CHEM351, Unit 2: protein folding + function (hemoglobin)

denaturation

loss of 3D structure that leads to loss of function

what are the ways that denaturation can occur?

chaotropic agents, ph changes, heat, and organic solvents

why do chaotropic agents cause denaturation?

they prefer to interact with the backbone, so they disrupt the folded structure

what are two chaotropic agents?

urea and guanidium hydrochloride

what did Anfisen's experiment reveal?

denaturation of ribonuclease A is reversible because primary structure is maintained

how did Anifsen perform his experiment?

he denatured ribonuclease A with b-mercaptoethanol and urea which disrupted the disulfide bonds, but when they were removed catalytic activity was restored

what determines a proteins final conformation?

hydrophobic collapse and secondary structure

hydrophobic collapse

describes the process of hydrophobic AA wanting to end up inside of a structure to provide entropic stability

kinetic traps

incorrect interactions formed while folding that require the structure to be destabilized to correct it

native structure

final structure of a protein that has the least free energy

what factors promote the formation of quaternary structures?

interface is stabilized since residues are less exposed, more binding sites for ligands, allosteric effects are possible, and folding is more efficient

binding

association of two molecules

ligand

molecules that bind to another molecule or proteins

fraction bound

the amount of ligands bound to a protein

Kd

dissociation constant, equilibrium constant for protein-ligand complex

K

association constant for protein-Iigand complex

cooperativity

how much the binding of an allosteric site facilitates binding for other sites

why does oxygen require a transporter?

it is not soluble in aqueous solutions

why does oxygen bind to the heme group?

it cannot bind directly to a metal or oxidation will occur, forming free radicals

prosthetic group

a group that is part of a protein but is not made of amino acids

heme group

protoporphyrin ring with iron bound

myoglobin

153 AA protein with one molecule of heme at the center

hemoglobin

myoglobin protein complexed to a heme group, bound to His93

HisE7

distal hemoglobin that stabilizes the o2 group

what is the equation for K?

K=PL/P*L

what is the equation for Kd?

Kd=P*L/PL

what is the fraction bound equation?

(PL)/(PL+P)

how do you calculate fraction bound without protein concentration?

(L)/(Kd+L)

how many subunits are in hemoglobin?

two alpha two beta

what are the two Hb concentrations?

T-state and R-state

T-state

tense state with more ion pairs, has low O2 affinity

R-state

relaxed state with less ion pairs, has high O2 affinity

what determines the conformation of Hb?

the O2 saturation present

what factors stabilize the T-state?

low pH and O2, high CO2

what factors stabilize the R-state?

high pH and O2, low CO2

2,3-biphosphoglycerate

protein present in RBC that reduces affinity for O2 by binding to the positive b subunits in the T-state

how will the body adjust to a high altitude?

forming more 2,3-BPG to bind to hemoglobin, which will release more oxygen to tissues