BIOL-540 M4 Protein-Ligand Interactions (cont.)

the heme prosthetic group consists of a ____ ring an a ____ ion.

porphyrin, iron (II)

the ⍺ and β globin subunits of hemoglobin have different versions used in different cell types or at different times in development. if all these versions were originally the result of gene duplication and divergent evolution, we would call them ____.

paralogs

the ⍺ and β subunits of hemoglobin and myoglobin have ____ structural similarity to each other.

high (low sequence similarity)

what is the [ligand] needed for half-maximal binding by the protein?

Kd

structural adaptation between protein and ligand in response to binding

induced fit

low-affinity form of a protein that binds its ligand less easily

tense (T) state

high-affinity form of a protein (like hemoglobin) that binds its ligand, such as oxygen, more easily

relaxed (R) state

predominant conformation of deoxyhemoglobin

T state (not oxygenated)

how is T state stabilized?

O2 binding to hemoglobin in the __ state triggers a conformational change to the __ state.

T, R

what 3 things occur when O2 binds to T-state hemoglobin?

- the 2 ⍺β subunit pairs slide past each other and rotate, decreasing ⍺1β2 and ⍺2β1 interactions

- pocket between β subunits narrows

- some ion pairs that stabilize the T state break, and new ones form in the R state

myoglobin has 153 AA residues and __ molecules of heme with bends named after the ____ segments they connect.

1, ⍺-helical

how many binding sites for O on myoglobin?

1

how many binding sites for O on hemoglobin?

4

in distal tissues, we want ____ affinity binding. in lungs, we want ____ affinity binding.

low, high

so oxygen will be released to tissues and binds in lungs

the hemoglobin hybrid binding curve for O2 is ____.

sigmoidal (not hyperbolic)

leghemoglobin is a plant oxygen-binding protein found in root nodules that contain bacteria that fix atmospheric nitrogen. this oxygen-binding curve is ____.

hyperbolic

binding of a ligand to one site affects the binding properties of another site on the same protein

allosteric protein (like hemoglobin)

ligands that bind to an allosteric protein to induce a conformational change

modulators

normal ligand and modulator are identical

homotropic

modulator is a molecule other than the normal ligand

heterotrophic

____ is a homotropic modulator of oxygen binding to hemoglobin.

oxygen

formula for equilibrium for a simple P-L interaction

formula for equilibrium for a protein with n binding sites

formula for association constant for a protein with n binding sites

formula for Y for a protein with n binding sites

true or false: at equilibrium, ratio of bound to free proteins (Y) is constant.

true; individual proteins are constantly binding/releasing ligands

[L] at which ½ of the available ligand-binding sites are occupied (Y = 0.5)

1/Ka (Kd)

as Kd increases, affinity for substrate ____/

decreases

when [L] = __, half of the ligand-binding sites are occupied.

Kd

hill equation

hill plot for binding O2 to hemoglobin

which molecule is a heterotrophic modulator of oxygen binding to hemoglobin?

2,3-biphosphoglycerate

a newly discovered protein has multiple subunits, each with a single ligand-binding site. binding of ligand to one site is completely independent of binding to any of the other sites. what is the hill coefficient?

= 1

a newly discovered protein has multiple subunits, each with a single ligand-binding site. binding of ligand to one site increases the binding affinity of other sites for the ligand. what is the hill coefficient?

> 1

there are 2 binding sites for acetylcholine on its receptor. what would be the shape of a hill plot if there were cooperativity of binding?

sigmoidal, with a sharp change in slope (similar to O binding to hemoglobin)

model of cooperative binding that states that all subunits are in the same conformation and the ligand binds more tightly to the R state

MWC model (aka concerted model)

model of cooperative binding that states that each subunit can be in either conformation and equilibrium is altered as additional ligands are bound, progressively favoring the R state

if nH = 1, ligand binding is...

not cooperative

if nH > 1, there is ____ cooperativity.

positive

if nH < 1, there is ____ cooperativity.

negative

in the model of cooperativity, __ state is low affinity and __ state is high affinity.

T, R

in peripheral tissues, O2 is released by ____.

hemoglobin

2 end products carried by hemoglobin after cellular respiration?

H+ and CO2

the binding of H+ and CO2 is ____ related to the binding of O2.

inversely

enzyme that catalyzes hydration of CO2 to bicarbonate

carbonic anhydrase

carbonic anhydrase catalyzes ____ of CO2 to bicarbonate.

hydration

the pull effect of pH and [CO2] on the binding and release of O2 by hemoglobin

bohr effect

according to the bohr effect, ____ hemoglobin is less likely to bind oxygen.

protonated

according to the bohr effect, oxygen in hemoglobin is less likely to bind ____.

free protons

bohr effect: when [O2] is high, hemoglobin ____ O2 and ____ H+.

binds, releases

bohr effect: when [O2] is low, hemoglobin ____ O2 and ____ H+.

releases, binds

the bohr effect is the phenomenon where hemoglobin's affinity for oxygen decreases when CO2 levels ____ or pH ____.

rise, drops

hemoglobin releases O2 more easily when CO2 is ____ and pH is ____.

high, low

hemoglobin binds O2 more tightly when CO2 is ____ and pH is ____.

low, high

how does CO2 binding to hemoglobin contribute to bohr effect?

when it binds, an H+ is released, leading to conformational shift that brings hemoglobin back to T state

2,3-bisphosphoglycerate (BPG) uses ____ modulation/

heterotophic allosteric

____ binds to hemoglobin on a distant site from the O2 binding site.

BPG

when BPG binds to distant site on hemoglobin, affinity for hemoglobin for O2 ____.

greatly reduces

as altitude increases, BPG concentration ____.

increases

true or false: BPG concentration greatly affects O2 affinity at pO2 present in lungs.

false; in peripheral tissues, has little effect in lungs

lowered oxygenation of peripheral tissues

hypoxia

hypoxia causes ____ in production of BPG.

increases

hypoxia creates a larger difference between ____ bound in lungs vs. peripheral tissues at lower pO2 in lungs.

O2 (hemoglobin gives more O2 to peripheral tissues; holds on to less)

BPG binds to the cavity between the __ subunits in the T state.

β (this cavity is lined with positively charged residues_

BPG binds to and stabilizes the __ state of hemoglobin.

T (lower affinity)

BPG leads to an ____ in Kd for oxygen in peripheral tissues.

increase

BPG leads to an increase in Kd for oxygen in ____ tissues.

peripheral

what is the actual, significant effect of 2,3-bisphosphoglycerate on O binding by hemoglobin?

increase in Kd for oxygen in peripheral tissues; more effective dumping from hemoglobin in peripheral tissues

a fetus synthesizes ____ hemoglobin.

a2𝛾2

difference in adult and fetal hemoglobin?

fetal hemoglobin has...

- lower BPG affinity

- higher O2 affinity

important for taking O2 from mom's blood

sickle cell disease is caused by a single ____ substitution.

amino acid; Glu6 to Val6 on β chains of hemoglobin

the shift from Glu6 to Val6 on β chains of hemoglobin produces a ____ patch on the surface.

hydrophobic (giving sickle shape)

deoxygenated sickle cell hemoglobin (HbS) becomes ____ and forms polymers that aggregate.

insoluble

true or false: normal hemoglobin (HbA) remains soluble upon deoxygenation.

true; sickle cell hemoglobin (HbS) is insoluble when deoxygenated