Packing, CavitiesSurface CalculationsMolecular Surface (Connolly)Alpha-shape theoryPacking DensitySlide 6Clefts/Active SitesSlide 8Slide 9De-stabilizing mutations in coreSide-chain contact profilesProtein-protein interactionsComplementarity of P-P interfacesSlide 14Electrostatic ComplementaritySlide 16Examples of P-P interactionsbeta-sheet extensionPPI TriviaCrystal-lattice ContactsProtein-Protein DockingSlide 22Slide 23Packing, Cavities•atomic radii, contact-distance profiles•cavities•P-P interactions•crystal contacts•solvent channels•de-stabilizing mutations in core (TS mutants?)•entropy effects on surfaceSurface Calculations•Lee & Richards - solvent accessible surface–expanded atom spheres, reentrant surfaces•typical water probe radius: 1.4A•computation: grid points vs. tangents (algebraic/analytic)Molecular Surface (Connolly)Alpha-shape theory•Voronoi methods•Liang and Edelsbrunner•pockets, pockets, depressions – depends on width of openingPacking Densityfrom Richards (1977)crambin (blue=vdw,red=interstitial)data on compressibility?Jie Liang and Ken A. Dill (BiophysJ, 2001). Are Proteins Well-Packed? •636 proteins; 1.4A probe radius •proteins are dense (like solids), yet atoms are arranged like liquids (without voids)•P=0.76 for hex-packed spheres•P=0.74 for protein interiors•distribution of number/size of voids is more variable, like a liquid•surface area scales linearly with volume, instead of A V-2/3Clefts/Active Sites•Liang Edelsbrunner, Woodward (1998)•Laskowski, Luscombe, Swindells, and Thornton (1996)De-stabilizing mutations in core•cavities, tolerance, re-packing•Serrano L., Kellis J., Cann P., Matouschek A. & Fersht A. (1992)•In barnase, 15 mutants were constructed in which a hydrophobic interaction was deleted •strong correlation between the degree of destabilization (which ranges from 0.60 to 4.71 kcal/mol) and the number of methylene groups deleted •average free energy decrease for removal of a completely buried methylene group was found to be 1.5±0.6 kcal/mol. This is additive. •double-mutants?•temperature-sensitive mutants?Side-chain contact profiles•Sippl – knowledge-based potentials •Subramaniam – PDF’s•dependence: radial distance, sequence separationProtein-protein interactions•flat and hydrophobic? •Janin•Jones and Thornton (1996), PNAS – data on flatness, H-bonds•which is predominant: H-bonds vs. salt-bridges vs. hydrophobic interactions?(homodimers)Complementarity of P-P interfaces•shape complementarity–measure “gaps” or voids–cavities at interfaces (Hubbard and Argos, 1994)•more common than in core•suggests complementarity doesn’t have to be perfect–surface normals:•R Norel, SL Lin, HL Wolfson and R Nussinov•LoConte, Chothia, and Janin (1999), JMB.–The average interface has approximately the same non-polar character as the protein surface as a whole, and carries somewhat fewer charged groups. –However, some interfaces are significantly more polar and others more non-polar than the average.–1/3 of interface atoms becomes completely buried; packing density is similar to core (like organic solids)–in high-res structures, remainder of space is filled in by water molecules (making H-bonds)–size for “typical” interfaces: 1600±400Å2Electrostatic Complementarity•McCoy et al. (1997)–defined two correlation coefficients between surfaces (summed over contacts): charge complementarity (ion pairs), and electrostatic potentials–depends on assignment of partial charges, solvation...–examine effect on G–charge correlations: -0.1..+0.1 (insignificant)–electrostatic potential correlations: 0.1..0.7 (significant)• steering and diffusion (Kozak et al., 1995)Jones and Thornton – Patch Analysis,PP-interface predictionExamples of P-P interactions• -lactamase/BLIP – one of the tightest•antibody-antigens (HYHel5)•SH2/SH3 and tyrosine kinases•PDZ domains•calmodulin•proteases, kinases (recognize+catalyze)beta-sheet extension•arylamine N-acetyltransferase (nat)–acetylates isoniazid in M.smegmatis–pdb: 1W6F–active in solution as both monomer and dimer–lower surface area, but many H-bondsPPI Trivia•obligate vs. transient complexes - affinity•differences between antigen-antibody, protease-inhibitor, and rest of complexes•induced conformational changes•allostery •evolutionary conservation at interfaces (Caffrey et al. 2004), •correlated mutations? mutational hot spots, evolutionary trace (Lichtarge)•why are homodimers so common? (Lukatsky et al, 2007)succinyl-CoA synthetasegreen=contactpurple=conservedCrystal-lattice Contacts•Carugo and Argos (1997)•small: 45%<100Å2, 8%>500Å2 •properties like rest of surface, so probably random•induced changes (rms)?Protein-Protein Docking•FTDOCK - Gabb, Jackson, and Sternberg (1997)–use Fourier transform to evaluate shape correlation function–correlation function includes shape and electrostatic complementarity of surfaces–try 6912 rotations, =15º=-15grid nodes within 1.8A ofprotein atom are “inside”•Multidock (Jackson, Gabb, Sternberg, 1998)–what about induced fit? alternative side-chain rotamers? domain rotations? –need refinement to do scoring of complexes, increase sensitivity to recognize correct interaction–add term for solvation energy (soft-sphere Langevin interactions between solvent grid points and surface side-chains)–sample different rotamers–Betts & Sternberg (1999) – induced fit at interfaces side-chain and backbone movements•GRAMM (Vakser) –low resolution protein docking–maybe removing details will help...–search 6D space for maximal surface overlap (20º rotations)–intermolecular overlap function–evaluated on a coarse 7Å gridPatchDock, FireDock (Nussinov &
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