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Protein Function

Protein Function. Structure will determine the function of the protein. Key ideas and terms. protein can bind a ligand in the binding site For an enzyme, the ligand is a substrate and they bind in what is called the active site ligand has to be the correct shape

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Protein Function

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  1. Protein Function Structure will determine the function of the protein

  2. Key ideas and terms • protein can bind a ligand in the binding site • For an enzyme, the ligand is a substrate and they bind in what is called the active site • ligand has to be the correct shape • ligand has to have the complementary charges and hydrophobicity or hydrophilicity

  3. Lock and Key Hypothesis • Protein and ligand have complementary shapes. • Interactions must also be complementary • If enzyme charge is negative, substrate must be positive • If pocket is nonpolar, ligand must be nonpolar • Antibodies

  4. Induced Fit • Induced Fit: when the protein and ligand bind, the protein may change conformation to allow for tighter binding • Frequently, both the ligand and the protein change conformation

  5. Examples: O2 binding proteins: myoglobin and hemoglobin • oxygen is not water soluble yet needs to be transported • diffusion is not effective • myoglobin is found primarily in muscle tissue • Hemoglobin is in the blood • Both proteins contain heme

  6. Heme Group consists of a Fe2+ and a protoporphyrin ring to help stabilize the iron(II) ion

  7. Heme Group the iron must be a 2+ to bind oxygen. The heme group is buried deep within the protein so that the iron is not oxidized to 3+ • there must be flexibility in the protein to allow for oxygen to attach and then let go • Iron has 6 coordination sites. • Four of them used by porphyrin. Unshared pairs on nitrogen complex to iron • Fifth and 6th for oxygen and protein • Heme is planar

  8. Myoglobin • has heme group • eight alpha helical segments • dense hydrophobic core • all but two polar groups on outside • room for only 4 water molecules • flat heme in pocket • iron coordinated to poryphorin and H • As well as the heme

  9. Myoglobin Binding Curve • Hyperbolic binding curve • Relatively insensitive to small changes in oxygen concentration

  10. Myoglobin • The P50 (oxygen partial pressure required for half saturation) for myoglobin is very low • Myoglobin has a high affinity for oxygen-an important characteristic for a protein that must extract oxygen from the small amounts present in blood. • At the oxygen concentration existing in the capillaries, the myoglobin in adjacent tissues is nearly saturated. • When cells are metabolically active, their internal PO2 falls to levels where myoglobin will lose(deliver) its oxygen.

  11. Hemoglobin Quaternary structure : 4 subunits • Each subunits is like myoglobin • Each subunit has heme group • 2 alpha chains; 2 beta chains • Few contacts between alpha and betas, more between alphas and betas

  12. Hemoglobin • Exists in two states • R state (high affinity for O2) • Where would this state be favored? • In the lungs • T state (low affinity for O2) (deoxyhemoglobin) • Where would this state be favored? • In the tissue • Sensitive to pressure changes • On oxygenation, one pair of subunits shifts with respect to the other by a rotation of 15 degrees.

  13. Hemoglobin • deoxy hemoglobin (T) oxy hemoglobin (R)

  14. Oxygen Binding to Heme in Hemoglobin • Fe is coordinated to a histidine in helix 8 of the Hb molecule • In deoxy form, porphyrin is puckered and Fe is out of the plane of the heme • When oxygen binds the Fe (at other coordination site) the Fe is pulled into the plane of the heme • This pulls on the histidine, which pulls on the helix, changing the shape of the molecule. His F8 Fe2+ 0.6 A O2

  15. Hemoglobin • Conformational changes in hemoglobin alter its binding ability • Binding of oxygen in one subunit causes conformational changes in the next subunit • This is called cooperative binding • This can happen because it is composed of 4 independent subunits • produces a different binding curve that is sigmoidal • The modulation of the affinity of a site for a ligand by ligand binding at another site is called Allostery.

  16. Hemoglobin Binding Curve

  17. Bohr Effect • Hemoglobin's affinity for oxygen is decreased in the presence of carbon dioxide and at lower pH. • Carbon dioxide reacts with water to give bicarbonate, carbonic acid free protons via the reaction: CO2 + H2O ---> H2CO3 ---> H+ + HCO3- • Protons bind at various places along the protein and carbon dioxide binds at the alpha-amino group forming carbamate. • This causes a conformational change in the protein and facilitates the release of oxygen.

  18. Bohr Effect • Blood with high carbon dioxide levels is also lower in pH (more acidic). (recall the equilibrium) • Conversely, when the carbon dioxide levels in the blood decrease (i.e. around the lungs), carbon dioxide is released, increasing the oxygen affinity of the protein.

  19. High CO2 in tissues Higher H+ Lower pH Affinity for O2 decreases O2 released to tissues T state favored Low CO2 in lungs Lower H+ Higher pH Affinity for O2 increases O2 binds hemoglobin R state favored Bohr Effect Summary

  20. Hemoglobin and CO poisoning • Other ligands can compete with oxygen for binding to hemoglobin • The binding of oxygen is affected by molecules such as carbon monoxide (CO) (For example from tobacco smoking, cars and furnaces). CO competes with oxygen at the heme binding site. • Hemoglobin binding affinity for CO is 200 times greater than its affinity for oxygen, meaning that small amount of CO can dramatically reduces hemoglobin’s ability to transport oxygen. • Hemoglobin also has competitive binding affinity for Nitrogen Dioxide and Hydrogen sulfide .

  21. Oxygen and Carbon Monoxide • Oxygen and carbon monoxide same size and shape. • Carbon monoxide, however has formal charge • Sticks to Fe better • Blocks oxygen binding

  22. Hemoglobin and 2,3 DPG • In people acclimated to high altitudes, the concentration of 2,3-diphosphoglycerate (2,3-DPG) in the blood is increased, which allows these individuals to deliver a larger amount of oxygen to tissues under conditions of lower oxygen tension. • This phenomenon, where molecule Y affects the binding of molecule X to a transport molecule Z, is called a heterotropic allosteric effect.

  23. Sickle Cell Anemia • Sickle cell disease is caused by an abnormal adult hemoglobin, called hemoglobin S. People with sickle cell disease make hemoglobin S instead of hemoglobin A. • Single amino acid substitution • glutamate changed to valine • To show condition, have to have mutation in both genes (Homozygous) • http://www.scinfo.org/sicklept.htm

  24. Sickle vs normal hemoglobin • first 9 amino acids of normal hemoglobin beta chain v h l t p e e k s • first 9 amino acids of sickle hemoglobin beta chain v h l t p v e k s Notice the single amino acid change?

  25. Sickle Cell Anemia • Position 6 is on outside of molecule • Glutamate is polar • Valine substitution causes “sticky spot” on outside of hemoglobin • Causes hemoglobin molecules to stick together • Forms long chains which cause red blood cell to sickle

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