Hydrogen bonding and biological specificity analysed by protein engineering



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BP 204 Focus Paper: Ligand Binding - Stroud:
P1) Fersht AR; Shi JP; Knill-Jones J; Lowe DM; Wilkinson AJ; Blow DM; et al. Hydrogen bonding and biological specificity analysed by protein engineering. Nature, 1985 Mar 21-27, 314(6008):235-8.
• Energetic accounting for a hydrogen bond

• Paired-to-unpaired hydrogen bonds to charged moieties

• Balancing the equation of hydrogen bonds versus solvent

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P2) Livnah O; Stura EA; Johnson DL; Middleton SA; Mulcahy LS; Wrighton NC; Dower WJ; Jolliffe LK; Wilson IA. Functional mimicry of a protein hormone by a peptide agonist: the EPO receptor complex at 2.8 A Science, 1996 Jul 26, 273(5274):464-71.
• Biology of selection: ‘phage display’; why dimerizing ligands selected?

• Avidity. Additivity of free energy contributors

• Buried surface areas and affinity

• Transmembrane signaling by cytokine receptors depends on dimerization

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P3) Robert C. Rizzo, De-Ping Wang, Julian Tirado-Rives, and

William L. Jorgensen Validation of a Model for the Complex of HIV-1

Reverse Transcriptase with Sustiva through Computation of Resistance Profiles

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P4) David E. Shaw,1,2* Paul Maragakis,1† Kresten Lindorff-Larsen,1† Stefano Piana,1† Ron O. Dror,1 Michael P. Eastwood,1 Joseph A. Bank,1 John M. Jumper,1 John K. Salmon,1 Yibing Shan,1 Willy Wriggers Atomic-Level Characterization of the Structural Dynamics of Proteins

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P5) Chimera tutorial: http://bit.ly/LBQr04

(http://www.cgl.ucsf.edu/chimera/data/tutorials/bp204/classdata.html)


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P6) Zhou, Y; Morals-Cabral, JH; Kaufman, A., MacKinnon, R. Chemistry of ion coordination and hydration revealed by a K+ channel-Fab complex at 2.0 Å resolution. Nature, 2001 Nov 01, (414):43-48.
• Basis for coordinating K+ ions – the pathway

• Compensation for lipids in the center

• Selectivity for K+ versus other ions

• Hydrophobic exit port

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P7). Erlanson,DA; Braisted, AC; Raphael, DR; Randal, Mike; Stroud,RM; Gordon, EM; Wells, JA. Site-directed ligand discovery. PNAS. 2000 Aug 15, 97(17)9367-9372.
• Facing problems for Drug Discovery

• Tethering – Why? How? Reducing potential

• Additivity of energy
Jencks WP. On the attribution and additivity of binding energies. Proc Natl Acad Sci U S A. 1981;78(7):4046-50.
• The conundrum of non-additivity

• Fragmenting Biotin

• Entropy reduction
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P8) Rasmussen SG, DeVree BT, Zou Y, Kruse AC, Chung KY, Kobilka TS, et al. Crystal structure of the beta2 adrenergic receptor-Gs protein complex. Nature. 2011;477(7366):549-55.

• Heterotrimeric G-protein complex

• Llama antibodies/nanobodies cubic lipidic phases

• Crystallogenesis

• Activation, interaction surface


Chun E, Thompson AA, Liu W, Roth CB, Griffith MT, Katritch V, et al. Fusion partner toolchest for the stabilization and crystallization of G protein-coupled receptors. Structure. 2012;20(6):967-76.
• Protein engineering for stability

• Size exclusion chromatography and flexibility

• Protein diffusion and crystallization

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Focus Papers: Enzymes and Catalysis: Miller/Gross

P13) Milburn MV, Tong L, deVos AM, Brunger A, Yamaizumi Z, Nishimura S, et al. Molecular switch for signal transduction: structural differences between active and inactive forms of protooncogenic ras proteins. Science. 1990;247


Kraulis PJ, Domaille PJ, Campbell-Burk SL, Van Aken T, Laue ED. Solution structure and dynamics of ras p21.GDP determined by heteronuclear three- and four-dimensional NMR spectroscopy. Biochemistry. 1994

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P14) Worthylake DK, Rossman KL, Sondek J. Crystal structure of Rac1 in complex with the guanine nucleotide exchange region of Tiam1. Nature. 2000;408
Aghazadeh B, Lowry WE, Huang XY, Rosen MK. Structural basis for relief of autoinhibition of the Dbl homology domain of proto-oncogene Vav by tyrosine phosphorylation. Cell. 2000;102(5):625-33.

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P15) Li P, Martins IR, Amarasinghe GK, Rosen MK. Internal dynamics control activation and activity of the autoinhibited Vav DH domain. Nat Struct Mol Biol. 2008;15(6):613-8.

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END OF COHERENT

Interaction: The C-Terminal Domain of GAP IS Not Sufficient for Full Activity

1) Kraut, J. [1988] How do enzymes work? Science 242, 533-540.
• Transition state theory

Basic kinetics

• Catalysis

• Enzyme kinetics

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2) Fastrez, J. & Fersht, A.R. [1973] Demonstration of the acyl-enzyme mechanism for the hydrolysis of peptides and anilides by chymotrypsin. Biochemistry 12, 2025-2034.
Serine proteases

• Covalent intermediates

• Multistep processing

• Free energy vs. reaction coordinate

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3) Abrahmsen, L., et al. [1991] Engineering subtilisin and its substrates for efficient ligation of peptide bonds in aqueous solution. Biochemistry 30, 4151-4159.
• Changing the properties of enzymes

• Microscopic reversibility

• Chemical reactivity of active site groups

• Additional examples of tailored enzymes

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4) Raines, R.T., et al. [1986] Reaction energetics of a mutant triosephosphate isomerase in which the active-site glutamate has been changed to an aspartate. Biochemistry 25, 7142-7154.
• Evolutionary perfection of enzyme catalysis

• Concerted reactions

• Diffusion control

Isotope effects

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5) Wells, T.N.C. & Fersht, A.R. [1986] Use of binding energy in catalysis analyzed by mutagenesis of the tyrosyl-tRNA synthetase. Biochemistry 25, 1881-1886.
• Specificity

• Limits of fidelity

• Proofreading mechanisms

• Binding energy in catalysis

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1) Kraut, J. [1988] How do enzymes work? Science 242, 533-540.


• Transition state theory

• Basic kinetics

• Catalysis

• Enzyme kinetics

_____________________________________________________________________________
2) Fastrez, J. & Fersht, A.R. [1973] Demonstration of the acyl-enzyme mechanism for the hydrolysis of peptides and anilides by chymotrypsin. Biochemistry 12, 2025-2034.
• Serine proteases

• Covalent intermediates

• Multistep processing

• Free energy vs. reaction coordinate

_____________________________________________________________________________
3) Abrahmsen, L., et al. [1991] Engineering subtilisin and its substrates for efficient ligation of peptide bonds in aqueous solution. Biochemistry 30, 4151-4159.
• Changing the properties of enzymes

• Microscopic reversibility

• Chemical reactivity of active site groups

• Additional examples of tailored enzymes

_____________________________________________________________________________
4) Raines, R.T., et al. [1986] Reaction energetics of a mutant triosephosphate isomerase in which the active-site glutamate has been changed to an aspartate. Biochemistry 25, 7142-7154.
• Evolutionary perfection of enzyme catalysis

• Concerted reactions

• Diffusion control

• Isotope effects

_____________________________________________________________________________
5) Wells, T.N.C. & Fersht, A.R. [1986] Use of binding energy in catalysis analyzed by mutagenesis of the tyrosyl-tRNA synthetase. Biochemistry 25, 1881-1886.
• Specificity

• Limits of fidelity

• Proofreading mechanisms

• Binding energy in catalysis

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Focus Papers for Protein Folding: Agard
1) Eriksson AE; Baase WA; Zhang XJ; Heinz DW; Blaber M; Baldwin EP; Matthews BW. Response of a protein structure to cavity-creating mutations and its relation to the hydrophobic effect. Science 255:178-183. (1992)
• Quantitative measurement of contribution of hydrophobic effect to protein stability

• Deletions result in cavities within the protein that compact to differing degrees

• Energetics of cavity formation comparable to hydrophobic effect

• Rationalizes energetic consequences of side chain mutations-after contradictions

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2) Riddle DS, Santiago JV, Bray-Hall ST, Doshi N, Grantcharova VP, Yi Q, Baker D Functional rapidly folding proteins from simplified amino acid sequences Nature Struct Biol 4:805-809 (1997)
• How complex a sequence “alphabet” is required to code a stable protein fold?

• Is the folding rate of small protein domains optimized through evolution?

• Use of phage display for simplifying proteins

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3) Hughson,F.M., Wright, P.E., Baldwin, R.L. Structural characterization of a partly folded apomyoglobin intermediate Science 249:1544-1548 (1990).
• Molten globules are thought to be critical intermediates along folding pathways

Structure of a molten globule

• Use of hydrogen exchange to study properties of folding reactions

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4) Chamberlain AK; Handel TM; Marqusee S. Detection of rare partially folded molecules in equilibrium with the native conformation of RNaseH Nature Structure Biology 3:782-7 (1996).
• Rare conformational states are accessible from the native state

• Correlation between relative stability and folding pathways

• Domains in a structure have different stabilities

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5) Netzer WJ, Hartl FU. Recombination of protein domains facilitated by co-translational folding in eukaryotes Nature 388:343-349 (1997)
• Comparison; protein folding machinery in prokaryotes and eukaryotes

• Fundamental differences may be adapted for folding of different kinds of proteins

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Focus Papers: Protein-Protein Interactions: Fletterick


1) Schuster SC; Swanson RV; Alex LA; Bourret RB; Simon MI. Assembly and function of a quaternary signal transduction complex monitored by surface plasmon resonance. 1993 Nature, 365(6444):343-7.
• Measuring association and dissociation of proteins

• Role of ATP driven phosphorylation and covalent modification in complex stability

• Role of ligand binding to receptor in promoting assembly

• SPR to derive binding constants

• Quaternary signal transduction complex controls chemotaxis

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2) Horton N; Lewis M. Calculation of the free energy of association for protein complexes. (1992) Protein Science 1(1):169-81.
• Thermodynamics of Protein Assembly

• Structural Change on complexation

• Empirical fitting of Atomic Interactions with Free Energy of Association

• Estimate of free energy of H bonds and charge interactions in protein complexes and role of hydrophobic effect

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3) Chothia C; Lesk AM; Tramontano A; Levitt M; Smith-Gill SJ; Air G; Sheriff S; Padlan EA; Davies D; Tulip WR; et al. Conformations of immunoglobulin hypervariable regions. Nature, 1989 Dec 21-28, 342(6252):877-83.
• Definition of IgG fold

• Definition of CDR’s and their conformations

• Target sites on antigens and Fab’s

• Structural changes, characterization of interfaces

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4) Clackson T; Ultsch MH; Wells JA; de Vos AM. Structural and functional analysis of the 1:1 growth hormone:receptor complex reveals the molecular basis for receptor affinity. Journal of Molecular Biology, 1998 Apr 17, 277(5):1111-28.
• Structure of Growth hormone with its receptor

• Mutagenesis and Affinity define important interfaces

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  1. Russo AA; Jeffrey PD; Patten AK; Massague J; Pavletich NP. Crystal structure of the p27Kip1 cyclin-dependent-kinase inhibitor bound to the cyclin A-Cdk2 complex. Nature, 1996 Jul 25, 382(6589):325-31.

• Multiple interactions build a three-protein complex

• Protein mimic of ATP

• Changes in protein structure on complex formation

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Focus Papers: Nucleic acid-Protein Interactions: Frankel


1) Weeks, K. M. and Crothers, D. M. Major groove accessibility of RNA. 1993 Science 261, 1574-1577.
• The major groove provides a prime recognition surface in nucleic acids

• RNA and DNA structures are very different

• Discontinuities in RNA helices make virtually all base pairs available for recognition

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2) Murray, J. B., Terwey, D. P., Maloney, L., Karpeisky, A., Usman, N., Beigelman, L., and Scott, W. G. The structural basis of hammerhead ribozyme self-cleavage. 1998 Cell 92, 665-673.
• RNAs adopt complex folds

• RNAs can perform chemical reactions

• Metal ions are important for structure and catalysis

• RNAs can undergo major conformational change

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3) Sclavi, B., Sullivan, M., Chance, M. R., Brenowitz, M., and Woodson, S. A. RNA folding at millisecond intervals by synchrotron hydroxyl radical footprinting. 1998 Science 279, 1940-1943.
• RNA folding is ordered but does not necessarily follow a single pathway

• Secondary structures (helices) assemble into higher order structures

• Kinetic traps are possible

• Folding time scales are similar to proteins

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4) Rastinejad, F., Perlmann, T., Evans, R.M., and Sigler, P.B. Structural determinants of nuclear receptor assembly on DNA direct repeats. 1995 Nature 375, 203-211.
• DNA-binding proteins often share common structural motifs

• The major groove, minor groove, and backbone provide specific recognition points

• Water molecules often are located at protein-nucleic acid interfaces

• Oligomeric arrangements can generate different specificities

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5) Price, S. R., Evans, P. E., and Nagai, K. Crystal structure of the spliceosomal U2B"-U2A' protein complex bound to a fragment of U2 small nuclear RNA. 1998 Nature 394, 645-650.
• RNA loops provide important recognition features

• Both RNA and protein often show induced fit upon binding

• Recognition surfaces can be remodeled to generate different specificities

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Epub 2000/09/28. PubMed PMID: 11007481.


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