BISC 330L Biochemistry Spring 2015 USC Lecture 10 Wednesday Feb 4 2015 Biochemistry Toolbox Synthesis of Peptides Determine protein structures 330 L Importance of Synthetic Peptides Serve as antigens for antibody production Used to isolate receptors Serve as drugs Allow to study 3 D structure of proteins Demonstrate the activity of synthetic peptide vasopression The chemistry of peptide synthesis 1 Selectivity between desired functional groups 2 Energy requirement Activation step Deprotection step Coupling step Solid Phase Peptide Synthesis Merrifield 1 Anchoring of C terminal 2 Deprotection of amino amino acid terminus 3 Coupling to next residue 4 Repeat complete peptide 5 Release peptide from resin Determine protein structure by X ray crystallography Protein crystal X ray source detector X ray wavelength bond length 1 1 5 The bottle neck is to grow protein crystals X ray source in house rotating anode synchrotron radiation Electrons scatter x rays Detector records diffracted X ray beams in direction and intensity Diffraction electron density map and structure model Phases Fourier transform 6 resolve main chain 2 8 4 side chains become visible 2 8 resolve hydrogen bonds 1 1 5 resolve atomic positions Determine protein structure by NMR in solution Chemical shifts reflects chemical environments Complex proton NMR spectrum in a protein 2D NMR nuclear Overhauser enhancement spectroscopy NOESY NOE magnetization transfer less than 5 1 r6 Mapping short proton proton distances throughout a protein Recall TCC contacting frequency map From short proton proton distances to 3D structure of the protein A population based approach to Model NMR structure A family of structures is generated In sufficient constraints NOESY derived distances are only approximate Protein molecules may vary slightly in solution This is also used in genome structure calculation using TCC data BISC 330L Reminders Next two lectures Chapter 4 material Flow of genetic information
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