Biochemistry Flashcards
Glycine
Gly, G
pKa: neutral
Group: small
Properties: not chiral; found in structural loops
Alanine
Ala, A
pKa: neutral
Group: polar
Serine
Ser, S
pKa: neutral
Group: polar
Properties: can form H-bonds; can be phosphorylated to introduce a negative charge
Threonine
Thr, T
pKa: neutral
Group: polar
Properties: can form H-bonds; can be phosphorylated to introduce a negative charge
Cysteine
Cys, C
pKa: slightly basic
Group: polar
Properties: forms disulfide bridges, important for 3 and 4 structure
Valine
Val, V
pKa: neutral
Group: nonpolar
Leucine
Leu, L
pKa: neutral
Group: nonpolar
Isoleucine
Ile, I
pKa: neutral
Group: nonpolar
Methionine
Met, M
pKa: neutral
Group: nonpolar
Properties: “start” amino acid (can also be found at other positions)
Proline
Pro, P
pKa: neutral
Group: nonpolar
Properties: the only cis-amino acid; side chain part of peptide bond; introduces kinks in α-helices; found in loops and turns
Phenylalanine
Phe, F
pKa: neutral
Group: nonpolar
Properties: aromatic
Tyrosine
Tyr, Y
pKa: neutral
Group: nonpolar
Properties: aromatic; can be phosphorylated to introduce a negative charge
Tryptophan
Trp, W
pKa: neutral
Group: nonpolar
Properties: aromatic
Aspartate
Asp, D
pKa: acidic
Group: negatively charged at physiological pH
Properties: side chain can form salt bridge
Glutamate
Glu, E
pKa: acidic
Group: negatively charged at physiological pH
Properties: side chain can form salt bridge
Asparagine
Asn, N
pKa: neutral
Group: polar
Properties: side chain can form H-bonds
Glutamine
Gln, Q
pKa: neutral
Group: polar
Properties: side chain can form H-bonds
Histidine
His, H
pKa: slightly acidic
Group: polar
Properties: aromatic; can be positively charged at acidic pH
Lysine
Lys, K
pKa: basic
Group: positively charged at physiological pH
Properties: side chain can form salt bridge; can be acetylated to mask the positive charge (important in DNA-protein interaction)
Arginine
Arg, R
pKa: basic
Group: positively charged
Properties: side chain can form salt bridge
Acid-base chemistry of AA
- At low (acidic) pH: full protonated
- When pH = pI: zwitterion
- At high (basic) pH: full deprotonated
- pI is determined by averaging the pKa values that refer to protonation and deprotonation of the zwitterion
Peptide bonds
Formation is a condensation (dehydration) rxn with a nucleophilic amino group attacking an electrophilic carbonyl; peptide bonds are broken by hydrolysis
Tertiary structure
3D structure stabilized by hydrophobic interactions, acid-base interactions (salt-bridges), H-bonding, and disulfide bonds
Quaternary structure
Interactions between subunits; heat and solutes can cause denaturation
Polyacrylamide gel electrophoresis (PAGE)
Proteins migrate through porous matrix according to size and charge; (1) native PAGE is used to analyze the protein in folded state (2) SDS-PAGE uses detergent to break all noncovalent interactions and analyzes the unfolded state
Reducing reagents
Can be used to break covalent disulfide bonds
Structural proteins
Generally fibrous; include collagen, elastin, keratin, actin, and tubulin
Motor proteins
Capable of force generation through a conformation change; include myosin, kinesin, and dynein
Cell adhesion molecules (CAM)
Bind cells to other cells or surfaces; include cadherins, integrins, and selectins
Enzyme-linked receptors
Participate in cell signaling through extracellular ligand binding and initiation of second messenger cascades
G protein-coupled receptors
Have a membrane-bound protein associated with a trimeric G protein; they also initiate second messenger systems
Binding site, impact on Km, impact on Vmax
Competitive: active site, increases, no change
Noncompetitive: allosteric site, no change, decreases
Mixed: allosteric site, increases/decreases, decreases
Uncompetitive: enzyme-substrate complex, decreases, decreases
Saturation kinetics
As substrate concentration increases, the reaction rate also increases until a maximum value is reached
v = vmax [S] / km + [S]
- At one-half Vmax, [S] = Km
Lineweaver-Burk
kcat = vmax / [enzyme]
Catalytic efficiency = kcat / Km
Ligases
Responsible for joining two large biomolecules, often of the same type
Isomerases
Catalyze the interconversion of isomers, including both constitutional and stereoisomers
Lyases
Catalyze cleavage without the addition of water and without the transfer of electrons; the reverse reaction (synthesis) is usually more biologically important
Hydrolases
Catalyze cleavage with the addition of water
Oxidoreductases
Catalyze oxidation-reduction reactions that involve the transfer of electrons
Transferases
Move a functional group from one molecule to another
Michaelis-Menten
Cooperative enzymes show a sigmoidal curve
Enzymes
Like all catalysts, lower the activation energy necessary for rxns; they do not alter the free energy or enthalpy change that accompanies the rxn nor the final equilibrium position; rather, they change the kinetics (rate) at which equilibrium is reached
Aldoses
Sugars with aldehydes as their most oxidized group
Ketoses
Sugars with ketones as their most oxidized group
D vs. L sugars
Sugars with the highest-numbered chiral carbon with the -Oh group on the right are D-sugars; those with the -OH on the left are L-sugar; D- and L-forms of the same sugar are enantiomers
Diastereomers
Differ at least one - but not all - chiral carbons
Also include: (1) epimers differ at exactly one chiral carbon (2) anomers are a subtype of epimers that differ at the anomeric carbon
Anomeric carbon
The new chiral center formed in ring closure; it was the carbon-containing the carbonyl in the straight-chain form
- α-anomers have the -OH on the anomeric carbon trans to the free -CH2OH group
- β-anomers have the -OH on the anomeric carbon cis to the free -CH2OH group
Mutarotation
One anomeric form shifts to another, with the straight-chain form as an intermediate
Monosaccharides
Single carbohydrate units and can undergo three main reactions: oxidation-reduction, esterification, and glycoside formation (the basis for building complex carbs and requires the anomeric carbon to link to another sugar)
- Sugars with an -H replacing an -Oh are termed deoxy sugars
Disaccharides
Sucrose (glucose-α-1,2-fructose), lactose (galactose-β-1,4-glucose), and maltose (glucose-α-1,4-glucose)
Cellulose
Main structural component of plant cell walls; main source of fiber in the human diet
Starches
Amylose and amylopectin; main energy storage forms for plants
Glycogen
A major energy storage form for animals
Reducing sugars
Any sugar with an anomeric carbon not bound in a glycosidic bond will react with reagents like Tollens’ and Benedict’s
Nucleotide vs nucleosides
Nucleosides contain a five-carbon sugar bonded to a nitrogenous base; nucleotides are nucleosides with one to three phosphate groups added; ATP is a high-energy nucleotide with an adenosine nucleoside
Watson-Crick Model
- DNA backbone is composed of alternating sugar and phosphate groups, and is always read 5’ to 3’
- There are two strands with antiparallel polarity, wound into a double helix
- A-T and A-U with two H-bonds
- C-G with three H-bonds
Chargaff’s rules
Purines and pyrimidines are equal in number in a DNA molecule; the amount of A equals T and vise versa
Euk. chromosome organization
In euk, DNA is wound around histone proteins to form nucleosomes, which may be stabilized by another histone protein
- DNA and its associated histones make up chromatin in the nucleus
Heterochromatin
Dense, transcriptionally silent DNA
Euchromatin
Less dense, transcriptionally active DNA
Telomeres
Are the ends of chromosomes; they contain high GC-content to prevent DNA unraveling
Centromeres
Hold sister chromatids together until they are separated during anaphase in mitosis; they also contain a high GC-content
Recombinant DNA
DNA composed of nucleotides from two different sources
DNA cloning
Introduces a fragment of DNA into a vector plasmid; a restriction enzyme (restriction endonuclease) cuts both the plasmid and the fragment, leaving them with sticky ends, which can bind
- Restriction enzyme sites are often palindromic
DNA replication
Is semiconservative: one old parent strand and one new daughter strand is incorporated into each of the two new DNA molecules
DNA polymerase
Synthesizes new DNA strands, reading the template DNA 3’ to 5’ and synthesizing the new strand 5’ to 3’
- The leading strand requires only one primer and can then be synthesized continuously
- The lagging strand requires many primers and is synthesized is discrete sections called Okazaki fragments
Origin of replication
Pro: one per chromosome
Euk: multiple per chromosome
Unwinding of DNA double helix
Pro: helicase
Euk: helicase
Stabilization of unwound template strands
Pro: single-stranded DNA-binding protein
Euk: single-stranded DNA-binding protein
Synthesis of RNA primers
Pro: primase
Euk: primase
Synthesis of DNA
Pro: DNA polymerase III
Euk: DNA polymerase α, δ, ε
Removal of primers
Pro: DNA polymerase I (5’ - 3’ exonuclease)
Euk: RNase H (5’ - 3’ exonuclease)
Replacement of RNA with DNA
Pro: DNA polymerase I
Euk: DNA polymerase δ
Joining of Okazaki fragments
Pro: DNA ligase
Euk: DNA ligase
Removal of positive supercoils ahead of advancing replication forks
Pro: DNA topoisomerases (DNA gyrase)
Euk: DNA topoisomerases
Synthesis of telomeres
Pro: N/A
Euk: Telomerase
Genomic libraries
Contain large fragments of DNA, including both coding and noncoding regions of the genome; they cannot be used to make recombinant proteins or for gene therapy
cDNA libraries (expression libraries)
Are generated by reverse transcribing mRNA of sample tissue. The resulting DNA library only includes exons of expressed genes; they can be used to make recombinant proteins or for gene therapy
PCR
An automated process by which millions of copies of a DNA sequence can be created from a very small sample by hybridization (the joining of complementary base pair sequences)
SNOW DROP
Southern - DNA
Northern - RNA
Western - proteins
Deoxyribonucleotides
Terminate the DNA chain because they lack a 3’ - OH group
Central dogma
DNA –> RNA –> proteins
Initiation and termination
Initiation: AUG (methionine)
Termination: UAA, UGA, UAG
- Redundancy and wobble (third base in the codon) allow mutations to occur without affecting the protein
Point mutations
Silent: no effect on protein synthesis
Nonsense (truncation): produce a premature stop codon
Missense: produce a codon that codes for a different AA
Frameshift: result from nucleotide addition or deletion and change the reading frame of subsequent codons
RNA is structurally similar to DNA except:
- Substitution of a ribose sugar for deoxyribose
- Substitution of uracil for thymine
- Single-stranded instead of double
Major types of RNA
mRNA: carries the message from DNA in the nucleus via transcription of the gene; travels into the cytoplasm to be translated
tRNA: brings in AA; recognizes the codon on the mRNA using its anticodon
rRNA: makes up much of the ribosome; enzymatically active
Transcription steps
- Helicase and topoisomerase unwind DNA double helix
- RNA polymerase II binds to TATA box within promoter region of gene
- hnRNA synthesized from DNA template (antisense) strand
Posttranscriptional modifcations
- 7-methylguanylate triphosphate cap added to 5’ end
- Polyadenosyl (poly-A) tail added to 3’ end
- Splicing done by spliceosome; introns removed and exons ligated together. Alternative splicing combines different exons to acquire different gene products
Translation steps
- Initiation, elongation, termination
- Posttranslational modifications: (1) folding of chaperones (2) formation of quaternary structure (3) cleavage of proteins or signal sequences (4) covalent addition of other biomolecules
Transcription factors
- Promoters are within 25 base pairs of the transcription start site
- Enhancers are more than 25 base pairs away from the transcription start site
Operons (Jacon-Monod model)
Are inducible or repressible clusters of genes transcribed as a single mRNA
Osmotic pressure
A colligative property, is the pressure applied to a pure solvent to prevent osmosis and is related to the concentration of the solution
∏ = iMRT
Passive transport
Simple diffusion: does not require a transporter; small, nonpolar molecules passively move from an area of high concentration to an area of low concentration until equilibrium is achieved
Osmosis: diffusion of water across a selectively permeable membrane
Facilitated diffusion: uses transport proteins to move impermeable solutes across the cell membrane
Active transport
Requires energy in the form of ATP (primary) or an existing favorable ion gradient (secondary); secondary active transport can be further classified as symport or antiport
Endocytosis and exocytosis
Methods of engulfing material into cells or releasing material to the exterior of cells; both via the cell membrane
Pinocytosis
Ingestion of liquid into the cell from vesicles formed from the cell membrane
Phagocytosis
Ingestion of solid material
Glucokinase
Present in liver and pancreatic β cells, responsive to insulin; phosphorylates glucose
Hexokinase
Present in all tissue; phosphorylates glucose to trap it in cells
Phosphofructokinase-1 (PFK-1)
Rate-limiting step
Phosphofructokinase-2 (PFK-2)
Produces F2-6-BP, which activates PFK-1
Glyceraldehyde-3-phosphate dehydrogenase
Produces NADH
3-phosphoglycerate kinase and pyruvate kinase
Perform substrate-level phosphorylation
The NADH produced in glycolysis:
Is oxidized aerobically by the mitochondrial electron transport chain and anaerobically by cytoplasmic lactate dehydrogenase
Glycolysis
Occurs in the cytoplasm of all cells, and does not require O2; yields 2 ATP per glucose
Pyruvate dehydrogenase
Converts pyruvate to acetyl-CoA; stimulated by insulin and inhibited by acetyl-CoA
ETC
- Takes place on the matrix-facing surface of the inner mitochondrial membrane
- NADH donates e to the chain, which are passed from one complex to the next; reduction potentials increase down the chain, until the electrons end up on O2, which has the highest reduction potential
- NADH cannot cross the inner membrane, so must use one of two shuttle mechanisms to transfer its e to energy carriers in the mitochondrial matrix: glycerol 3-phosphate shuttle or the malate-aspartate shuttle
Oxidative phosphorylation
The proton-motive force is the electrochemical gradient generated by the ETC across the inner mitochondrial membrane; the intermembrane space has a higher concen. of protons than the matrix; this gradient stores energy, which can be used to form ATP via chemiosmotic coupling
Summary of energy yield of carbohydrate metabolism processes:
Glycolysis: 2 NADH and 2 ATP
Pyruvate dehydrogenase: 1 NADH (2 NADH per molecule of glucose because each glucose forms two molecules of pyruvate)
CAC: 3 NADH, 1 FADH2, and 1 GTP (6 NADH, 2 FADH2, and 2 GTP per molecule of glucose)
- Each NADH: 2.5 ATP; 10 NADH form 25 ATP
- Each FADH2: 1.5 ATP
- GTP converted to ATP
= 32 ATP per molecule of glucose; 30-32 per molecule is the commonly accepted range
ATP synthase
Enzyme responsible for generating ATP from ADP and an inorganic phosphate (Pi)
Glycogenesis (glycogen synthesis)
The building of glycogen uses two main enzymes:
- Glycogen synthase, which creates α-1,4 glycosidic links between glucose molecules; it is activated by insulin in the liver and muscles
- Branching enzyme, which moves a block of oligoglucose from one chain and connects it as a branch using an α-1,6 glycosidic link
Glycogenolysis
Breakdown of glycogen using two main enzymes:
- Glycogen phosphorylase, removes single glucose 1-phosphate molecules by breaking α-1,4 glycosidic links. In the liver, it is activated by glucagon to prevent low blood sugar. In exercising skeletal muscle, it is activated by epinephrine and AMP to provide glucose for the muscle itself
- Debranching enzyme, moves a block of oligoglucose from one branch and connects it to the chain using an α-1,4 glycosidic link
Gluconeogenesis
Occurs in both the cytoplasm and mito; predominantly in the liver.
- Most is just reverse of glycolysis, using same enzymes
Three irreversible steps of glycolysis must be bypassed by different enzymes:
- Pyruvate carboxylase and PEP carboxykinase bypass pyruvate kinase
- Fructose-1,6-bisphosphatase bypasses phosphofructokinase-1
- Glucose-6-phosphatase bypasses hexokinase/glucokinase
Pentose phosphate pathway
Occurs in the cytoplasm of most cells, generating NADPH and sugars for biosynthesis; rate-limiting enzyme is glucose-6-phosphate dehydrogenase, which is activated by NADP+ and insulin and inhibited by NADPH
Metabolic states
- Postprandial/well-fed (absorptive): insulin secretion is high and anabolic metabolism prevails
- Postabsoptive (fasting): insulin secretion decreases while glucagon and catecholamine secretion increases
- Prolonged fasting (starvation): dramatically increases glucagon and catecholamine secretion; most tissues rely on FAs
Tissue-specific metabolism
- Liver: maintains blood glucose through glycogenolysis and gluconeogenesis; processes lipids, cholesterol, bile, urea, and toxins
- Adipose: stores and releases lipids
- Resting muscle: conserves carbohydrates as glycogen and uses free FAs for fuel
- Active muscle: may use anaerobic metabolism, oxidative phosphorylation, direct phosphorylation (creatine phosphate), or FA oxidation
- Cardiac muscle: uses FA oxidation
- Brain: uses glucose except in prolonged starvation, when it can use ketolysis
Lipid transport
Via chylomicrons, VLDL, IDL, LDL, and HDL
Cholesterol metabolism
- Cholesterol may be obtained through dietary sources or through synthesis in the liver
- The key enzyme in cholesterol biosynthesis is HMG-CoA
- Palmitic acid, only FA that humans can synthesize
- Fatty acid oxidation occurs in the mito. following transport by the carnitine shuttle, via β-oxidation
- Ketone bodies form (ketogenesis) during a prolonged starvation state due to excess acetyl-CoA in the liver; ketolysis regenerates acetyl-CoA for use as an energy source in peripheral tissues
