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principles of protein structure and folding
- Levels of structure
- Primary structure: sequence of amino acids, covalently attached by peptide bonds.
- Secondary structure: local backbone interactions involving hydrogen bonding.
- α helix
- Right handed.
- N-H group of nth residue H-bond with C=O group of n-4 residue.
- Core of helix tightly packed, held together by van der Waals.
- R groups stick outward (and downward).
- β-sheet
- Can be antiparallel (more stable) or parallel (less stable).
- H-bond between backbones (N-H and C=O) of adjacent chains.
- R groups stick out above and below the sheet.
- Other helices: thinner or wider than the alpha helix can form.
- Tertiary structure: 3D structure determined by both backbone and R group interactions.
- Quaternary structure: interactions between different chains (subunits).
- Folding
- Primary structure determines secondary structure, which determines tertiary structure.
- This principle is proved by the RNase experiment: denatured RNase refolds to its active form.
- Interactions:
- Covalent: peptide bond, disulfide bond. Eg. Subunits of hemoglobin.
- Hydrogen bonding: backbone and R groups.
- Electrostatic: +charged R group (basic) with -charged R group (acidic).
- Complex with metal ions or prosthetic groups. Eg. Heme group of globins.
- Hydrophobic interactions: nonpolar residues packed inside protein, shielded from water, driven by more favorable entropy (thermodynamics).
- Chaperones:
- Assist in correct folding of proteins.
- Prevent unfolded proteins from unwanted interactions with other proteins.
- Prevent protein from folding into wrong, alternative conformations.
- Prions: abnormally folded proteins that induce their normal counterparts to misfold.
- Mad cow disease (spongiform encephalopathy disease in humans).
- Fibrous proteins:
- Chains don't fold back on themselves, adopt a fiberous conformation.
- Water insoluble.
- Globular proteins:
- Chains fold back on themselves, adopt a spherical conformation.
- Water soluble.
enzymes: kinetics, reaction mechanisms
- Kinetics:
- Review premed kinetics vs thermodynamics here.
- Kinetics is about how fast a reaction occurs.
- How fast a reaction occurs depend on the activation energy.
- Enzymes lower the activation energy so a reaction proceeds faster.
- Enzymes increase the rate for both the forward and the reverse reaction.
- Enzymes do not change ΔG.
- Michaelis-Menten enzyme kinetics:
- E + S ↔ ES → E + P
- E is enzyme. S is substrate. ES is enzyme-substrate complex. P is product.
- E + S → ES: k1
- E + S ← ES: k-1
- ES → E + P: k2
- V = Vmax[S]/KM + [S]
- KM = k-1 + k2/k1
- When [S] = KM, V = 1/2 Vmax
- Lineweaver-Burk double-reciprocal plot:
- y-axis = 1/V
- y-intercept = 1/Vmax
- Competitive inhibitors don't change Vmax, so y-intercept is unchanged. Plot rotates left about the y-intercept.
- x-axis = 1/[S]
- x-intercept = -1/KM
- Noncompetitive inhibitors don't change KM, so x-intercept is unchanged. Plot rotates left about the x-intercept.
- Reaction mechanisms:
- Induced fit model:
- substrate bind to active site.
- enzyme changes conformation as to stabilize the transition state.
- substrate becomes transition state, then turn into products.
- Lock and key model is outdated. If substrate were to bind enzyme like a key, then no catalysis would occur.
- Mechanisms:
- Acid-base catalysis.
- Covalent catalysis (eg. Formation of Schiff base).
- Metal ion catalysis (metal ion bind substrates, mediate redox reactions, and stabilize negative charges).
- Electrostatic catalysis (charge distribution of active site guide substrate binding).
- Proximity and orientation (brings reactants close together, in the right orientation for reaction to occur).
- Transition state binding.
- Cofactors give enzymes the ability to catalyze certain reactions that the protein alone won't suffice.
- Metal ions:
- Zn2+. Deficiency causes delayed wound healing, hypogonadism, decreased adult hair, may predispose to alcoholic cirrhosis.
- Coenzymes (organic molecules): eg. NAD+.
- Vitamins are commonly coenzyme precursors.
-
| Coenzyme |
Reaction |
Vitamin precursor |
Disease if deficienty |
| Biotin |
Carboxylation |
|
Dermatitis, alopecia, enteritis (excessive egg white ingestion) |
| Thiamine pyrophosphate |
Aldehyde transfer |
Thiamine (B1) |
Beriberi (Ber1Ber1) |
| Flavin |
Redox |
Riboflavin (B2) |
Cheilosis, corneal vascularization |
| Nicotinamide (NAD+) |
Redox |
Nicotinamide (NAD+) |
Pellagra (Diarrhea, Dermatitis, Dementia, Death) |
| Coenzyme A |
Acyl transfer |
Pantothenate (B5) |
Dermatitis, enteritis, alopecia, adrenal insufficiency |
| Pyridoxal phosphate |
Amino group transfer |
Pyridoxine (B6) |
Convulsions, hyperirritability, peripheral neuropathy (inducible by INH and oral contraceptives) |
| Cobalamin (B12) |
Alkylation |
Cobalamin (B12) |
Macrocytic, megaloblastic anemia, neurological symptoms |
| Tetrahydrofolate |
One-carbon group transfer |
Folic acid |
Macrocytic, megaloblastic anemia |
| Lipoic acid |
Acyl transfer |
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- Prosthetic groups (permanently associated with enzyme): eg. Heme.
- Cofactors are changed during the reaction (eg. oxidized or reduced), and need to be changed back for the reaction to occur again.
structural and regulatory proteins: ligand binding, self-assembly
- Hemoglobin binds heme group.
- Collagen self assembly into triple helix.
- RNase fully assembles into active enzyme after being denatured.
- Self-assembly of viral coat protein.
regulatory properties
- Covalent modification.
- Phosphorylation:
- Kinases and phosphorylases attach phosphate groups to proteins.
- Phosphatases remove phosphate groups from proteins.
- Phosphate ester can be formed on amino acid residues that have an -OH group: Serine, Threonine, Tyrosine.
- Methylation.
- Hydroxylation.
- Acetylation.
- Glycosylation.
- Zymogens: enzymes that are inactive until cleaved.
- Binding of activators or inhibitors.
- Regulate the production of the enzyme.
- Negative feedback: products inhibit the production of enzyme.
- Substrates stimulate production of enzyme (eg. Lac operon)
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