Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Models for Allosteric Behavior

Monod, Wyman, Changeux (MWC) Model: allosteric proteins can exist in two states: R (relaxed) and T (taut) In this model, all the subunits of an oligomer must be in the same state T state predominates in the absence of substrate S S binds much tighter to R than to T

Biochemistry 2/e - Garrett & Grisham

More about MWC

Cooperativity is achieved because S binding increases the population of R, which increases the sites available to S Ligands such as S are positive homotropic effectors Molecules that influence the binding of something other than themselves are heterotropic effectors

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Hemoglobin

A classic example of allostery Hemoglobin and myoglobin are oxygen transport and storage proteins Compare the oxygen binding curves for hemoglobin and myoglobin Myoglobin is monomeric; hemoglobin is tetrameric Mb: 153 aa, 17,200 MW Hb: two alphas of 141 residues, 2 betas of 146

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Hemoglobin Function
Hb must bind oxygen in lungs and

release it in capillaries
When a first oxygen binds to Fe in heme of Hb, the heme Fe is drawn into the plane of the porphyrin ring This initiates a series of conformational changes that are transmitted to adjacent subunits

Biochemistry 2/e - Garrett & Grisham

Hemoglobin Function
Hb must bind oxygen in lungs and

release it in capillaries
Adjacent subunits' affinity for oxygen increases This is called positive cooperativity

Biochemistry 2/e - Garrett & Grisham

Myoglobin Structure

Mb is a monomeric heme protein Mb polypeptide "cradles" the heme group Fe in Mb is Fe2+ - ferrous iron - the form that binds oxygen Oxidation of Fe yields 3+ charge - ferric iron -metmyoglobin does not bind oxygen Oxygen binds as the sixth ligand to Fe See Figure 15.26 and discussion of CO binding

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

The Conformation Change

The secret of Mb and Hb! Oxygen binding changes the Mb conformation Without oxygen bound, Fe is out of heme plane Oxygen binding pulls the Fe into the heme plane Fe pulls its His F8 ligand along with it The F helix moves when oxygen binds Total movement of Fe is 0.029 nm - 0.29 A This change means little to Mb, but lots to Hb!

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Binding of Oxygen by Hb
The Physiological Significance
Hb must be able to bind oxygen in the lungs Hb must be able to release oxygen in capillaries If Hb behaved like Mb, very little oxygen would be released in capillaries - see Figure 15.22! The sigmoid, cooperative oxygen binding curve of Hb makes this possible!

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Oxygen Binding by Hb
A Quaternary Structure Change When deoxy-Hb crystals are exposed to oxygen, they shatter! Evidence of a structural change! One alpha-beta pair moves relative to the other by 15 degrees upon oxygen binding This massive change is induced by movement of Fe by 0.039 nm when oxygen binds See Figure 15.32

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

The Bohr Effect

Competition between oxygen and H+ Discovered by Christian Bohr Binding of protons diminishes oxygen binding Binding of oxygen diminishes proton binding Important physiological significance

See Figure 15.34

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Bohr Effect II
Carbon dioxide diminishes oxygen binding Hydration of CO2 in tissues and extremities leads to proton production These protons are taken up by Hb as oxygen dissociates The reverse occurs in the lungs

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

2,3-Bisphosphoglycerate
An Allosteric Effector of Hemoglobin In the absence of 2,3-BPG, oxygen binding to Hb follows a rectangular hyperbola! The sigmoid binding curve is only observed in the presence of 2,3-BPG Since 2,3-BPG binds at a site distant from the Fe where oxygen binds, it is called an allosteric effector

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

2,3-BPG and Hb
The "inside" story...... Where does 2,3-BPG bind? "Inside"
in the central cavity

What is special about 2,3-BPG?

Negative charges interact with 2 Lys, 4 His, 2 N-termini

Fetal Hb - lower affinity for 2,3-BPG, higher affinity for oxygen, so it can get oxygen from mother

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham

Biochemistry 2/e - Garrett & Grisham