRNA but not for protein synthesis
No mRNA modifications or splicing
Transcription & translation coupled
mRNA polycistronic
Uses purine-rich Shine-Dalgarno sequence in small ribosomal subunit to position mRNA, AUG, and tRNA_i^Met
N-formylmethionine (fMet) is the first AA
- Recognized by body as foreign

Question 51

Q

Prokarytote Translation

Initiation

Answer

A

Oriented using a Shine-Delgarnosequence

Positions mRNA on small ribosome
Purine-rich
6-10 bases upstream from initiating AUG

Question 52

Q

Streptomycin

(and related aminoglycosides)

Answer

A

High concentrations

Binds S-12 of bacterial small ribosomal subunit
Interferes with normal binding of fMet-tRNA_i^Met to P site
Inhibits translation initiation

Low concentrations

Causes misreading of mRNA
Results in the wrong AA being inserted

Resistant strains of bacterial have altered ribosome that prevent drug binding.

Question 53

Q

Tetracycline

Answer

A

Binds to the small bacterial ribosomal subunit.

Interefers with binding of incoming aminoacyl-tRNA to the A-site.

Inhibits translation elongation.

Question 54

Q

Erythromycin

Answer

A

Interacts with large subunit of baterial ribosome.

Sterically hinders the exit tunnel.

Prevents release of nascent polypeptide.

In resistant strains, a single base of rRNA methylated and drug cannot bind.

Question 55

Q

Chloramphenicol

Answer

A

Inhibits peptidyl transferase activity.

Prevents transfer of growing peptide chain onto next AA residue.

Affects both prokaryotic and mitochondrial protein synthesis.

Reserved for severe cases of infection.

Question 56

Q

Diphtheria Toxin

Mechanism

Answer

A

Toxin binds to plasma membrane and enters the cell.
Catalyzes ADP ribosylation of EF-2 ⇒ inhibits translocation activity
Inhibits elongation phase of protein synthesis

Question 57

Q

Co-Translational Targeting

Overview

Answer

A

All translation of nuclear genes begins on free ribosomes in cytosol.

Presence of ER-targeting signal on protein’s N-terminus targets peptide/ribosome pair to ER ⇒ becomes fixed.

Location of translation determines where protein will be ultimately trafficked and likely types of protein processing.

Question 58

Q

Signal Sequence / Leader Sequence

Answer

A

16-30 AA residue at N-terminus of protein

Has a hydrophobic core of 6-12 AA and one or more positively charged AA

Cleaved upon entry into the ER

Question 59

Q

Translocon

Answer

A

Water-filled channel in the ER membrane through which the nascent polypeptide chain passes.

Question 60

Q

Translational ER Targeting

Mechanism

Answer

A

Translation starts on free ribosome in cytosol
Signal sequence on N-terminal emerges from ribosome
Interacts with signal recognition particle (SRP)
- Temporarily stops translation ⇒ elongation arrest
SRP targets ribosome-nascent chain complex to a docking protein on cytosolic face of RER
Upon docking, GTP ⇒ GDP promotes insertion of nascent peptide chain into open translocon channel
SRP released
- Requires GTP hydrolysis
Signal sequence removed by signal peptidase
Elongation resumes
When translation complete, ribosome released and translocon closes

Question 61

Q

Fates of Proteins

Synthesized on RER

Answer

A

Resident ER proteins
Formation of integral/transmembrane proteins
Default pathway
- No additional ER specific signals
- Moved to Golgi for ultimate constitutive secretion
- Additional tagging can target proteins to lysosomes or regulated secretory pathway

Question 62

Q

ER Resident Protein

Targeting

Answer

A

Possess retrieval signal sequences.

Allows proteins to be retrieved from Golgi and returned to the ER.

Most common is C-terminal KDEL (Lys-Asp-Glu-Leu)

Travel in COPI coated vesicles.

Ex. Disulfide isomerase

Question 63

Q

Intergral Membrane Protein

Targeting

Answer

A

Assembled at the RER
- Majority are targeted and inserted into ER during synthesis
Includes a stop transfer sequence
Transmembrane domain
- hydrophobic core
- can be a single domain or cross >20x
Translocon
- Can open the pore so polypeptide chain translocated into the ER lumen
- Can open laterally so hydrophobic regions enter the lipid bilayer
Proteins can remain localized in ER membrane or transported in vesicles to other cellular membranes
- Default is the plasma membrane

Question 64

Q

Golgi Targeting

Answer

A

Travel from ER ⇒ cis face of Golgi in COPII coated vesicles
Move from cis ⇒ trans face to reach Trans-Golgi Network (TGN)
- Various modifications occur during transit
Proteins sorted and placed into different vesicles in TGN
- Depends on tagged signal sequences

Answer 63

A

Completion of N-linked glycosylation and O-linked glycosylation
- By glycosyl transferases
Glycosaminoglycan chains added to core proteins ⇒ proteoglycans

Answer 64

A

Mannose-6-phosphate (M6P) ⇒ lysosomes
- In clathrin coated vesicles
Signals directing to secretory vesicles ⇒ regulated secretory pathway
- In specialized secretory cells
Secreted and plasma membrane proteins selectively directed to apical or basolateral domains
- In polarized cells
- Requires specific signals

Answer 65

A

Proteins segregated and concentrated into clathrin-coated vesicles within the trans-Golgi network (TGN).

Constitutive Pathway

Default pathway
Delivered from TGN ⇒ plsama membrane constitutively unless diverted elsewhere or retained in Golgi
Operates in all cells
- Many soluble proteins
- Supplies plasma membrane with new components

Regulated Secretion Pathway

Proteins diverted to secretory vesicles
- Concentrated and stored
Extracellular signal stimulates secretion
In specialized secretory cells only
Small molecules can be actively transported from cytosol into preformed secretory vesicles via similar mechanism
- Ex. Histamine and neurotransmitters
- Often bound to specific macromolecules to store at high concentration without extra osmotic pressure
  - Ex. Histamine ↔︎ Proteoglycans

Answer 66

A

Membrane-bound compartments inside cell.

Major sorting compartment.

Molecules transported from trans-Golgi membrane ⇒ endosomes in clathrin coated vesicles.

Classified as early, sorting, or late depending on stage post-internalization.

Endosomes can continue to develop into lysosomes or recycle back to Golgi.

Endocytotic vesicles can enter pathway ⇒ lysosomes or return to plasma membrane.

Answer 67

A

Protein undergo N-linked glycosylation in ER lumen
Terminal mannose residue phosphorylated by phosphotransferase in Golgi ⇒ mannose-6-phosphate
M6P recognized by specific Golgi receptors
Receptor-protein complex buds off in clathrin-coated vesicle
Vesicle fuses with endosomes
Low pH ⇒ dissociation of glycoprotein from M6P receptor
Vesicle w/ fully processed lysosomal enzymes bud off from endosomes & fuse with lysosomes

Answer 68

A

AR lysosomal storage disease

Deficient phosphotransferase ⇒ no M6P tag

Material cannot be broken down ⇒ inclusions

Answer 69

A

Proteins made by cytosolic ribosomes:

Retained in the cytosol (default pathway)

Targeted to nucelus, mitochondria, or peroxisomes by specific signals

Answer 70

A

Short stretch of basic amino acid residues.

Ex. Lysine or Arginine

Integral to the protein ⇒ not cleaved.

Usually sequestered until transport required ⇒ conformational change exposes NLS

Answer 71

A

Protein translated by free ribosomes
Nuclear localization signal bound by importin
Protein-importin complex docks and translocated through nuclear pore complex
Inside the nucleus, protein released facilitated by Ran-GTP
Ran-GTP carries importin back to cytosol
Hydrolysis of GTP releases importin

Answer 72

A

Nuclear export signals (NES) binds to exportin and Ran-GTP
Trimeric complex transported through nuclear pore
Hydrolysis of GTP in cytosol dissociates complex

Answer 73

A

Transport through nuclear pore driven by concentration gradient of Ran-GDP in cytosol and Ran-GTP in nucleus.

Answer 74

A

Due to mutations in the NLS of SRY protein.

SRY unable to enter nucleus ⇒ no testis-developmental signals

XY individuals born as phenotypic females

Answer 75

A

Amphiphatic alpha helix

Basic residues on one side
Hydrophobic residues on the other

One or more signal sequences direct protein to the co rrect mitochondrial sub-compartment.

Answer 76

A

Protein translated by free ribosomes
Chaperones bind nascent protein maintaining in unfolded state
- Requires ATP
Matrix-targeting signal sequence on N-end recognized by translocase in the outer membrane (TOM complex)
N-terminus moved into intermembrane space
Signal sequence binds translocase in the inner membrane (TIM complex)
Protein moved into the matrix
- Protein spans both matrices for a short time
Signal sequence removed by signal peptidase in the matrix
Mature protein forms within the matrix with help of chaperones
- Requires ATP

Answer 77

A

Internal signals, rather than N-terminal.

Some with stop-transfer signals.

Can target proteins to outer membrane, inner membrane, and inner membrane space.

Answer 78

A

Peroxisomal proteins selectively imported from the cytosol via transmembrane tansport.

Most with a peroxisomal targeting sequence (PTS) at the C-terminus⇒ 3-amino acid (Ser-Lys-Leu)

Some with target sequence at the N-terminus.

Import requires peroxins

Acts as both chaperones and receptor.

Answer 79

A

AR disorder

Inability to correctly target proteins to matrix of peroxisomes.

Neurological impairment leads to death at an early age.

Answer 80

A

Pinocytosis
Receptor-mediated Endocytosis
Phagocytosis

Answer 81

A

Cell ingests fluids, molecules, and particles.

Performed in virtually every cell type.

Contitutive and nonspecific.

Vesicles do not contain a clathrin coat.

Process:

Substances to be taken in contacts extracellular surface of plasma membrane.
Surface becomes indented.
Small pinocytoic vesicles dynamically form in the plasma membrane.

Answer 82

A

Cargo protein binds specific receptors forming a clathrin-coated pit
Pit buds from plasma membane forming a coated vesicle
Vesicles lose their coat within the cell and fuse with early endosomes
1. Early endosomes are the main sorting station
Endosomes mature and pH ↓
1. Patches of membrane invaginate into lumen forming intralumenal vesicles ⇒ multivesciular bodies
Ligands dissociate from receptors
Most receptors recycled to plasma membrane via transport vesicles
Ligands digested in lysosomes.

Answer 83

A

Sometimes receptors/ligands can be transcytosed to another part of the plasma membrane.

Ex. epithelial cells in lamina propria move Ab from basolateral domain to apical domain.

Answer 84

A

Ex. of receptor mediated endocytosis

LDL binds LDL receptor on cell surface
LDL/LDL receptor internalized in clathrin-coated pits ⇒ clathrin-coated vesicles
Vesicles lose coat and fuse with early endosomes
Low pH in endosome ⇒ dissociation of LDL from receptor
Continue through endosomal pathway to lysosomes
LDL hydrolyzed to free cholesterol

Answer 85

A

Phagocytic cell engulfs pathogen in phagosome
Phagosome fuses with lysosome ⇒ phagolysosome
- TACO (tryptophanaspartate-containing coat protein) coat must be removed before fusion
- Internalized membrane components recycled
Pathogen digested in lysosomes

Process called heterophagy.

Answer 86

A

ex. TB bacillus

Avoids digestion by preventing TACO coat from being removed.

Can sometimes be killed via autophagy.

Answer 87

A

Can escape from phagosomes.

Bacteria secretes a protein that destroys phagosome membrane.

Answer 88

A

Degradation pathway for cellular proteins and organelles.

3 general types:

Macroautophagy
Microautophagy
Chaperone-mediated autophagy

Answer 89

A

portions of cytoplasm or whole organelles surrounded by a vacuole
forms a double-membraned sac ⇒ autophagosome
outer membrane fuses with lysosome ⇒ autolysosome
inner membrane and contents digested

Answer 90

A

Non-specific cytoplasmic proteins enter via invagination of lysosomal membrane.

Answer 91

A

Specific proteins with targeting signals directed into lysosomes
Aided by heat-shock chaperone proteins
Requires specific receptors on lysosomal surface
Significant protein degradation mechanism in liver and kidney
Activated during starvation

Answer 92

A

Dysfunctional metabolism of sphingolipids.

Due to defects in autophagy.

Accumulation of large amounts of cholesterol and lipids in lysosomes.

Answer 93

A

Disposal of excess proteins stored in secretory vacuoles by fusion with lysosomes.

Rare process occuring in few cell types.

Happens when it will likely a long time before products needed again.

Ex. mammotrophs of anterior pituitary & prolactin.

Answer 94

A

Forms rope-like structure

Characteristic banding pattern

High tensile strength

Type I, II, and III collagens

Answer 95

A

Fibril forming

90% of body collagen

Found in:

Dermis

Tendons / Ligaments

Bone

Fascia

Organ capsules

Cornea

Answer 96

A

Forms a 3-D mesh

Ex.

Type IV Collagen ⇒ anchoring plaques in basal lamina

which connects to

Type VII Collagen ⇒ anchoring fibrils of lamina reticularis

Answer 97

A

Flexible collagens with interrupted helices.

Bind to fibrils and connects then to ECM.

Collagen types IX and XII

Answer 98

A

Triple helix formed of 2 identical α1 chains and an α2 chain.

Rich in proline and glycine
- Proline ⇒ kinks peptide chain for helix formation
- Glycine ⇒ allows tight turns
Select prolines and lysines hydroxylated
- Site for stabilizing interchain H-bonds
3-residue repeating motifs
- Gly-Pro-X
- Gly-X-Hyp

α chains ⇒ triple helix (procollagen) ⇒ tropocollagen ⇒ fibrils ⇒ fibers ⇒ fascicles ⇒ tendons/ligaments

Answer 99

A

COL genes transcribed ⇒ α1 and α2 chains
- mRNA processed
- Moved into cytosol via gated transport
mRNA translated by free ribosomes ⇒ makes preproprotein
- Signal sequence ↔︎ signal recognition particle ⇒⇒⇒ RER
- Nascent polypeptide enters translocon ⇒⇒⇒ RER lumen
- Signal peptide cleaved by signal peptidase
  - Converts preproprotein ⇒ proprotein
Select proline and lysine residues hydroxylated by two ascorbate dependent enzymes
- Pro ⇒ hydroxyproline by proline hydroxylase
- Lys ⇒ hydroxylysine by lysyl hydroxylase
Select hydroxy lysines receive O-linked glycosylation with galactose or glucose
Triple helix forms from C-terminus to N-terminus ⇒ procollagen
- Stabilized by intra- and interchain hydrogen and disulfide bonds
- Aided by chaperones
Procollagen packaged into secretory vessicles
- Constitutively secreted via default pathway
In ECM, hydrophilic N and C-terminal propeptides cleaved by procollagen peptidases ⇒ forms tropocollagen
Tropocollagen aggregate into fibrils d/t hydrophobic effect
- Fibrils stabilized by lysyl oxidase
  - Copper-requiring enzyme
  - Cross-links the fibrils via staggered covalent bonds
Fibrils aggregate to form mature collagen fibers

Answer 100

A

Dietary Vit C deficiency

Impaired function of proline and lysyl hydroxylases

Initial symptoms:
- Fatigue, malaise, gum inflammation
Progressive symptoms:
- Depression
- Swollen and bleeding gums
- Loosening or loss of teeth
- Eecchymoses
Secondary iron deficiency anemia
- Due to blood loss and ↓ nonheme iron absorption
↑ risk with malabsorption disorders, cancers, end-stange renal disease

Answer 101

A

90% of cases due to mutations in COL1A1 or COL1A2 genes.

Results in abnormal collagen structure.

Type I OI
- Mildest form
- Due to mutation resulting in abnormal protein that does not leave ER or form procollagen
  - nonsense or frameshift
Types II-IV OI
- More severe
- Caused by missense mutation replacing glycine residues
  - Alpha-chains cannot form tight turns
- Steric hinderance and bulge in triple helix during procollagen formation
- Protein degraded

Answer 102

A

Due to mutations in either collagen genes or mutations in proline/lysyl hydroxylases.

Joint hypermobility
Skin hyperextendibility
Atrophic scar formation
Arterial, intestinal, and/or uterine fragility

Answer 103

A

Mutation in gene critical for regulating copper level.

X-linked recessive disorder

Copper accumulates in kidney and intestines
Inadequate Cu in other tissues, esp brain
Impact function of copper-containing enzymes
- Lysyl oxidase involved cross-linking of collagen fibrils
Sx.
- Sparse, brittle, and twisted hair
- Failure to thrive
- Lack of muscle tone
- Seizures
- Progressive brain deterioration
Often do not live past 3 y/o

Answer 104

A

All the chemical changes that are involved in the occurance and continuance of life.

Ability to accomplish these changes at constant body temperature requires enzymatic catalysis & thermodynamic coupling of endergonic and exergonic processes.

Answer 105

A

The series of steps involved in the breakdown or synthesis of major biological constituents.

Cycle = pathway that regenerates the initial substrate

Catabolic and anabolic pathways linked via ATP.

Answer 106

A

Degradative (complex to simpler)

Oxidative

Exergonic

ATP generating

Often requires NAD⁺ or FAD

Answer 107

A

Sum of the pathways that are involved in synthesis and growth.

Reductive

Energy consuming

ATP utilization

Often requires NADPH

Answer 108

A

Expression of enthalpic change.

Gives the normalized total heat production per unit time.

Answer 109

A

Measured in a resting state (awake laying still)

Energy requirements for:

involuntary muscle work
maintenance of osmotic gradients
maintenance of body temperature
turnover and synthesis of cell constiuents

Answer 110

A

ΔG = ΔH - TΔS

H= enthalpy S= entropy T= temp in kelvins

The energy change occuring under conditions of constant pressure and temperature.

Additive function and independent of pathway.

Total ΔG of a process can be expressed as the sum of ΔG changes of individual steps.

Answer 111

A

Coupling of an exergonic and endergonic reaction so that net ΔG is negative and reaction can occur.

A common intermediate must exist.

In an enzyme catalyzed reaction, the common intermediate may not be free and may only exist on the enzyme.

Answer 112

A

The relative affinity of a molecule, atom, or ion for electrons taken under standard conditions where reactants and products are at unit activity (~1 M) then compared to the proton/hydrogen electrode (H+ and ½ H₂)

Often expressed as values corrected to pH 7 (Eº’)

Interpreted as the relative affinity of the system for electrons as compared to that of a proton:

Negative reduction potential indicates a weaker affinity for electrons than a proton.
Positive reduction potential indicates a stronger affinity.

Answer 113

A

Dependent on the ratio of oxidant to reductant agent concentration in a cell.

Answer 114

A

E = Eº’ + 2.3 RT/nF log [oxidant] / [reductant]

R = gas constant

F = Faraday constant

Relationship of reduction potential to ratio of oxidant to reductant agents in a cell.

Answer 115

A

Describes a bond which has a large negative standard Gibb’s free energy (ΔGº’) of hydrolysis

Minimum of -7.0 kcal/mol

Indicated with a ~

Answer 116

A

Contains two phosphoanhydride bonds which have a ΔGº’ of hydrolysis of approximately -7.3 kcal/mol each.
Biological utilization of these high energy bonds require the ATP form.
One or both bonds may be used in a reaction.
Phosphoryl group(s) of ATP can be transferred to acceptor molecules to generate activated intermediates for metabolism.
Can function as coenzyme-cosubstrates.

Answer 117

A

Use of both phosphoanhydride bonds of ATP is the functional equivalent of using 2 ATP’s.

Reactions coupled to the hydrolysis of the pyrophosphate (PP_i) product to 2 orthophosphates (P_i) by pyrophosphatase which drives reactions forward.

Answer 118

A

Catalyzes the reversible reaction:

AMP + ATP ⇔ 2 ADP

Answer 119

A

Total concentration of adenine nucleotide pool essentially constant, however, ratio of adenine nucleotides vary with metabolic state of the cell.

Concentrations of ATP, ADP, and AMP in a cell are in rapid equilibrium due to activity of adenylate kinase.

The cosubstrate pool communicates between and significantly influences the different pathways of the cell which utilizes those cosubstrates.

AMP level most sensitive parameter to change in pool and usually initiates the responses to decreased ATP levels.

Answer 120

A

Energy charge = ½ • ( [ADP] + 2[ATP] ) / ( [AMP] + [ADP] + [ATP])

A stoichiometric expression of the mold fraction of high energy phosphate bonds present relative to the maximal high energy bonds possible.

0 = nucleotides are totally in the form of AMP

1 = nucleotides are totally in the form of ATP

Answer 121

A

ΔG_ATP→ADP = ΔGº’_ATP→ADP + RT ln ( [ADP] x [P_i] / [ATP] )

Actual ΔG for ATP hydrolysis in a cell is dependent upon and may be calculated from the concentrations of ATP, ADP, and inorganic phosphate.

May be very different from the ΔGº’.

In a resting cell may be as high as = 15 kcal/mol which is equivalent to an energy charge of approximately 0.9.

“High energy charge”, “large negative ΔG of ATP hydrolysis”, and “resting cell” all indicate that ATP is plentiful.

Answer 122

A

Pathways for synthesis and breakdown of the same constituent are never the same, although individual steps may be reversible and utilized in both pathways.
Opposing pathways typically occur in seperate cellular compartents and/or different tissues or organs.
Regulation of pathway enzyme activity by:
- product inhibition
- allosteric regulation
- covalent regulation i.e. phosphorylation
Changes in the rate of synthesis/degradation of an enzyme or its mRNA.

Answer 123

A

Glucose transported across cell membranes by:
1. Facilitated diffusion via GLUT transports down their concentration gradients.
2. In renal and intestinal epithelium, transported against its concentration gradient by Na⁺-glucose co-transporters (SGLTs)

Answer 124

A

Glucose-dependent tissues such as RBC’s and brain have low K_m insulin-independent GLUT1 or GLUT3 transports respectively.
In peripheral tissues such as muscle which are glucose-independent, GLUT4 transports have a low KM but is insulin-dependent ⇒ allows cross regulation.
The liver which does not rely on glucose for energy, uses the GLUT2 transporters have a high K_M for glucose but is insulin-independent.
- Limits glucose uptake to conditions when blood glucose levels are high.
- Allows the transporter to act as a sensor of high blood glucose levels.

* Normal fasting blood glucose levels are 3.9 - 5.5 mmol/L.

Answer 125

A

Located in muscle and adipose tissue.
Relatively low K_m for glucose so would transport around normal fasting levels.
Insulin-dependent glucose transporter:
- When insulin is absent, the transporters are removed from the plasma membrane and sequestered into vesicles.
  - Functional but not in the membrane.
- Insulin signaling stimulates the movement of the transporter from internal stores to the plasma membrane.
In skeletal muscle, exercise stimulates GLUT4 translocation to the plasma membrane through AMP-activated protein kinase (AMPK) via unknown mechanism.
- Long-term exercise also increases the amount of GLUT4 in the muscle cell.

Answer 126

A

Binding of insulin to the α-subunit of its receptor activates a tyrosine kinase domain resulting in auto-cross-phosphorylation of tyrosine residues in the β-subunits.
Negative charge of the phosphates causes IRS (insulin receptor substrate) proteins to bind to the β-subunit.
IRS proteins phosphorylated at two Tyr residues by the kinase activity of activated insulin receptor.
Phosphorylated-IRS dissociate from the receptor then bind to and activate proteins with SH2 domains i.e. PI-3-kinase (Phosphatidylinositol-3-kinase).
PI-3-kinase phosphorylates PIP2 to PIP3.
PIP3 activates PDK-1 (phosphoinositide-dependent kinase).
PDK-1 activates downstream effectors Akt and PKB which results in the movement of GLUT4 to the cell surface in adipose and muscle, increasing glucose uptake.

Akt/PKB also:

phosphorylates and inactivates GSK3 (glycocen synthase kinase 3) resulting in increased glycogenesis.
Activate amino acid uptake and protein synthesis
Increase lipid synthesis
Inhibit gluconeogenesis
decrease cAMP levels by activating phosphodiesterase
various gene expression modulations both +/-
- Increases protein synthesis by activating a kinase (mTOR) that ultimately results in the activation of eIF4 and EF2 from protein translation.

Answer 127

A

Phosphorylation of glucose prevents back diffuse out of the cell via the transporter and commits it for use in that cell.
Hexokinases
- Found in tissues such as muscle and brain.
- Has a low K_M for glucose
  - Can phosphorylate other monosaccharides but affinity for glucose considerably higher
- Show product inhibition by glucose-6-phosphate.
Glucokinase
- Found in liver and pancreatic β-cells
- Has a high K_M for glucose
- Shows no direct product inhibition

Answer 128

A

Despite being monomeric, glucokinase displays sigmoidal kinetics towards glucose.
The inflection point of the glucokinase enzyme curve is such that small changes in blood glucose levels causes significant changes in enzymatic activity.
When blood glucose levels are high the hepatic glucokinase becomes significantly more active.
Hexokinase, however, is fully saturated a normal concentrations of blood glucose.

Answer 129

A

glucose + 2 NAD⁺ + 2 ADP + 2 P_i

→

2 pyruvate + 2 NADH + 2 ATP + 2 H₂O + 2 H⁺

Answer 130

A

Priming steps

Two ATP-linked phosphorylations
Functions to prevent diffusion of the intermediates of the pathway out of the cell.

Oxidation/reduction with the production of NADH
* Results in the generation of ATP
Re-oxidation of the NADH produced.

Answer 131

A

The synthesis of ATP involving two coupled reactions linked by a common intermediate containing a high-energy bond.

Answer 132

A

Irreversible phosphorylation of glucose at carbon-6 by hexokinase using ATP•Mg²⁺ to produce glucose-6-phosphate.
- Hepatic isozyme is glucokinase.
- Traps glucose because cell membrane is impermeable to phosphate esters.
- Commits glucose to intracellular metabolism but NOT glycolysis.
Freely reversible aldose-ketose isomerization of glucose-6-phosphate to fructose-6-phosphate by phosphoglucose isomerase.
Irreversible phosphorylation of fructose-6-phosphate to fructose-1,6-bisphosphate using ATP by phosphofructokinase (PFK1).
- True commitment step and primary regulatory site for glycolysis.
Fructose-1,6-bisphosphate split into two triose phosphates: dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP) by aldolase.
- DHAP freely interconverted to GAP by triose phosphate isomerase.
- Only GAP enters next stage of glycolysis.
Glyceraldehyde-3-phosphate dehydrogenase catalyzes the reversible oxidation of glyceraldehyde-3-phosphate to 1,3-bisphosphoglycerate.
- NAD+ converted to NADH **
Phosphate from the carboxyl group from 1,3-bisphosphoglycerate is reversibly transferred to ADP to produce ATP and 3-phosphoglycerate by 3-phosphoglycerate kinase.
- First substrate-level phosphorylation.
- Additional mutase found in erythrocytes which transfers carbonyl phosphate of 1,3-bisphosphoglycerate to carbon-2 to produce 2,3-bisphosphoglycerate (2,3-BPG).
3-phosphoglycerate converted to 2-phosphoglycerate by phosphoglycerate mutase.
2-phosphoglycerate converted to phosphoenolpyruvate (PEP) by enolase with loss of H₂O.
Phosphate of PEP is transferred to ADP by pyruvate kinase (PK) to produce pyruvate and ATP.
- Irreversible
- Has a ΔGº’ of -14 kcal/mol
NADH produced is recycled back to NAD+
- In aerobic conditions via malate shuttle.
- In anerobic conditions pyruvate converted to lactate via lactate dehydrogenase.

Answer 133

A

Most important regulatory step in glycolysis because:

It is the commitment step for the glycolytic pathway.
It is allosterically modulated by many metabolic intermediates and products.

Answer 134

A

Allosteric effectors act by influencing the equilibrium between the active and inhibited forms of the enzyme.

Binds preferentially to one form or the other and stabilizes that form.

Activators ⇒ high energy signals
- Citrate
- ATP
- Fructose-2,6-Bisphosphate***
Inhibitors ⇒ low energy signals
- ADP
- AMP

Answer 135

A

Fructose-2,6-bisphosphate (F-2,6-P₂) is a positive heterotropic effector of PFK1.

F-2,6-P₂ is synthesized and degraded by a multifunctional enzyme with both:

kinase activity ⇒ PFK2
phosphatase activity ⇒ F-2,6-Bisphosphatase

Answer 136

A

Multifunctional enzyme reponsible for synthesis and degradation of F26BP.

PFK2 & F26BPase

Phosphorylation of either domain inhibits its catalytic activity.

Liver:

PFK2 enzyme a substrate for cAMP-dependent PKA.

Phosphorylation site for liver PFK2 isozyme lies within the kinase domain.

Glucagon ⇒ phosphorylation ⇒ ↓ kinase activity & ↑ phosphatase activity ⇒ ↓ [F26BP] ⇒ ↓ PFK-1 activity

Skeletal Muscle:

PFK2/F26BPase isozyme has no phosphorylation sites.

Is not covalently regulated.

Answer 137

A

Hepatic PFK1 is primarily regulated by the [F-2,6-BP]
The primary function of F-2,6-BP in the liver is to make PFK1 sensitive to regulation by glucagon and other hormones.
The liver does not consume glucose as fuel during times of need – rather it makes glucose for use by other tissues.

Answer 138

A

Covalent Regulation

(Liver Only)

Fasted state:
- Glucagon → cAMP → PKA → phosphorylated PK ⇒ PK inhibited
High glucose:
- Insulin → ↑ phosphatase activity → dephosphorylation of PK ⇒ PK activation

Allosteric Regulation

(All cells)

Fasted state:
- Acetyl CoA
- Long chain fatty acid
- Alanine
High energy state:
- ATP
- F-1,6-BP

Answer 139

A

Glycolysis occurring under anaerobic conditions.

Answer 140

A

Pyruvate + NADH → lactate + NAD⁺

Catalyzed by lactate dehydrogenase.

Occurs in mammalian tissues specifically skeletal muscle.

Allows glycolysis to continue under anerobic conditions.

Lactate enters the blood and is ultimately reconverted to glucose in the liver via gluconeogenesis.

Answer 141

A

Occurs in microorganisms such as yeast.

1. Pyruvate → CO₂ + acetaldehyde

Catalyzed by pyruvate decarboxylase.

2. Acetaldehyde + NADH → ethanol + NAD^{<strong>+</strong>}

Catalyzed by alcohol dehydrogenase.

Answer 142

A

Electrons are transferred from cytosolic NADH to oxaloacetate forming malate and NAD⁺.
Malate enters the mitochondrial inner membrane via malate/α-ketoglutarate transporter.
Inside the matrix, malate is reoxidized by malate dehydrogenase and NAD⁺ to form OAA and NADH.
OAA is converted to aspartate via a transamination reaction.
Aspartate is transported back to the cytosol via a glutamate/aspartate transporter.
In the cytosol the aspartate undergoes transamination to reform OAA.

*Shuttle is readily reversible: important in gluconeogenesis.

Answer 143

A

Couples the cytosolic oxidation of NADH with the mitochondrial reduction of FAD.

Cytoplasmic NADH utilized by glycerol-3-phosphate dehydrogenase to convert dihydroxyacetone phosphate to glycerol-3-phosphate.
Glycerol-3-phosphate is then converted back to DHAP by mitochondrial version of the dehydrogenase which resides on the inner mitochondrial membrane. FAD is reduced to FADH₂.
Electrons from FADH₂ are trasferred to the electron carrier Q which enters the respiratory chain as QH₂.

Answer 144

A

Alternate pathway from glucose-6-phosphate
Primary function is generation of NADPH
- Important in neutralization of ROS
- Used to support fatty acid biosynthesis (cytosol)
Serves as a source for ribose-5-phosphate synthesis
- Used in synthesis of nucleic acids
Used to produce glyceraldehyde-3-phosphate
- Fed into TCA and ETC cycles
Pathway can be divided into two sections:
- Oxidative
- Non-oxidative

Answer 145

A

Rate-limiting and regulated step in PPP

Stimulated by G6P and NADP⁺

NADPH is a competitive inhibitor

Answer 146

A

Most common of all enzyme deficiency-related diseases
X-linked
Cuts off cell’s supply of NADPH
Affects RBC’s most who cannot produce new proteins
Oxidative stress leads to hemolysis and anemia
- Infection most common
- Drugs, chemicals, certain foods
  - Fava beans
  - Anti-malarials
  - Certain antibiotics
  - Naphthalene
Heterozygotes have some resistance against malaria
- Therefore gene prevalent in the Mediterranean and Africa

Answer 147

A

Glutathione and the glutathione peroxidase system is the principal antioxidant defense system in mammalian cells
Glutathione is a tripeptide which contains a central Cys residue.
- Reduced form (GSH)
- Oxidized form (GSSG)

Answer 148

A

Glucose preferred but cells will also use other sugars
Fructose and galactoase are significant in the diet
Mannose important components of glycoproteins
Metabolism of other sugars are fed into glycolytic pathways
Sugars must be phosphorylated by the cell before they can be used.
- Hexokinase/glucokinase can phosphorylate other monnosaccharides but their K_M are significantly higher than glucose’s
- Galactokinase present in most cells
- Fructokinase in liver only

Answer 149

A

Fructokinase (found primarily in the liver) phosphorylates fructose at the 1 position producting fructose-1-phosphate
- No mechanism exists to convert F-1-P to G-1-P
Aldolase B (liver isozyme of aldolase) can split both F-1-P and F-1,6-P₂ to form dihydroxyacetone phosphate (DHAP) and glyceraldehyde.
Glyceraldehyde formed can be:
- Phosphorylated by triose kinase to form glyceraldehyde-3-phosphate → enter glycolysis
- Reduced to glycerol by alcohol dehydrogenase then phosphorylated by glycerol kinase to form glycerol-3-phosphate which is then converted to DHAP by glycerol phosphate dehydrogenase.
- DHAP converted to GAP by isomerase.

Answer 150

A

Due to deficiency of fructokinase
Relatively benign
Leads to accumulation of fructose in the urine

Answer 151

A

Autosomal recessive
Absence of aldolase B (liver isoform)
- Results in accumulation of fructose-1-P in the liver
- Depletes levels of ATP and P_i
  - Low P_i levels inhibit glycogenolysis
  - Low ATP levels inhibit gluconeogenesis
  - Low P_i activates AMP deaminase in muscle
    - Results in increased purine catabolism and hyperuricemis → gout
  - Low P_i prevents phosphorylation of ADP so adenylate kinase will convert 2 ADP → ATP + AMP
    - AMP degraded to urate
Symptoms include:
- Vomiting
- Hypoglycemia
- Jaundice
- Metabolic acidosis
- Coma

Answer 152

A

Most tissues are cabable of metabolizing galactose.

Galactokinase phosphorylates galactose at the 1 position producing galactose-1-P.
Galactose-1-phosphate uridylyltransferase (GALT) switches galactose for glucose from UDP-glucose producing UDP-galactose.
UDP-galactose can interconverted to UDP-glucose by UDP-glucose-4-epimerase.

Answer 153

A

Most common cause of galactosemia
Symptoms include:
- Failure to thrive
- Liver damage
- Bleeding
- Sepsis
- Cataracts - later on
If galactose restricted diet provided within the first 10 days the most severe complications can be avoided:
- Neonatal death
- Liver failure
- Intellectual disability
Children with galactosemia remain at risk for developmental delays and problems with speech and mother function.

Answer 154

A

Rare disorder
Morbidity limited to cataract formation

Answer 155

A

Derived from ATP and pantothenic acid (Vit B5)
Contains a reactive thiol group (-SH) that is covalently linked to the acetyl group via a thioester bond
Acetyl~CoA readily donates acetyl groups to other acceptors

Answer 156

A

Member of the α-ketoacid dehydrogenase family
Large multienzyme complex which contains 3 enzymes in multiple copies
Very efficient due to close spatial proximity of complexes

Enzyme subunits

E1: pyruvate dehydrogenase (aka pyruvate decarboxylase) #30
E2: dihydrolipoyl transacetylase #60
- Acts as a swinging arm between E1 & CoA preventing substrate from diffusing away ⇒ substrate channeling
E3: dihydrolipoyl dehydrogenase #12
Contains two stoichiometric coenzymes/cosubstrates
- NAD
- CoA
Contains three catalytic coenzyme prosthetic groups
- thiamine pyrophosphate (TPP)
- FAD
- lipoic acid
Contains two regulatory enzymes associated with but not part of the complex
- PDH kinase
- PDH phosphatase

Answer 157

A

Pyruvate is decarboxylated and the acetyl group is attatched to thiamine pyrophosphate (TPP) coenzyme of pyruvate decarboxylase (E1).
Acetyl group transferred to the lipoic acid covalently bound to dihydrolipoyl transacetylase (E2),
The acetyl group, bound as a thioester to the side chain of lipoic acid, is transferred to free CoA.
The sulfhydryl form of lipoic acid is oxidized by FAD-dependent dihydrolipoyl dehydrogenase (E3) to regenerate the oxidized lipoic acid.
FADH₂ on E3 is reoxidized to FAD as NAD⁺ is reduced to NADH₂ + H⁺.

Answer 158

A

Allosteric regulation

Inhibitors (feedback inhibition)
- Acetyl~CoA
- NADH

Covalent regulation

(main mechanism)

PDH inactivated by phosphorylation by PDH kinase
PDH activated by dephosphorylation by PDH phosphatase

PDH kinase and PDH phoshatase are in turn allosterically regulated:

PDH kinase
- activated (high-energy signals)
  - ATP
  - acetyl-CoA
  - NADH
- inhibited
  - pyruvate
PDH phosphatase
- activated
  - ↑ [Ca²⁺]
    - important in skeletal muscle

Answer 159

A

Niacin (B₃) → NAD+/NADH

Thiamine (B1) → TPP

Lack of either will reduce/block activity of PDH complex.

↓ ATP synthesis

CNS dependent on ATP ⇒ reduced CNS function

Answer 160

A

Arsenic in the form of trivalent arsenite (AsO33-) forms a stable complex with thiol groups of lipoic acid.

Lipoic acid unable to serve as coenzyme for PDH.

Answer 161

A

Acetyl~CoA + 3 NAD⁺ + FAD → 2 CO₂ + 3 NADH + FADH₂

8 steps
- 4 oxidations that produce NADH or FADH₂
- one substrate-level phosphorylation producing GTP
Except for succinate dehydrogenase which is embedded in the inner mitochondrial membrane all enzymes are located in the matrix

Answer 162

A

Formation of citrate.
- Condensation of acetyl~CoA and oxaloacetate (OAA) catalyzed by citrate synthase.
- Reaction made irreversible by the hydrolysis of the thioester bond of acetyl~CoA.
- Facilitated by an enzyme-bound intermediate, citroyl~CoA
- Citrate synthase inhibited by citrate via competitive inhibition
Isomerization of citrate to isocitrate.
- Citrate isomerized to isocitrate by aconitase.
- ΔGº’ = 6.3 kJ/mol but reaction pushed to the right in vivo because product rapidly consumed in next step.
Oxidation of isocitrate to α-ketoglutarate and CO₂.
- Irreversible decarboxylation of isocitrate to α-ketoglutarate by isocitrate dehydrogenase.
  - Inhibited by ATP and NADH.
  - Stimulated by ADP and Ca²⁺
- Loss of CO₂ make the reaction irreversible
- NADH produced
Oxidation of α-ketoglutarate to succinyl~CoA and CO₂.
- Catalyzed by α-ketoglutarate dehydrogenase complex.
  - Same family as PDH dehydrogenase.
  - Cosubstrates (NAD and CoA)
  - Coenzymes (TPP, FAD, and lipoic acid)
  - Inhibited by succinyl~CoA and NADH
  - Activated by Ca²⁺
  - Not regulated by phosphorylation/dephosphorylation.
- NADH produced
- Loss of CO₂makes reaction irreversible.
Conversion of succinyl~CoA to succinate.
- Succinate thiokinase catalyzes the hydrolysis of the thioester bond in succinyl~CoA paired to the conversion of GDP to GTP.
- GTP interconvertible to ATP by nucleoside diphosphate kinase.
Oxidation of succinate to fumarate.
- Catalyzed by succinate dehydrogenase
- FADH₂ made
- Enzyme located on inner mitochondrial membrane.
- ΔGº’ = 0 so reaction can go either way but pushed to the right in vivo because fumarate used in next step.
Regeneration of oxaloacetate.
1. Fumarate converted to malate by fumarase.
2. Malate converted to oxaloacetate by malate dehydrogenase
  - NADH made
    - Rxn has a ΔGº’ = 29.7 kJ/mol but pulled to the right because OAA used by citrate synthase in step 1

Answer 163

A

Regulated almost exclusively at the three irreversible steps.

Citrate synthase competitively inhibited by citrate.
Isocitrate dehydrogenase
- Inhibited by ATP and NADH
- Stimuated by ADP and Ca⁺⁺
α-ketoglutarate dehydrogenase
1. Inhibited by succinyl~CoA and NADH
2. Activated by Ca⁺⁺

Answer 164

A

TCA cycle intermediates can be used for other reactions such as:

amino acid synthesis
fatty acid synthesis
gluconeogenesis

Answer 165

A

Intermediates of the TCA cycle can be provided for by other metabolic pathways, specifically amino acid metabolism.

Answer 166

A

Inner mitochondrial membrane bound complex.

Consists of four seperate protein complexes.

Each complex accepts or donates electrons to/from mobile electron carriers (coenzyme Q and cytochrome C).

ETC pumps protons from matrix to intermembrane space to form a proton gradient.

Answer 167

A

Complex 1: NADH dehydrogenase
- Flavin mononucleotide (FMN) coenzyme accepts two electrons from NADH to become FMNH₂.
- FMNH₂ transfers electrons to CoQ to form CoQH₂.
- Enzyme contains iron-sulfur (Fe-S) centers which acts as intermediate electron carriers.
Complex 2: Succinate dehydrogenase
- Electrons from succinate of TCA cycle transferred via Fe-S centers to FAD to form FADH₂.
Complex 3: Cytochrome bc₁ reductase
- Electrons from CoQH₂ (from complex 1) are used to reduce cytochrome c which acts as electron carrier.
- Contains Fe-S centers which act as intermediates.
- Contains heme group which shifts from Fe³⁺ to Fe²⁺ and back as electrons move through.
Complex 4: Cytochrome oxidase
- Contains two different heme groups
  - Heme a
  - Heme a₃
- Contains two Cu ions which act as intermediates
- Electrons from cytochrome c transferred to heme a via one of the Cu centers, then from heme a to heme a_<em>3</em>via other Cu center, and finally to O₂ to form H₂O.

Answer 168

A

Oxidized form ubiquinone reduced to ubiquinol after accepting electron.

Accepts electrons from complex I and II of the ETC.

Answer 169

A

Contain a heme group in which the iron shifts from Fe³⁺ to Fe²⁺ oxidations states and back as electrons move to and from the heme group.

All contain the porphyrin ring with differing side chains.

Answer 170

A

Measures the ease with which an electron can be added or removed.

The more positive the value of E’º, the greater the tendancy of the oxidant in the redox pair to accept electrons.

Electrons flow from the redox pair with the more negative E’º to one with a less negative/more positive E’º.

Answer 171

A

The redox potential (E’º) is related to the free energy change in the reaction by the Farady constant (F).

ΔG’º = -nFΔEº

n = number of electrons transferred

ΔEº = difference in reduction potentials of the overall reaction

Answer 172

A

In the cell, the effective reduction potential (E) depends on the concentration of reactants.

Answer 173

A

Due to the reduction potentials of each complex the electrons always flow downhill.

Answer 174

A

Used to block electron flow in ETC.

Carriers befor the block become reduced and those after remain oxidized.

Answer 175

A

ATP synthase uses the proton gradient set up by ETC to produce ATP.

Dependent on the integrity of the inner mitochondrial membrane.

No high-energy intermediate exists (as with substrate-level phosphorylation).

Answer 176

A

When more electrons enter the ETC than can be immediately passed to O₂ a state of oxidative stress exists.

Highly reactive superoxide free radicals such as superoxide (•O₂^-) are generated.

Mitochondria have systems to eliminate free radicals:

Superoxide (•O₂^-) is converted to peroxide (H₂O₂) by superoxide dismutase.
Peroxide converted to water by glutathione peroxidase.

NADPH used by glutathione reductase to regenerate glutathione peroxidase.

Answer 177

A

The coupling of ATP synthesis and electron transport.

Neither process can occur without the other.

Answer 178

A

The stoichiometry of ATP synthesis relative to the substrate that is oxidized.

The amount of ATP formed per ½ O₂ consumed (or per pair of electrons.

Approaches 3 for NADH.

Closer to 2 for FADH₂ (because succinate oxidation bypasses complex I)

Answer 179

A

For each pair of electrons that travel through the ETC and transferred to O₂:

Complex 1 pumps out 4 protons

Complex 3 pumps out 4 protons

Complex 4 pumps out 2 protons

Total of 10 protons per electron pair.

NADH + 11 H⁺_M + ½ O₂ → NAD⁺ + 10 H⁺_IM + H₂O

Answer 180

A

Proton pumping by ETC across the inner mitochondrial membrane sets up a pH gradient (ΔpH) and a transmembrane potential (Δψ) with matrix side negative.

Can be calculated as

ΔG = 2.3 RT ΔpH + F Δψ

In general, transport of a pair of electrons through the ETC generates ~ 53 kcal.

Answer 181

A

The proton motive force drives ATP synthesis as the protons flow passively back through the inner mitochondrial matrix down their concentration gradient through a proton pore in the ATP synthase.

The ~ 53 kcal produced per pair of electrons can drive the synthesis of 3 ATPs (using ~ 22 kcal) with the remaining energy used to drive ancillary reactions or dissipated as heat.

ADP + P_i + nH⁺_IM → ATP + H₂O + nH⁺_M

Value of n depends on the structure of the ATP synthase and varies between species (~3.3 - 5 protons per ATP)

Answer 182

A

F₁F_o ATPase

O stands for oligomycin which blocks the proton pore

F-type ATPase which functions as a reversible ATP-driven proton pump

F₁ portion contains the ATPase domain.
- Contains 9 subunits α₃β₃γδε
- α and β subunits for the knoblike catalytic section
- γ subunit forms a shaft that connects with F_o
F_o complex consists of a, b, and c subunits
- 8 c-subunits form the c-ring in vertebrates
  - Rotation of C-ring induces conformational changes in the β subunits of F₁ that drive ATP synthesis
  - Requires 8 protons moving across the membrane with each rotation and produces 3 ATP’s per turn

Answer 183

A

Takes 11 H⁺ to generate 3 ATP’s
- 8 H⁺ to give a complete c-ring rotation
- Additional 3 H⁺ to bring 3 phosphates into the matrix via the mitochondrial phosphate transporter
From 1 NADH: 10 H⁺ pumped so (10/11) x 3 ATPs produced = 2.75 ATPs per NADH
From 1 FADH₂(Succinate): 6 H⁺ pumped so (6/11) x 3 ATPs produced = 1.64 ATPs per FADH₂
Glycolysis generates 2 net ATPs and 2 NADH
PDH generates 1 NADH/pyruvate → 2 NADH/glucose
TCA generates:
- 3 NADH/pyruvate → 6 NADH/glucose
- 1 FADH₂/pyruvate → 2 FADH₂/glucose
- 1 ATP/pyruvate → 2 ATP/glucose
OxPhos:
- 10 NADH → 27.5 ATP
- 2 FADH₂ → 3.3 ATP

Going all the way through oxidative phosphorylation generates:

~ 31 ATP by OxPhos
2 ATP for glycolysis
2 ATP for TCA

TOTAL OF ~35 ATP/GLUCOSE

Answer 184

A

CN^- or CO

blocks electron transport and subsequently ATP synthesis

Answer 185

A

Phosphorylation inhibitor

Blocks ATP synthesis and thus electron transport

Answer 186

A

dinitrophenol (DNP)

uncoupling protein 1 (UCP1 aka thermogenin)

Causes collapse of the proton motive force releasing energy in the form of heat rather than ATP.
Electrons free to move through the ETC.
Increases oxygen consumption.

Answer 187

A

ETC and ATP synthesis coupled so increasing or decreasing one does the same to the other.

Answer 188

A

All the transport processes associated with oxidative phosphorylation which utilize the proton motive force or transmembrane potential.

Mostly used to transport molecules across the impermeable inner mitochondrial membrane.

Agents that disrupt the PMF will reduce transport.
Transport of these substances will conversely reduce the PMF available for ATP synthesis.
Electroneutral transport systems driven by concentration gradients alone.
Electrogenic transport systems utilize concentration gradients as well as the transmembrane potential.
- positively charged molecules enter matrix more easily
Adenine nucleotide translocase
Phosphate translocase
Transporters for other charged metabolites such as pyruvate, malate, citrate, etc.
Ca⁺⁺ and asparate also transported in a PMF-dependent fashion

Answer 189

A

Antiporter that exchanges ADP^3- from the intermembrane space for ATP^4- from the matrix.

Net transport of 1 negative charge out of the matrix (electrogenic) stimulated by the matrix-negative transmembrane potential.

Inhibited by atractyloside.

Answer 190

A

Symporter that moves one phosphate (H₂PO₄^-) and one proton (H⁺) into the matrix.

Driven by the proton gradient established by ETC.

Electronically neutral.

Answer 191

A

Complex containing ATP synthase, adenine nucleotide translocase, and phosphate translocase can be isolated by gentle disruption of the inner mitochondrial membrane suggesting that these three proteins are spatially integrated.

Answer 192

A

A transmembrane protein embedded in the inner mitochondrial membrane.

Uses the proton gradient to drive:

NADH + NADP⁺ + H⁺_IM → NAD⁺ + NADPH + H⁺_M

NADH is used by ETC so its levels are related to the potential for ROS generation.

Production of NADPH by transhydrogenase will increase as more electrons travel down ETC.

NADPH indirectly used in ROS neutralization.

Answer 193

A

Mitochondria contain their own genome which contains 37 genes.

13 encode subunits of respiratory chain proteins.

Answer 194

A

Maternally inherited.

Defects in oxidative phosphorylation most commonly associated with mutations in mitochondrial genes.

Presumably due to generation of reactive oxygen species.

Affects tissues with high requirement for ATP such as brain, liver, and skeletal/cardiac muscle.

Answer 195

A

Triggered by:
- External signals acting via a receptor
- Oxidative stress
- Heat shock
- Viral infection
Exposure to stimulus induces formation of large pores in the outer mitochondrial membrane called permeability transition complex
- Allows release of cytochrome c into the cytosol
Cytochrome c in associated with apoptosis protease activating factor 1 (Apaf-1) activate a family of cytosolic proteases (the caspases) that degrade proteins and lead to cell death.

Answer 196

A

Major short-term storage form of carbohydrates in animals.
- For times of metabolic need.
Branched chain homopolysaccharide of α-D-glucose
- Primary bond is α-1,4-glycosidic linkages.
- Every 8-10 glucose residues there is a branch attached via a α-1,6-glycosidic linkage
Stored in the cytoplasm as large hydrated granules.
- Each granule contains as many as 55,000 glucose units.
Found in the cytoplasm of liver and skeletal muscle cells primarily.

Answer 197

A

Diet
Degradation of glycogen
Gluconeogenesis

Answer 198

A

Functions to maintain the blood glucose concentration particularly during the early stage of a fast.
Glucose rapidly released from liver glycogen.
- Glucose able to enter systemic circulation due to presence of glucose-6-phosphatase in the liver.
Hepatic glycogen stores ~24 hour supply of glucose.
Liver also able to synthesize glucose via gluconeogenesis.

Answer 199

A

Serves as a fuel reserve for synthesis of ATP that will power muscle contraction.
Muscle glycogen is not available to other tissues because muscle lacks glucose-6-phosphatase.

Answer 200

A

Phosphorylation
- Glycolysis utilizes glucose-6-phosphate.
Create a nucleotide sugar
- Glycogenesis utilizes UDP-glucose.

Alternate activation methods allows for both pathways to occur at the same time.

Answer 201

A

Occurs in the cytoplasm and can be divided into two stages:
1. Chain synthesis
2. Chain branching

I. Chain Synthesis

A. Synthesis of UDP-glucose.

Glucose is phosphorylated to glucose-6-phosphate.
- By glucokinase in hepatic tissue.
- By hexokinase in peripheral tissue.
Glucose-6-phosphate converted to glucose-1-phosphate by phosphoglucomutase.
Glucose-1-phosphate reacts with UTP to form UDP-glucose and PP_i which is catalyzed by glucose 1-phosphate uridylyltransferase (aka UDP-Glc pyrophosphorylase)
- Hydrolysis of PP_i → 2 P_i by pyrophosphatase makes the reaction energetically favorable and irreversible.

B. Requirement of primer to initiate glycogen synthesis.

⇒Glycogen synthase cannot initiate glycogen synthesis de novo and can only add glucose to an existing chain, therefore, glycogenesis requires a primer.

Glycogen fragment can serve as a primer for glycogen synthase which attaches glucosyl residues to existing chain using UDP-glucose.
The protein glycogenin (homodimer) can prime glycogen synthesis and attach glucose residues through auto-glucosylation ⇒ serves as both substrate and enzyme in its role as primer.
a. The glycosyltransferase activity of glycogenin transfers the first molecules of glucose from UDP-glucose to a specific tyrosine side-chain (tyr-194) on itself.
b. After at least 4 (about 7) glucose residues have been added, glycogen synthase takes over.
- Glycogenin remains within the glycogen granule.

C. Elongation of glycogen chains.

Glycogen synthase transfers glucose from UDP-glucose to the non-reducing end of the growing chain via α-1,4-linkages between the -OH group on C-1 of the activated sugar and the C-4 of the accepting sugar.

⇒ Glycogen synthase is the rate-limited and regulated enzyme of glycogenesis.

⇒ There are liver & muscle isozymes.

⇒ UDP released when α-1,4-glycosidic bond is formed can be converted to UTP by nucleoside diphosphate kinase + ATP.

Answer 202

A

Occurs in the cytoplasm and can be divided into two stages:
1. Chain synthesis
2. Chain branching

II. Chain branching

A. Catalyzed by “branching enzyme” called glucosyl 4:6 transferase.

Glucosyl 4:6 transferase cleaves an α-1,4-glycosidic bond from the non-reducing end of the glycogen chain producing a 6-8 glucosyl residues fragment.
Enzyme then transfers the fragment to another residue on the linear chain via an α-1,6-glycosidic bond.
Resulting new non-reducing end and old non-reducing ends can be further elongated by glycogen synthase.
After further elongation, new chains of 6-8 residues can be transferred to make additional branches.

Answer 203

A

Increases solubility of glycogen molecule.
- Stored as hydrated granules.
Increases number of non-reducing ends to which new glucosyl residues can be added to or removed from glycogen.
- Facilitates fast breakdown of glycogen into glucose when energy needed.

Answer 204

A

Occurs primarily in the cytoplasm of liver and skeletal muscle cells.
Involves 2 stages:
1. Shortening of chains
2. Removal of branches

Answer 205

A

Glycogen phosphorylase uses Pi to cleave the α-1,4-glycosidic bonds between glucose residues at the non-reducing ends of the glycogen chains releasing glucose-1-phosphate.
⇒Enzyme is a homodimeric exoglucosidase
⇒Requires pyridoxal phosphate (PLP) coenzyme (derivative of Vit B6)
⇒Liver and muscle isozymes
- Glycogen phosphorylase stops attacking α-1,4-glycosidic bonds four glucosyl residues from an α-1,6-branch point.
- Resulting structure is called a limit dextrin.
Glucose-1-phosphate is converted to glucose-6-phosphate by phosphoglucomutase.
Next step in glycogenolysis depends on the tissue:
- In liver: glucose-6-phosphate is hydrolyzed to free glucose and P_i by glucose-6-phosphatase.
  ⇒Free glucose able to leave the liver and enter the blood stream.
- In peripheral tissues: glucose-6-phosphate will be oxidized in the glycolytic pathway to produce energy.

Answer 206

A

Catalyzed by a debranching enzyme which is a bifunctional protein with two catalytic activities.

4-α-D-glucantransferase activity transfers the outer three glucosyl residues of the limit dextrin to a non-reducing end by breaking and formation of α-1,4 bonds leaving one glucosyl residue in α-1,6 linkage.
⇒ 4,4 transferase activity
α-1,6 linkage is then cleaved hydroytically by the amylo-α-1,6 glucosidase activity of debranching enzyme releasing free glucose (non-phosphorylated).
Co-operative and repetitive action of phosphorylase and debranching enzymes results in complete hydrolysis of glycogen to yield glucose-1-phosphate and free glucose in a 12:1 ratio.

⇒Free glucose is quickly phosphorylated in muscle for intracellular use.
⇒Phosphorylated glucose is already trapped inside muscle cells.

Answer 207

A

Key regulatory enzymes of glycogen metabolism:

Glycogen synthase (synthesis)

&

Glycogen phosphorylase (degradation)

Each is controlled by:

Hormone-induced covalent modification through phosphorylation or dephosphorylation of ser residues.
* Way of responding to the needs of the body as a whole.
Allosteric effectors
* Way of responding to the need of a particular tissue at a particular time.

Answer 208

A

I. Regulation by covalent modification:

Glycogen phosphorylase exists in two forms:

A or active form: phosphorylated

B or inactive form: dephosphorylated

A and B forms are interconverted by:

Phosphorylase kinase
Produces active phosphorylated A-form.

&

Phosphoprotein phosphatase-1

Produces inactive dephosphorylated B-form.

*The A form of glycogen phosphorylase is more active because phosphorylation causes a conformational change in the enzyme which shifts the equilibrium of the enzyme between its T-state (taut and inactive) and R-state (relaxed and active) towards the active R-state. The B-form is inactive because the taut state is favored.

Phosphorylase kinase (regulatory enzyme) is itself regulated by phosphorylation/dephosphorylation.

A and B forms are interconverted by:

Protein Kinase A (PKA)
Produces active phosphorylated A-form.

&

Phosphoprotein phosphatase-1

Produces inactive dephosphorylated B-form.

Answer 209

A

Hormonal signals that activate PKA include Glucagon and Epinephrine.

Phosphorylase kinase (regulatory enzyme)

&

Glycogen phosphorylase (regulated enzyme of glycogenolysis)

are phosphorylated in response to hormonal signals that are transduced via cAMP which activates protein kinase A.

PKA also results in the inhibition of phosphoprotein phosphatase-1 by phosphorylating and activating protein phosphatase inhibitor (A-form).

This enzyme is used to maintain the level of phosphorylation until hormone signal changes.

Active protein kinase A has a short half-life because seperation of regulatory and catalytic subunits revealed PEST sequences which marks protein for degradation.

Answer 210

A

Peptide hormone released from the alpha-cells of the pancreas when blood glucose is low.

Glucagon binds to plasma membrane receptors on liver cells but not muscle.

Stimulates glycogen degradation via PKA-mediated activation of phosphorylase kinase.

Activated during periods of fasting thus making glucose available to tissues.

Answer 211

A

Glycogen synthase exists in two forms:

A or active form: dephosphorylated

B or inactive form: phosphorylated

Several kinases phosphorylate glycogen synthase A to the inactive B form including:

cAMP dependent PKA*
Phosphorylase kinase*
Glycogen synthase kinase-3*
AMP-dependent kinase*

B-form converted back into the active A-form by

Phosphoprotein phosphatase-1.

PKA also results in the inhibition of phosphoprotein phosphatase-1 by phosphorylating and activating protein phosphatase inhibitor (A-form).

Answer 212

A

cAMP regulates glycogen metabolism through the simultaneous activation of glycogenolysis and inhibition of glycogenesis.

Also displays amplification: 1 hormone activates many adenyl cyclase, makes many cAMP, each activates many PKA, each phosphorylates many phosphorylase kinase, each phosphorylates many phosphorylase ect.

Answer 213

A

Released by pancreas in response to high blood glucose levels.

Has the opposite affect of glucagon/epinephrine.

Activates glycogen synthesis and inhibits degradation in liver and muscle.

Promotes inhibition of several protein kinases and activation of phosphoprotein phosphatase.
- Causes subsequent dephosphorylation of glycogen synthase (activating) and dephosphorylation of phosphorylase kinase and phosphorylase (inactivating).
Promotes conersion of cAMP to 5’ AMP by activating a phosphodiesterase.
- Causes subsequent decrease in active PKA.

Answer 214

A

Allosteric effectors are superimposed onto covalent regulation in order to meet the needs of the tissue.
Enzymes of glycogen metabolism including glycogen phosphorylase kinase, glycogen phosphorylase, and glycogen synthase are regulated in an allosteric effectors acting in a non-covalent manner.
- Postive allosteric effectors bind to a regulatory site on the R (active) form of the enzyme stabilizing it and pulling the equilibrium towards the R-form.
- Negative allosteric effectors bind to the T (inactive) form and stabilizes that pulling equilibrium towards T-form.
Effectors include:
- Ca²⁺ and AMP which are signs of low energy
- Glucose and glucose-6-phosphate which are signs of high energy

Answer 215

A

Released in times of energy need.

Ca²⁺ binds to the calmodulin-like δ-subunits of dephosphorylated (B-form) phosphorylase kinase causing conformational change which activates the catalytic γ-subunits in the absence of phosphorylation ⇒ stabilizes the R-state

Ca²⁺ also required for maximal activation of phosphorylase kinase a.

Phosphorylase kinase subsequently phosphorylates and inhibits glycogen synthase.

End result of a rise in [Ca²⁺]_in is increased degradation and decreased synthesis of glycogen.

Contracting muscle
- Ca²⁺ released in in response to nerve impulses.
- [Ca²⁺]_in activates sarcoplasmic glycogenolysis by activating phosphorylase kinase.
Liver cells
- Epinephrine binds to α-adrenergic receptors, activating phospholipase-C, and generating IP₃ and DAG from PIP₂.
  - IP₃ causes release of Ca²⁺ from the SER.
    - [Ca²⁺]_in activates glycogenolysis by activating phosphorylase kinase⇒ activates glycogen degradation.
  - DAG activates PKC which phosphorylates and inactivates glycogen synthase ⇒ inhibites glycogen synthesis.

Side note: Ca²⁺ also activates mitochondrial events:

Activates PDH phosphatase thereby activating PDH and pyruvate degradation.
Allosteric effector of isocitrate dehydrogenase and α-ketoglutarate dehydrogenase thereby activating TCA cycle.

Answer 216

A

AMP to ATP ratio reflects the energy state in muscle cells.
- Increase in AMP signals low energy and need for glycogen degradation.
AMP functions as an allosteric activator of muscle phosphorylase b.
- Directly activates the dephosphorylated form of myophosphorylase b.

Answer 217

A

When glucose is plentiful, heptatic glycogenolysis is decreased by glucose itself acting as an allosteric inhibitor of hepatic phosphorylase a.
The glucose-6-phosphate fromed from glucose is an allosteric activator of glycogen synthase b in both liver and muscle, thus increasing glycogenesis.
- For this reason, glycogen synthase b is sometimes designated “D” because it is dependent on glucose-6-phosphate for activity while synthase a is designed “I” for independent.

Answer 218

A

Von Gierke Disease

Glucose-6-phosphatase deficiency
- Liver and kidney
Severe fasting hypoglycemia hallmark
Major Findings
- Hepato/renomegaly
- Fasting hypoglycemia
- Lactic acidemia
- Hyperuricemia
- Hyperlipidemia

Answer 219

A

Pompe Disease

aka

Generalized Glycogenosis

Lysosomal acid α-glucosidase deficiency
Infantile, juvenile, and adult-onset forms
Affects all organs but skeletal/cardiac most
Cardiomegaly/myopathy in infantile forms
Muscle weakness in later forms
Enzyme replacement therapy has reduced mortality

Answer 220

A

Cori Disease

aka

Limit Dextrinosis

Debranching enzyme deficiency (both actions)
IIIa : affects liver and muscle
IIIb: affects liver only
Milder hepatomegaly
Muscle weakness
Accumulated glycogen has abnormal structure with shorter chains
May cause liver fibrosis or cirrhosis

Answer 221

A

Andersen Disease

aka

Amylopectinosis

Branching enzyme deficiency in liver
Progressive hepatomegaly
Accumulated glycogen has abnormal structure with longer chains and no branches
Progressive liver cirrhosis in infantile form can be lethal

Answer 222

A

McArdle Disease

Muscle glycogen phosphorylase deficiency
Infantile and Adult forms
Exercise intolerance and muscle cramps
Symptoms usually first appear in adolescence
Failure of blood lactate to rise after anaerobic exercise

Answer 223

A

Hers disease

Liver glycogen phosphorylase deficiency
Hepatomegaly
Benign in general
Mild fasting hypoglycemia

Answer 224

A

Tarui Disease

Muscle PFK-1 deficiency
Affects muscle and RBC’s
More severe exercise intolerance and muscle cramps
RBC’s show some percentage of normal activity

Answer 225

A

The synthesis of glucose from smaller precursors such as:
- pyruvate
- lactate
- glycerol
- most amino acids
  - Alanine ⇒ pyruvate
  - Glutamine ⇒ α-ketoglutarate
  - Except: leucine and lysine
Serves to stabilize glucose levels in the blood during fasting after glycogen stores have been depleted
Occurs in the liver and to some extent the kidneys
Requires specialized gluconeogenic enzymes to bypass the irreversible steps of glycolysis

Answer 226

A

Pyruvate + HCO₃^- → oxaloacetate
by pyruvate carboxylase + ATP

Reaction is anaplerotic because it yields a citric acid cycle intermediate OAA.
2. Oxaloacetate → PEP + CO₂
by phosphoenolpyruvate carboxykinase (PEPCK) + GTP

Required to bypass the highly exergonic reaction catalyzed by pyruvate kinase during glycolysis.

Answer 227

A

Biotin-containing multifunctional protein
Biotin coenzyme linked to a lysine residue

During course of reaction:

Carbon dioxide is first bound to the biotin in one active site forming a high energy carboxy-biotin species.
Long flexible side chain swings the carboxy-biotin to the second active site.
Carboxy-group transferred to pyruvate.

Answer 228

A

Allosterically activated by acetyl CoA.

Acetyl CoA is a product of fatty acid metabolism.

Links gluconeogenesis and fat catabolism.

Acetyl CoA also inhibits pyruvate dehydrogenase by activating PDH kinase.

Decreased glycolysis, increased gluconeogensis, and increased fatty acid catabolism by the liver when blood glucose low.

Compartmentalized in the mitochrondria.

Answer 229

A

Fructose-1,6-bisphosphate → fructose-6-phosphate

by fructose-1,6-bisphosphatase

By passes the reaction of PFK1 in glycolysis.

fructose-1,6-bisphosphatase

Allosterically inhibited by F-2,6-P₂ and AMP

Same allosteric activators of PFK1 = Reciprocal regulation

Answer 230

A

Glucose-6-phosphate → glucose

by glucose-6-phosphatase

By passes reaction of hexokinase/glucokinase.

Allows glucose to leave the cell and enter the blood.

Answer 231

A

Glucose-6-Phosphatase is a membrane bound enzyme located in the ER.

Active site faces into the lumen of ER.

Glucose-6-P must enter then glucose and P_i must leave.

Deficiency causes glycogen storage disease Ia.

Found only in liver and kidney.

There is no direct control of the enzyme but has a high KM for glucose-6-P so only functions when concentrations high.

Answer 232

A

Glycolysis converts 1 glucose into 2 molecules of pyruvate.

Produces 2 NADH and 2 ATP

Gluconeogenesis converts 2 pyruvates into 1 glucose

Consumes 2 NADH and 6 ATP or GTP

Two processes are tightly regulated so that only once may proceed at a time.

Allosteric Regulation

Important sites for control are the irreversible reactions

pyruvate kinase - F-1,6-P₂ (+) , ATP (-)
pyruvate carboxylase - Acetyl CoA (+), ADP (-)
phosphofructokinase - AMP (+) , F-2,6-P₂ (-)
fructose bisphosphatase - AMP (-) , F-2,6-P₂ (-)

Hormonal Regulation

Glucagon → increased cAMP → phosphorylation of hepatic pyruvate kinase and PFK 2 → decrease glycolysis
Insulin → decreased cAMP → increase hepatic glycolysis

Transcriptional Regulation

Glucagon → stimulates expression of PEPCK and maybe glucose-6-phosphatase through phosphorylation of cAMP response element (CREB) by PKA
Insulin → stimulates expression of PFK1, pyruvate kinase, PFK2, enzymes of glycolysis

Answer 233

A

Gluconeogenesis requires both mitochrondrial and cytosolic enzymes.

Pyruvate carboxylase is a mitochondrial enzyme while the other gluconeogenic enzymes are largely cytoplasmic.

Pyruvate must be transported into the mitochondria where it is converted to OAA.

OAA_mito ⇒ Malate_mito ⇒ Malate_cyto ⇒ OAA_cyto

Gluconeogenesis consumes NADH in the cytosol in the conversion of 1,3-bisphosphoglycerate to glyceraldehyde-3-P but the NADH/NAD⁺ ratio is normally very low so glycolysis usually preferred.

Answer 234

A

Occurs in the liver
Lactate converted to pyruvate in the cytosol by LDH
- Yields NADH ⇒ export of reducing equivalents fro mthe mitochondria is not nescessary
Pyruvate enters mitochondria where it is converted to OAA by pyruvate carboxylase
OAA can either:
- Be converted to aspartate by transamination in mitochondria ⇒ leave mitochondria via aspartate shuttle ⇒ converted back to OAA in the cytosol
- Be converted to PEP by mitochrondrial PEP carboxykinase ⇒ PEP leaves mitochondria

Answer 235

A

Recycles glucose carbons from lactate in order to maintain blood glucose levels.

Lacate produced via anaerobic glycolysis travels to the liver.

There it is used for gluconeogenesis and resulting glucose is released back in to the blood.

(Liver hopes the next time the glucose will be used oxidatively for more energy.)

Answer 236

A

Hypoxanthine

Answer 237

A

Orotic Acid

Answer 238

A

Built on a ribose-5-phosphate.

Occurs primarily in cytosol of hepatocytes.

Answer 239

A

11 step process

Requires a large amount of ATP

Ribose-5-P ⇒ PRPP by PRPP synthetase
- Regulated but not committed step
Add N from glutamine to PRPP by PRPP glutamyl amidotransferase
- Committed step
- Highly regulated
- Rate-limiting
Add glycine backbone
Add C from N¹⁰-THF
Add N from glycine for ring 2
Ring 1 closure
Add C from respiratory CO₂
Attach aspartate R group N to C of respiratory CO₂
Keep only N from aspartate and release backbone as fumarate
Add C from N¹⁰-THF
Close ring 2 to form inosine monophosphate (IMP)

Answer 240

A

Catalyzes the regulated but not committed step of purine de novo synthesis.

Feed back inhibition by purine ribonucleotides (mainly AMP and GMP)

Activated by P_i

Answer 241

A

Catalyzed the committed and rate-limiting step of purine de novo synthesis.

Inhibited by end-products (purine ribonucleotides, inactive dimer)

Activated by PRPP and active monomer

High [PRPP] able to overcome end-product inhibition.

Answer 242

A

IMP converted to AMP or GMP in seperate two step reactions.

IMP ⇒ AMP

IMP + Asp ⇒ Adenylosuccinate monophosphate (ASMP)

ASMP ⇒ AMP + Fumarate

IMP ⇒ GMP

IMP ⇒ XMP

XMP + Gln ⇒ GMP + Glutamate

Answer 243

A

Reciprocal regulation ensures appropriate levels of AMP and GMP.

1st step in IMP ⇒ AMP inhibited by AMP and requires GTP.

2nd step in conversion of IMP ⇒ GMP inhibited by GMP and requires ATP.

Answer 244

A

Meets the needs of the cell.

Answer 245

A

Maintains adequate and balanced concentrations of deoxyribonucleotides.

Catalyzed by ribonucleotide reductase.

Requires reduced thioredoxin coenzyme.

RNR has 3 sites:

Active site ⇒ reduces ribonucleotide to deoxyribonucleotide
Activity site ⇒ on/off switch
- ATP activates
- dATP inactivates
Substrate specificity site ⇒ binds a specific positive allosteric effector (NTP or dNTP) to control reductive of a given NDP
- C ⇒ U ⇒ G ⇒ A
- Ex. Binding of dCTP makes enzyme specific for UDP

RNR inhibited pharmacologically by hydroxyurea.

Answer 246

A

Guanine ⇒ Xanthine by Guanase

Adenosine ⇒ Inosine by Adenosine Deaminase

Inosine ⇒ Hypoxanthine by Purine nucleoside phosphorylase

Hypoxanthine ⇒ Xanthine by Xanthine Oxidase

Xanthine ⇒ Uric acid by Xanthine Oxidase

Answer 247

A

Free purine bases or nucleosides can be reutilized.

Important in non-hepatic tissues.

Base salvage:

Adenine phosphoribosyltransferase (APRT)

Adenine + PRPP ⇒ AMP + PP_I

Hypoxanthine-guanine phosphoribosyltransferase (HGPRT)

H or G + PRPP ⇒ IMP or GMP + PP_i

Nucleoside salvage:

Adenosine kinase

Adenosine + P_i ⇒ AMP

Answer 248

A

Salvage inhibits de novo synthesis:

Decreases [PRPP] ⇒ base salvage only

Generates nucleoside monophosphates that inhibit the amidotransferase ⇒ base and nucleoside salvage

Answer 249

A

Adenosine Deaminase (ADA) Deficiency

⊗ ADA ⇒ ⊗ adenosine catabolism ⇒ ↑ [dATP] ⇒ ⊗ RNR ⇒ developmental arrest and/or apoptosis of lymphocytes

Results in severe combined immunodeficiency syndrome ⇒ ADA-SCIDS

Answer 250

A

Purine nucleoside phosphorylase deficiency

Rarer and less severe than ADA deficiency.

Primarily affects T-cells.

Answer 251

A

X-linked
Hypoxanthine-Guanine Phosphoribosyltransferase (HGPRT) deficiency
- ↑ [PRPP] ⇒ ↑ [uric acid]
Symptoms:
- hyeruricemia with gout
- compulsive self-mutilation
- aggressive behavior
- developmental delay
- profound motor impairment
- early death from renal failure

Answer 252

A

Build the base directly then attach to a sugar phosphate.

Answer 253

A

Pyrimidine ring made then linked to ribose phosphate.

CAD catalyzed first 3 reactions.

UMP synthase catalyzes reactions 5 & 6.

Orotic acid is the first pyrimidine formed
- Regulated and rate-limiting step is synthesis of carbamoyl phosphate by CPS II.
- Committed step is synthesis of carbamoyl aspartate by aspartate transcarbamoylase.
Orotic acid + PRPP ⇒ OMP (first nucleotide)
OMP decarboxylated to UMP

Answer 254

A

OMP ⇒ UMP

UMP phosphorylated by kinases to UDP and UTP
UTP aminated using Gln to CTP and Glu by CTP synthetase

Answer 255

A

⊕ UTP

⊖ CTP

Answer 256

A

UDP ⇒ dUDP by RNR

dUMP arises from both dUD(T)P and dCMP

dUDP ⇒ dUTP by kinase

dUTP ⇒ dUDP ⇒ dUMP by dUTPase

dCMP ⇒ dUMP by deamination

Answer 257

A

dUMP methylated at the 5 position by thymidylate synthase ⇒ TMP

Thymidylate synthase inhibited by 5FU.

Uses N⁵N¹⁰-methylene THF as 1 C donor.

THF oxidized to DHF as methylene group reduced to methyl group.

DHF reduced back to THF by dihydrofolate reductase + NADPH.

Dihydroflocate reductase inhibited by methotrexate.

Traps folate as DHF ⇒ ↓ thymidylate synthesis ⇒ ↓ DNA synthesis

Answer 258

A

5-FU converted to nucleotide monophosphate by addition of PRPP.

Deoxy from of nucleotide inhibits thymidylate synthase.

Answer 259

A

Folic acid analog

Inhibits dihydrofolate reductase.

↓ thymidylate synthesis ⇒ ↓ DNA synthesis

Used to treat some cancers and psoriasis.

Answer 260

A

Trifunctional protein.

Catalyzes the first 3 steps of pyrimidine synthesis:

Carbamoyl phosphate synthetase II
Aspartate transcarbamoylase
Dihyrorotase

Answer 261

A

Carbamoyl phosphate synthetase II is the regulated enzyme

⊕ PRPP

⊖ UTP

Answer 262

A

UMP synthase deficient

Orotate phosphoribosyl transferase
Orotate decarboxylase

↑ [orotic acid] and ↑ excretion of OA

Symptoms are failure to grow and develop and anemia.

Treat with uridine:

Uridine → UMP →→ UTP → CTP

Requires functional salvage pathway.

UTP inhibits CPSII of pyrimidine de novo synthesis so decreases production of OA also.

Answer 263

A

All products are water soluble.

There are no known pathologies of salvage or degradation.

Nucleotide → Nucleoside → Free base:

Cytosine deaminated to uracil.

Uracil → β-alamine, CO₂ and NH₄

Thymine → β-aminoisobutyrate, CO₂ and NH⁺

Some salvage of nucleosides and bases to nucleotides.

Answer 264

A

Thymine → β-aminoisobutyrate, CO₂ and NH⁺

β-aminoisobutyrate is unique to thymine degradation and urinary excretion used as a measure of DNA turnover

Answer 265

A

Disorder of purine metabolism.

Requires:

Hyperuricemia

Precipitation of uric acid or monosodium urate crystals in tissues

Inflammatory response

Tophi may be seen.

Answer 266

A

Uric acid deprotinated at physiological pH to urate

Urate + sodium ions ⇒ monosodium urate

Answer 267

A

Hyperuricemia
- Necessary precondition for gout but not enough
- All patients with gout show hyperuricemia but only minority of people with hyperuricemia experience gout
Formation and deposition of urate crystals
- Tendency for crystal deposition varies
- Factors may precipitate crystal formation
  - ↓ temperature
    - Explain why gout most commonly affects great toe (podagra)
  - ∆ pH
  - trauma
  - stress
Inflammatory response to the crystals

Answer 268

A

Asymptomatic hyperuricemia

↓

Acute intermittent attacks

↓

Chronic tophaccous gout

May see urolithiasis.

Answer 269

A

Gout is the major manisfestation of an innate disorder.

>90% of cases hyperuricemia due to ↓ excretion not ↑ production

Most cases idiopathic.

Known molecular defects:

PRPP synthetase with increased activity ⇒ super synthetase
Partial deficiency of HGPRT

Answer 270

A

Gout is a secondary manifestation of an underlying disorder.

Total deficiency (<1.5% of normal) of HGPRT seen in X-linked Lesch-Nyhan syndrome.
Increased cell turnover
- cancers
- hemolytic diseases
- psoriasis
- chemotherapy
Decreased renal excretion
- lead poisoning
Increased conversion of ATP ⇒ AMP
- ETOH abuse
- Type I glycogen storage disease
- Inborn errors of fructose metabolism
- Hypoxia

Answer 271

A

Therapy to reduce inflammation.

No effect on urate concentrations.

NSAIDS
- Indomethacin
- Aspirin contraindicated
  - competes with urate for renal excretion
Colchicine
- Inhibits phagocytosis of urate crystals
  - Inhibits microtubule assembly
- Many side effects
- Low dose used prophylactically
Steroids
- Glucocorticoids

Answer 272

A

Life-long therapy to decrease urate concentration.

No direct effect on inflammation.

Increasing renal excretion ⇒ uricosuric agents
- Probenecid
  - Inhibits renal reabsorption of urate
Decreasing urate synthesis
- Xanthine oxidase inhibitors
  - Allopurinol
  - Febuxostat
Uricase
- urate to more soluble allantoin

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Exam 2: Biochemistry Flashcards

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