lecture5_9_16_08 - Lecture 5 Chapter 7: Stryer Overview:...

Info iconThis preview shows page 1. Sign up to view the full content.

View Full Document Right Arrow Icon
This is the end of the preview. Sign up to access the rest of the document.

Unformatted text preview: Lecture 5 Chapter 7: Stryer Overview: myoglobin & hemoglobin structure/function allostery cooperativity BPG regulation Hill plots Homework: 7.4, 7.6, 7.8, 7.9 Themes in protein function • structure dictates function • proteins are often flexible & dynamic • protein activity/function requires interactions with other molecules protein-ligand ligand = metal ion, small molecule, protein, DNA, RNA, etc. binding site structure dictates function • conformational changes in one or both of the interacting molecules induced fit allostery cooperativity Oxygen-binding proteins myoglobin (Mb) in muscle - oxygen storage monomeric 153 aa, 16.7 kDa hemoglobin (Hb) in blood - oxygen transport tetrameric (!2"2), 64.5 kDa 2 ! subunits, 141 aa each 2 " subunits, 146 aa Why do we need these proteins? What are the roles of heme and Fe? How do they bind/release oxygen? Oxygen-binding proteins contain heme protoporphyrin IX 4 pyrrole rings Oxygen-binding proteins contain heme Fe2+ Oxygen (O2) binding to myoglobin (Mb) general binding curve binding curve for Mb-O2 Kd = [L] at which 50% of binding sites are full fraction of binding sites P-L interaction Kd = myoglobin - 8 helical chains [P][L] [PL] Ka = [PL] [P][L] [L] measured as partial pressure low Kd, high affinity high Kd, low (poor) affinity PL P+L dissociation/association constants units? Ligand Kd values units - molarity (M) or inverse molarity (M-1) 1 # 10-1 1 # 10-2 1 # 10-3 1 # 10-4 1 # 10-5 1 # 10-6 1 # 10-7 1 # 10-8 1 # 10-9 1 # 10-10 1 # 10-11 1 # 10-12 1 # 10-15 0.1 M 0.01 M 0.001 M 0.0001 M 0.00001 M 0.000001 M 0.0000001 M 0.00000001 M 0.000000001 M 0.0000000001 M 0.00000000001 M 0.000000000001 M 0.000000000000001 M 100 mM weak 10 mM 1 mM 100 µM 10 µM 1 µM good binding 100 nM 10 nM strong binding 1 nM 100 pM 10 pM 1 pM 1 fM O2 vs. CO binding to heme O2 binding to heme in class tutorial Role of distal amino acid residues in O2 binding Hemoglobin (Hb) and O2 • Hb binds oxygen in the lungs and releases it in the capillaries • Hb without O2 is called deoxyhemoglobin • first O2 binds and initiates a conformational change that is transmitted to adjacent subunits • adjacent subunits’ affinity for O2 increases • example of positive cooperativity • Hb tetramer has two conformational states - T state (‘tense’), low O2 affinity - R state (‘relaxed’), high O2 affinity - binding O2 changes the protein from T -> R state Hemoglobin (Hb) and O2 Hemoglobin (Hb) vs. Myoglobin (Mb) Limited sequence similarity, but similar structure Hemoglobin (Hb) vs. Myoglobin (Mb) Deoxyhemoglobin interaction at interface between unlike subunits ion pairs stabilize T state Limited sequence similarity Hemoglobin (Hb) and O2 Hemoglobin (Hb) and O2 Note: !2"2 dimer shifts relative to !1" 1 dimer see tutorial compare curves of Hb and Mb with O2; note differences in P50 value sigmoidal curvation is indicative of cooperativity What is the advantage to Hb? Think about the function of Hb compared to Mb! Structural basis for ligand-binding cooperativity in a multisubunit protein Hemoglobin (Hb) and O2 cooperativity: what is the benefit? consider response of Hb/dioxygen binding during exercise Hemoglobin (Hb) and O2 Hemoglobin (Hb) and O2 binding models Hemoglobin (Hb) and O2 Hemoglobin (Hb) and O2 Hemoglobin (Hb) and BPG BPG (2,3-bisphosphoglycerate) regulation of hemoglobin binding to O2 Hemoglobin (Hb) and BPG T State BPG reduces the affinity of Hb for O2 (one BPG binds each Hb tetramer & stabilizes the low O2 affinity T state) R State see tutorial Hemoglobin (Hb) and BPG Hemoglobin (Hb) and BPG How is fetal Hb different from maternal Hb? What is the significance? Hemoglobin (Hb) and CO2 Hb can bind H+ and CO2 and serves to carry them from the tissues where they are generated to the lungs and kidneys where they are excreted At low pH (high [H+ ])and high [CO2], Hb has reduced affinity for O2 O2 and H+ have inverse binding affinities for Hb Bohr effect Chemical basis for the Bohr effect In deoxyHb, three amino acid residues (Lys, His, and Asp) form two salt bridges that stabilize the T quaternary structure. Think about pH dependence of the bridges. Carbon dioxide and pH Carbon dioxide and pH What is the role of carbonic anhydrase (CH. 9)? Hemoglobin (Hb) and sickle-cell anemia Sickle-cell anemia results from a single mutation in the normal Hb protein! Glu -> Val at position 6 in ! chain Val creates a hydrophobic ‘sticky’ patch on Hb, causing proteins to associate abnormally, leading to long, fibrous aggregates this gives the erythrocyte a ‘sickled’ appearance, leading to capillary blockage Hemoglobin (Hb) and sickle-cell anemia Hemoglobin (Hb) and sickle-cell anemia Hemoglobin (Hb) and sickle-cell anemia What is the relationship of sickle-cell anemia to malaria? leads to aggregation Hemoglobin (Hb) and !-thalassemia Binding models See pp. 199-200 in your textbook for equation derivations The “Hill” plot AHSP: "-hemoglobin stabilizing protein ...
View Full Document

This note was uploaded on 08/04/2010 for the course CHM 6620 taught by Professor Dr.christinechow during the Fall '08 term at Wayne State University.

Ask a homework question - tutors are online