Kyle’s Section BIBC100 – Structural [email protected] Summer 2005 Week 4 OutlineI.Hemoglobin / MyoglobinA. Hill Plotsi. T-state, allosteric change, R-stateii. Hill coefficient (cooperativity index); n = 3 for hemoglobiniii. n > 1 => positive cooperativityiv. 0 < n < 1 => negative cooperativityB. Allosteric Regulationi. Hb vs. Stripped Hbii. Effects of CO2, Biphosphoglycerate, and pH on hemoglobina. CO2 reduces binding affinity of O2b. BPG also reduces binding affinityc. Drops in pH ( increasing acidity) decrease binding affinitya. “The Bohr Effect”II. MetalloproteinsA. Carbonic Anhydrasea. H2O + CO2 => HCO3- + H+b. Zn at active site, coordinated by histidine residuesB. Calmodulina. Regulated by changes in calcium levelsb. Activate Ca/CaM complex can further activate or inactivate other proteins and enzymesc. Helix – turn – helix binding motif for calcium (EF Hand)d. Physiological range of [Ca2+] = 0.2-0.5i. This is well below saturation levelsIII. Ion ChannelsA. Porina. Beta-Barrel structureb. Porin monomers & porin trimersc. Tyrosine – often found at plasma membrane aqueous solution interfaced. E-Coli – porins allow disaccharides like maltose to pass throughB. Nicotinic Acetylcholine Receptor ( nAChR)a. A ligand gated ion channel found at the neuromuscular junction and other neuronal synapsesb. Activation, or the opening of the channel, requires 2 acetylcholine molecules to be bound simultaneouslyi. Allows for Na+ to flow in and depolarize the cellc. Pentameric Structure – alpha2, beta, gamma, deltai. Each monomer has 4 transmembrane spanning alpha-helicesii. M1, M2, M3, M4iii. This pentamer forms a circular structureiv. Acetylcholine binds to the alpha subunitv. Topology Diagram – N-terminal and C-terminal are both located extracellularly.vi. M2 helix contains Leu residues that block the channel when inactivatedvii. M2 helix turns when ligand binds, and rotates Ser residues towards the center of the channelviii. Hydropathy PlotC. K-channel a. Channels like this must remove the hydration shell around ionsb. Voltage gated – S4 helix has + charged residues which respond to changes in current flowc. Non-voltage gated – only S1 and S2 helicesd. Pore Loop is conserved in both structurese. Structurei. Often tetramericii. Selectivity filter, central cavity, central poreiii. 2 alpha helices oriented with ∂- dipole end pointing towards thecentral cavityiv. Seven main sites for ions to pass through the channelf. Na+ is only bound to 2 of the 4 possible C=O carbonyl groups, and thus is energetically unfavorable enough to not allow the ion to pass through the K+ channel, in-spite of its smaller sizei. K+ is large enough to interact with all 4 carbonylsIV. Molecular Motion of MacromoleculesA. Time Scalesa. Short - bond stretching, angle bendingb. Medium - surface side chain motion, loop motionc. Long - folding in small peptides, helix coil transitiond. Very Long - protein foldingB. Degrees of Freedoma. A 4-carbon backbone will have 6 degrees of freedomi. 3 for bond stretchingii. 2 for bond angle bendingiii. 1 for the torsion angleC. Gramicidin Aa. Dimmerb. N-terminals meet at the center of the membraneD. Peptide Self-assembly - * understand this slide *E. Protein Foldinga. Nucleation, Condensation, Molten Globule, Tertiary Fold, Quaternary Fold, Global Energy MinimumF. Folding Funnels – changes in the amount of entropyG. Chaotropic Agents – chemicals that induce disorder or unfolding in proteinsa. Ureab. Guanidinium HClH. Reducing Agents – these break disulfide bridges found in tertiary protein structurea. B-mercaptoethanolb. Dithiothreitol (DTT)I. p53 - tumor suppressor gene which is inactivated when mutateda. mutated proteins are more susceptible to chaotropic agents like
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