Exam 1 Flashcards
Heme pathway mnemonic
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Glycine + Succinyl CoA –> ALA
Enzyme and byproducts?
ALA Synthase, CoA-SH and CO2
ALA –> PBG
enzyme and byproducts?
ALA Dehydrase, 2H2O
PBG –> Bilane
enzyme
PBG Deaminase
Bilane –> Uro’gen III
Enzyme
Uro’gen III cosynthase
Uro’gen III –> Copro’gen III
Enzyme and byproducts?
Uro’gen III decarboxylase, 4CO2
Copro’gen III –> Proto’gen IX
Enzyme and byproducts?
Copro’gen III decarboxylase, 2CO2
Proto’gen IX –> Protoporphyrin IX
enzyme and byproducts?
Proto’gen IX dehydrogenase, 6H+
Protoporphyrin IX –> Heme
enzyme and byproducts?
Ferrocheletase, 2H+, adds Fe2+
From 1-8, which intermediates in the heme synthesis pathway are the most hydrophobic
1 - least hydrophobic
8 - most hydrophobic
4 ways of ALA synthase inhibition by heme
1) Heme binds to ALA synthase
2) Heme binds repressor in nucleus (inhibits transcription of ALA synthase)
3) Heme blocks translation of ALA synthase
4) Heme blocks ALA synthase import into mitochondria
Heme oxygenase pathway enzymes
1) Heme Oxygenase
2) Biliverdin reductase
What are the notable cofactors and byproducts in the heme oxygenase pathway and at what step are they used?
It uses O2 and produces CO in the production of Heme to Biliverdin IXa
3 acute porphyrias
AIP - Acute Intermittent Porphyria
HC - Hereditary Coproporphyria
VP - Variegate Porphyria
Chronic porphyria
Porphyria Cutanea Tarda
3 photosensitive porphyrias
HC, PCT, VP
What intermediates build up as a result of PCT?
Only Uro’gen III
PCT enzyme
Uro’gen III Decarboxylase
HC enzyme
Copro’gen III Decarboxylase
VP enzyme
Proto’gen IX Dehydrogenase
AIP enzyme
PBG Deaminase
Gilbert’s Syndrome
Inhibition of BRGT/UGT promoter, results in increased BR-Albumin
What is a stasis
Defect in liver secretion
Crigler-Najjar (two types)
Mutation in BRGT/UGT coding regions, results in increased BR-Albumin
Type I is homozygous = fatal
Type II is heterozygous
Conjugated Hyperbilirubinemia
Liver stasis, BRDG leaks back into the blood, eliminated in dark orange urine
Neonatal Jaundice
Lack of BRGT/UGT expression in first 5-14 days, since BR is unconjugated, it is not transported into liver and BR-Albumin builds up in the blood. Treated by using light to break down BR-Albumin
Kernicterus
BR-Albumin buildup due to lack of BRGT/UGT expression. Since blood-brain barrier not complete, becomes toxic to nervous system, build-up in basal ganglia
Porphyria Cutanea Tarda illness course
1) Found in Hep C patients where Iron is deposited in the liver, leading to inflammation and production of ROS.
2) ROS then causes Uro’gen III to become inhibitor of Uro’gen III decarboxylase.
3)Uro’gen III becomes Uroporphyrin, creating an immune response (rash, lesions, etc.)
Prolyl and Lysyl Hydroxylase
Hydroxylate Proline and Lysine using Ascorbate (Vitamin C) on alpha chains of pre-pro collagen in ER
Lysosomal storage disease is characterized by:
accumulation of sphingolipids in the lysosome
Galactosyl and Glucosyl transferase
Glycosylate hydroxyLYSINE in ER, first Galactose, and then sometimes Glucose gets added
collagen alpha chains amino acids
33% Glycine, 17% Proline
What cleaves procollagen and where does it happen?
Procollagen peptidase, outside the cell
Ehlers-Danlos syndrome is a result of:
Lack of Lysyl Hydroxylase, resulting in fragile collagen and stretchy skin
Lysyl Oxidase (LOX)
1) Cross-links collagen fibers using Copper, creating aldehydes on Lysines and Hydroxylysines
2) cross-links tropoelastins to create insoluble elastin fiber
Osteogenesis Imperfect is a result of:
mutations in the alpha1 and alpha2 chains used to create the procollagen triple helix
Elastin turnover is catalyzed by:
Elastase
Alpha1 Antitrypsin role
Inhibits Elastase, otherwise Elastin would be destroyed and skin would become stiff
Rac1/PIP2
Activate WASP/WAVE complex in response to extracellular signals to begin reorganization of actin
WASP/WAVE complex
Stimulate actin nucleation via activation of Arp2/3 complex
Arp2/3 complex
stimulate actin nucleation and creation of new filaments
Formin proteins
Cap + ends of growing actin filaments to prevent further extension
Rac1
Regulates extension of the leading edge
Rho GTPase
RhoA
Regulates detachment of trailing edge, inhibits Rac1
Rho GTPase
Does NMII move towards the + or - end
+ end of action filament via hydrolysis of one ATP
Proteolysis by MMPs can generate
signalling molecules
Proteoglycan functions
1) serve as a reservoir for growth factors
2) facilitate contact between proteins and cell-surface receptors
3) protect proteins from proteolytic degradation
Fibronectin
1) Used by cells to migrate
2) Remodeling of fibronectin can send a signal for integrity to remodel and vice versa
3) 2 monomers linked by disulfide bonds
Tay-Sachs
An inherited form of lysosomal storage disease which progressively destroys neurons in the brain and spinal cord. characterized by a cherry-red spot.
Peroxisomes
responsible for beta-oxidation and synthesis of certain glycerolipids.
X-linked adrenoleukodystrophy
Peroxisomal disorder: FAs can’t get to the peroxisome for breakdown
Zellweger syndrome
biogenesis disorder
Glycerophospholipid structure
Polar head (choline, serine), phosphate and glycerol group connection, 2 fatty acid tails
Intermediate filaments function
mechanical integrity, and providing a scaffold for signaling molecules
IF assembly
two monomers form a coiled-coil dimer, two coiled-coil dimers form a staggered tetramer, two tetramers pack together, eight tetramers twist together into a ropelike filament
Actin filament function
Important for cell shape, locomotion
microtubule assembly
hollow cylinders of 13 protofilaments made up of alpha/beta dimers
Can ATP be a co-substrate?
Only if it provides energy. If Pi is part of the end product, then it is a regular substrate
Cofactors are:
small, nonprotein molecules
Holoenzyme
Enzyme with a cofactor
Apoenzyme
Enzyme without a cofactor
Organic cofactors are called:
coenzymes
Does ∆G provide information about the rate of the reaction?
No
∆G equation
∆G = ∆Gº’+RTln([Products]/[Reactants])
Relatively small changes in ∆G‡ lead to
relatively large changes in overall reaction rate
What does the Induced Fit theory explain that the Lock and Key theory does not?
Transition state stabilization, they both explain enzyme specificity but experimental data supports induced fit
Four strategies besides binding energy (major pathway) to form and stabilize the transition state
1) Covalent catalysis: covalent bond forms between substrate and enzyme
2) General acid-base catalysis: molecule (not H2O) becomes a proton donor/acceptor
3) Catalysis by approximation: reactants are aligned and held close together
4) Metal ion catalysis
First order
The rate of product formation is directly proportional to concentration of reactant
Second order
The rate of product formation is directly proportional to the product of the concentration of two reactants
Michaelis-Menten equation
Vo = Vm[S]/Km+[S]
equation for fraction of active sites filled
Vo/Vmax = [S]/Km+[S] = fraction of active sites filled
5 assumptions of the Michaelis Menten equation
1) ES complex exists
2) no back reaction
3) initial velocities at t=0
4) steady state for [ES]
5) oversaturation of substrate
catalytic efficiency equation
Kcat/Km
- perfect enzymes have the highest ratio
- if substrates have a similar ration, the one with the highest Kcat is usuallly the best
2 types of sequential reactions
1) ordered sequential: A, then B, then P then Q
2) random sequential: B then A, then P then Q
ping-pong reaction
A binds, then P releases, then B binds, then Q releases
What are transition-state analogs
A form of competitive inhibitor that represent the transition state and bind the enzyme more tightly (more potent inhibitors)
effect of uncompetitive inhibitor on kcat/km
stays the same
effect of competitive and noncompetitive on kcat/km
Decreases
Inhibitor if the relative rate decreases with increasing [S]
Uncompetitive Inhibitor
Inhibitor if the relative rate increases with increases [S]
Competitive Inhibitor
Inhibitor if the relative rate stays the same with increasing [S]
Noncompetitive Inhibitor
Equation for % inhibition
(1-relative rate)*100
Relative rate: Vo,i/Vo
Vo,i = initial reaction rate with inhibitor
Vo = initial reaction rate without inhibitor
Equation for relative rate
Relative rate: Vo,i/Vo
Vo,i = initial reaction rate with inhibitor
Vo = initial reaction rate without inhibitor
3 types of irreversible inhibitors
1) Group-specific reagents
2) Affinity labels
3) Suicide inhibitors
1 - least specific
3 - most specific
Group-specific reagents
Type of irreversible inhibitor: covalently interact with a specific enzyme side chain to inactivate the enzyme
Affinity labels
Type of irreversible inhibitor: Structurally similar to the substrate and bind active-site residues
Suicide Inhibitors
Type of irreversible inhibitor: substrate analog that binds the active site and tricks the enzyme into catalysis, creating an intermediate which results in permanent inactivation. MOST SPECIFIC
4 methods of enzymatic regulation
1) Allosteric control: sigmoidal
2) Regulatory proteins
3) Reversible covalent modifications
4) Proteolytic activation
Homotropic effector
Type of allosteric enzymatic control where the effector is the same as the substrate and binds near the active site, almost always increasing activity
Heterotropic effector
Type of allosteric enzymatic control, effector is different from the substrate, “true” allosteric site and interaction increases or decreases activity
What occurs when a positive effector binds to an allosteric site?
Activity is increased
What occurs when a negative effector binds to an allosteric site?
Activity is decreased
Explain the ATCase example
similar to Hgb, R state allows for more substrate binding. CTP is negative effector and stabilizes T, ATP is positive effector and stabilizes R, Aspartate is the substrate and stabilizes R
PKA example
Inhibitory: Needs cAMP to bind to allow PKA to leave, allowing the enzyme active site to be open and active
Reversible covalent modifications
Kinases and Phosphatases: overall cycle (phosphorylation+dephosphorylation) hydrolyzes one ATP and the overall ∆G is negative
Proteolytic activation
1) IRREVERSIBLE
2) OUTSIDE of the cell
3) NO ATP
VIIIa
Antihemophillic factor: lack of this leads to classic hemophilia
ACTIVATED BY THROMBIN, leading to positive feedback
Intrinsic pathway
activated by damage of vasculature, involves anti hemophilic factor (lack of this is classic hemophilia)
Extrinsic pathway
Activated by external trauma
VII is activated by THROMBIN
Contains tissue factor X
Thrombin
- Most important enzyme, cleaved from prothrombin,
- can act on VIIIa and VII as positive feedback to upregulate intrinsic and extrinsic pathways respectively
- can activate factor C to shut down the pathway
- can act on transglutaminase to promote cross-linking of fibrin into hard clot
How is the soft clot formed?
fibrinogen is cleaved to fibrin
How is the hard clot formed?
Fibrin is catalyzed by transglutaminase to form a cross-linked hard fibrin clot
Characteristics of the blood clotting cascade
1) Signal amplification at each step
2) rapid response to trauma
3) cascade of zymogen activations
Heparin mechanism
Anticoagulant, Irreversible inactivation of Thrombin
(via antithrombin III)
tPA
Clot Buster
degrades fibrin clots via activation of plasminogen (serine protease)
Tyrosin Kinase Inhibitor
Imitates ATP and binds the ATP-binding pocket to prevent phosphorylation of substrate
TKI resistance
mutations can develop in proteins locking tyrosine kinase into active conformation
Branched chain amino acids
Valine, Leucine, Isoleucine, sometimes Alanine included
Cluster together for stability, hydrophobic
Mnemonic for uncharged polar amino acids
CYST NQ
Which three amino acids favor alpha helix formation?
Methionine, Alanine, Leucine
Each turn of the Alpha helix is about how many residues?
3.6
Peptide bonds H-bond with residues how many away?
4
Why are disordered protein regions notable?
They allow for interactions with multiple other proteins, such as p53 which can bind four different proteins with this region
Hsp70 functions:
co-translationally
Hsp60 functions:
post-translationally
Ubiquitination steps
1) E1 picks up Ub
2) E1 primes E2/3 with Ub
3) E2/3 binds target protein and attaches Ub
4) cycles through
How does E1 pick up Ub
Ub is bound to a cysteine via a thioester bond using the hydrolysis of ATP to AMP
How does E1 transfer Ub to E2/3
Moves from E1 cysteine to E2/3 cysteine
How does E2/3 transfer Ub onto the target protein
Target protein contains a lysine side chain degradation signal. Ub is transferred to the epsilon amino group via an iso-peptide bond
How many Ub for protein to go to proteasome
4
BiP
Binding protein: Hsp70 chaperone
- hydrolyzes ATP as proteins brought into ER
- recognizes misfolded proteins by binding amino acids usually found on the interior
PDI
promotes formation of disulfide bonds
PPI
facilitates interconversion of proline from cis to trans to facilitate protein folding
Calnexin and Calreticulin
Play a role in glucose tagged proteins attempting to exit ER to Golgi
UPR (Unfolded Protein Response)
1) increased ER protein folding
2) up regulation of chaperone and protein degrading genes
3) apoptosis
PrP-c
3 alpha helices
2 beta strands
one unstructured region
PrP-sc
multiple stacked beta strands
protease resistant core
amyloid fibril
Increased pH shifts curve to ___ and stabilizes ___ state
Left, stabilizes R state
Increased CO2 shifts curve to ___ and stabilizes ___ state
Right, stabilizes T state
Why does fetal Hb have a higher affinity for O2?
Less bound 2,3-BPG
Sickle cell disease
substitution on the beta chain, causing HbS to self-associate and cell to sickle
Alpha Thalassemia (HbH) mechanism and effects
Defect in alpha chain genes, binds O2 with high affinity and no cooperatively
Beta Thalassemia mechanism and effects
Defect in beta chain, lack of O2 cooperativity, tetramers tend to be insoluble and precipitate
Methemeglobinemia
Fe2+ oxidized to Fe3+ leading to high O2 affinity
Glutathione
Sacrificial molecule to keep ROS low and prevent inactivation of methemoglobin reductase
Methemglobin reductase
Uses NADH to reduce Fe3+
Actin filament structure
- 2 stranded helical polymers of actin subunits form each filament
- 3 types: alpha, beta, gamma
- each filament is made from one type
