Biochem exam 2

protein function

transport, storage, signaling

Ligands

molecules bound to proteins not chemically modified

Substrates

molcules bound to proteins chemically modifies

“Y”

fraction of binding sites occupie; Y = ^[L]/_{Kd + [L]}

K_d

rate of dissociation; K_d = ^[P][L]/_[PL]

On graph, K_d equals

[L] where 50% of binding sites occupied

K_d > 10um

weak binding, lower affinity

K_d < 10um

strong binding, higher affinity

Distal Histidine

prevents Fe oxidation by crowding and makeing O2 bent

Myoglobin

8-alpha helixes; only reaches tertiary; 1 heme group; stores O2 so new O2 can enter cell spontaneously by diffusion

Myoglobin binding curve

hyperbolic so no cooperativity; high O2 affinity

Hemoglobin

O2 transport from lungs to tissues; 2 alpha + 2 beta subunits; a1B2 strong interactions; 4 heme groups

T state

low affinity

R state

high affinity

Cooperativity

first binding increases affininty at remaining sites

T state interactions

ionic interactions between His and Asp stabilize; when O2 binds, beta subunits slide past each other, R-groups rearrange, ioninc interactions weaken

Hill Equation

Y = ([L]ⁿ)/(Kdⁿ + [L]ⁿ)

Hill constant (n_H)

degree of cooperativity between subunits

n_H = 1 : no cooperativity

n_H > 1 : positive cooperativity

n_H < 1 : negative cooperativity

Cooperativity concerted model

only 2 possible conformations for all 4 subunits; no intermediate states; all subunits change simultaneously; equilibrium between T and R states; related to lock and key model

Cooperativity sequential model

many possible T and R conformations in one Hgb protein; each binding event cause conformatinoal change that gradually increases affinity; related to induced fit model

Allosteric protein

ligand binding at one site affects binding properties of different site on same protein

Homotropic

normal ligand is allosteric regulator (ex: O2 binding to Hb)

Heterotropic

different ligand affects binding of normal ligand (ex: BPG binding to Hb)

pH affect on O2 binding

lower pH = lower affinity

Bohr Effect

how pH change can affect O2 affinity 

Carbamino Terminal Residue

in lower pH; forms addition salt bridges due to ’”-” stabilizing T state

BCG

negative heterotropic regulaor; binds to “+” charged central cavity of Hb; stabilizes T state

BCG important for

O2 release at high altitudes

Enzyme

increase reaction rate of forward and backward reaction; don’t affect equilibria

transition state

point where decay to substrate or product equally likely

activation energy

difference between graound state and transition state energy

reactiion intermediate

species on reaction pathway that has finite chemical lifetime (ex: ES and EP complexes)r

rate-limiting steps

step with highest activation energy; determines overall rate of reaction

rate equation for first order

v = k[S]; k = s^-1

rate equation for second order

v = k[S₁][S₂]; k = m^-1s^-1

How enzymes increase reaction rate

1) bring substrate in close proximity and correct orientation

2) stabilize transition state; enzyme binds tightly to transition state using binding energy to lower activation barrier (ΔG released and used to decrease energy needed to reach transition state)

3) General acid-base catalysis: due to pH changes in environment

4) Covalent catalysis: use R-groups of aa to facilitate peptide bond breakage

5) Metal ion catalysis: metal ion can bound to enzyme and help enzyme perform reaction that aa usually can’t

Lock and key model

debunked; slows down reaction instead speed up cause would need more energy from ES to transition state

Optimized binding energy in transition state

only room for substrate in active site, so all water removed; need to replace H2O H-bonds with H-bonds of aa in binding site

Specificity

can differentiate substrate from comepeting molecule; more exergonic ΔG_B = greater specificity

Componenets that determine magnitude of ΔG transition state

1) molecule entropy in solution (remove randomness; once substrate bound, don’t depend on randomness for reaction to proceed)