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Pressure Perturbation Calorimetry

Pressure Perturbation Calorimetry. Volumetric Properties of Proteins. Protein folding. Billions of possible conformations Always (almost) form the active conformation. 1gfp.pdb. Protein Folding. Hydrophobic side chains on outside Hydrophilic side chains on inside. Hydrogen bonding.

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Pressure Perturbation Calorimetry

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  1. Pressure Perturbation Calorimetry Volumetric Properties of Proteins

  2. Protein folding • Billions of possible conformations • Always (almost) form the active conformation 1gfp.pdb

  3. Protein Folding • Hydrophobic side chains on outside • Hydrophilic side chains on inside

  4. Hydrogen bonding • Peptide hydrogen bond • Enthalpy or Entropy Driven???

  5. Salt bridges • Enhanced when buried in low dielectric constant hydrophobic interior of protein

  6. Pressure Perturbation • Apply pressure pulse to DSC(???)

  7. Property Measured • Thermal expansion coefficient • Coefficient of thermal expansion • Isobaric coefficient of thermal expansion •  = 1/V (V/ T)P • In solution includes changes in solute and solvent

  8. Structure Makers/Breakers • Hydrophobic sidechains: structure makers /T > 0 • Hydrophyllic sidechains: structure breakers /T < 0

  9. Chymotrypsinogen and Ribonulease A • Globular proteins: lots of hydrophilic sidechains • RNase and chymotrypsinogen exhibit α vs temperature behavior characteristic of structure-breakers Pressure Perturbation Calorimetry (PPC) Application Note, Microcal Corp.

  10. Chymotrypsinogen and Ribonulease A • native protein unfolding • solvent accessible surface area and α increase Pressure Perturbation Calorimetry (PPC) Application Note, Microcal Corp.

  11. References • 1. T.V. Chalikian & K.E. Breslauer (1996) Biopolymers 39, 619-626. • 2. J.F. Brandts, R. Oliveira, & C. Westort (1970) Biochemistry 9, 1038-1047. • 3. K. Sasahara & K. Nitta (1999) Protein Sci. 8, 1469-1474. • 4. S.A. Hawley (1971) Biochemistry 10, 2436-2442. • 5. H. Bull & K. Breese (1973) Biopolymers 12, 2351-2358. • 6. T.V. Chalikian, M.Totrov, R. Abagyan, & K. E. Breslauer (1996) J. Mol. Biol. 260, 588-603. • 7. D. Eisenberg & W. Kauzmann “The Structure and Properties of Water” Oxford Univ. Press, New York 1969. • 8. P. von Hippel & K-Y. Wong (1964) Science 145, 577-580. • 9. B.M. Baker & K.P. Murphy (1998) Methods Enzymology 295, 294-315. • 10. F.M. Richards in “Protein Folding (Ed. T.E.Creighton)” p. 53, Freeman, New York. • 11. R. Palma & P.M.G. Curmi (1999) Protein Sci. 8, 913-920. • 12. L-N. Lin, V. Plotnikov, J.M. Brandts, S. Williston, & J.F. Brandts, to be submitted.

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