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Thermodynamic signatures in macromolecular interactions involving conformational flexibility

  • Anja Menzel , Piotr Neumann , Christian Schwieger und Milton T. Stubbs EMAIL logo
Veröffentlicht/Copyright: 8. Juli 2014
Biological Chemistry
Aus der Zeitschrift Biological Chemistry Band 395 Heft 7-8

Abstract

The energetics of macromolecular interactions are complex, particularly where protein flexibility is involved. Exploiting serendipitous differences in the plasticity of a series of closely related trypsin variants, we analyzed the enthalpic and entropic contributions accompanying interaction with L45K-eglin C. Binding of the four variants show significant differences in released heat, although the affinities vary little, in accordance with the principle of enthalpy-entropy compensation. Binding of the most disordered variant is almost entirely enthalpically driven, with practically no entropy change. As structures of the complexes reveal negligible differences in protein-inhibitor contacts, we conclude that solvent effects contribute significantly to binding affinities.

Keywords: crystal structure; enthalpy entropy compensation; isothermal titration calorimetry; protein flexibility; thermodynamics

Dedicated to Professor Dr. Gerhard Klebe on the occasion of his 60th birthday.

Corresponding author: Milton T. Stubbs, Institut für Biochemie und Biotechnologie, Martin-Luther-Universität Halle-Wittenberg, Kurt-Mothes-Straße 3, D-06120 Halle/Saale, Germany, e-mail:
aPresent address: Institut für Rechtsmedizin, Otto-von-Guerike-Universität, Leipziger Straße 44, D-39120 Magdeburg, GermanybPresent address: Institut für Mikrobiologie und Genetik, Georg-August-Universität, Justus-von-Liebig-Weg 11, D-37077 GÖttingen, Germany

Acknowledgments

This work was supported by the DFG Graduiertenkolleg 1026 ‘Conformational transitions in macromolecular interactions’.

  1. 1

    The disorder is reflected in higher temperature factors of the final model in this region; a more detailed analysis of these has not been carried out because the link between crystallographic B factors and dynamics is questionable, demonstrated recently for a set of representative proteins (Reichert et al., 2012)

References

Bode, W. and Huber, R. (1991). Proteinase-protein inhibitor interaction. Biomed. Biochim Acta 50, 437–446.10.1007/978-3-642-76412-7_9Suche in Google Scholar

Bolognesi, M., Pugliese, L., Gatti, G., Frigerio, F., Coda, A., Antolini, L., Schnebli, H.P., Menegatti, E., Amiconi, G., and Ascenzi, P. (1990). X-ray crystal structure of the bovine alpha-chymotrypsin/eglin c complex at 2.6 Å resolution. J. Mol. Recognit. 3, 163–168.10.1002/jmr.300030405Suche in Google Scholar

Breiten, B., Lockett, M.R., Sherman, W., Fujita, S., Al-Sayah, M., Lange, H., Bowers, C.M., Heroux, A., Krilov, G., and Whitesides, G.M. (2013). Water networks contribute to enthalpy/entropy compensation in protein-ligand binding. J. Am. Chem. Soc. 135, 15579–15584.10.1021/ja4075776Suche in Google Scholar

Burgering, M.J.M., Orbons, L.P.M., vanderDoelen, A., Mulders, J., Theunissen, H.J.M., Grootenhuis, P.D.J., Bode, W., Huber, R., and Stubbs, M.T. (1997). The second Kunitz domain of human tissue factor pathway inhibitor: cloning, structure determination and interaction with factor Xa. J. Mol. Biol. 269, 395–407.10.1006/jmbi.1997.1029Suche in Google Scholar

Chen, Z. and Bode, W. (1983). Refined 2.5 Å X-ray crystal structure of the complex formed by porcine kallikrein A and the bovine pancreatic trypsin inhibitor. Crystallization, Patterson search, structure determination, refinement, structure and comparison with its components and with the bovine trypsin-pancreatic trypsin inhibitor complex. J. Mol. Biol. 164, 283–311.10.1016/0022-2836(83)90078-5Suche in Google Scholar

Cho, S., Swaminathan, C.P., Bonsor, D.A., Kerzic, M.C., Guan, R., Yang, J., Kieke, M.C., Andersen, P.S., Kranz, D.M., Mariuzza, R.A., et al. (2010). Assessing energetic contributions to binding from a disordered region in a protein-protein interaction. Biochemistry 49, 9256–9268.10.1021/bi1008968Suche in Google Scholar

Chodera, J.D. and Mobley, D.L. (2013). Entropy-enthalpy compensation: role and ramifications in biomolecular ligand recognition and design. Annu. Rev. Biophys. 42, 121–142.10.1146/annurev-biophys-083012-130318Suche in Google Scholar

Emsley, P. and Cowtan, K. (2004). Coot: model-building tools for molecular graphics. Acta Cryst. D60, 2126–2132.10.1107/S0907444904019158Suche in Google Scholar

Frigerio, F., Coda, A., Pugliese, L., Lionetti, C., Menegatti, E., Amiconi, G., Schnebli, H.P., Ascenzi, P., and Bolognesi, M. (1992). Crystal and molecular structure of the bovine alpha-chymotrypsin-eglin c complex at 2.0 A resolution. J. Mol. Biol. 225, 107–123.10.1016/0022-2836(92)91029-OSuche in Google Scholar

Ghai, R., Falconer, R.J., and Collins, B.M. (2012). Applications of isothermal titration calorimetry in pure and applied research – survey of the literature from 2010. J. Mol. Recognit. 25, 32–52.10.1002/jmr.1167Suche in Google Scholar PubMed

Heinz, D.W., Hyberts, S.G., Peng, J.W., Priestle, J.P., Wagner, G., and Grutter, M.G. (1992). Changing the inhibitory specificity and function of the proteinase inhibitor eglin c by site-directed mutagenesis: functional and structural investigation. Biochemistry 31, 8755–8766.10.1021/bi00152a011Suche in Google Scholar

Hipler, K., Priestle, J.P., Rahuel, J., and Grutter, M.G. (1992). X-ray crystal structure of the serine proteinase inhibitor eglin c at 1.95 Å resolution. FEBS Lett. 309, 139–145.10.1016/0014-5793(92)81082-WSuche in Google Scholar

Hyberts, S.G., Goldberg, M.S., Havel, T.F., and Wagner, G. (1992). The solution structure of eglin c based on measurements of many NOEs and coupling constants and its comparison with X-ray structures. Protein Sci. 1, 736–751.10.1002/pro.5560010606Suche in Google Scholar PubMed PubMed Central

Kabsch, W. (2010). XDS. Acta Cryst. D66, 125–132.10.1107/S0907444909047337Suche in Google Scholar PubMed PubMed Central

Koh, C.Y. and Kini, R.M. (2009). Molecular diversity of anticoagulants from haematophagous animals. Thromb Haemost. 102, 437–453.10.1160/TH09-04-0221Suche in Google Scholar PubMed

Krissinel, E. and Henrick, K. (2007). Inference of macromolecular assemblies from crystalline state. J. Mol. Biol. 372, 774–797.10.1016/j.jmb.2007.05.022Suche in Google Scholar PubMed

Ladbury, J.E. (2010). Calorimetry as a tool for understanding biomolecular interactions and an aid to drug design. Biochem. Soc. Trans. 38, 888–893.10.1042/BST0380888Suche in Google Scholar PubMed

Ladbury, J.E., Klebe, G., and Freire, E. (2010). Adding calorimetric data to decision making in lead discovery: a hot tip. Nat. Rev. Drug Discov. 9, 23–27.10.1038/nrd3054Suche in Google Scholar PubMed

Laskowski, M. and Qasim, M.A. (2000). What can the structures of enzyme-inhibitor complexes tell us about the structures of enzyme substrate complexes? Biochim. Biophys. Acta 1477, 324–337.Suche in Google Scholar

Leslie, A.G.W. (2006). The integration of macromolecular diffraction data. Acta Cryst. D62, 48–57.10.1107/97809553602060000675Suche in Google Scholar

McCoy, A.J., Grosse-Kunstleve, R.W., Adams, P.D., Winn, M.D., Storoni, L.C., and Read, R.J. (2007). Phaser crystallographic software. J. Appl. Cryst. 40, 658–674.10.1107/S0021889807021206Suche in Google Scholar PubMed PubMed Central

McPhalen, C.A., Schnebli, H.P., and James, M.N. (1985). Crystal and molecular structure of the inhibitor eglin from leeches in complex with subtilisin Carlsberg. FEBS Lett. 188, 55–58.10.1016/0014-5793(85)80873-5Suche in Google Scholar

Murshudov, G.N., Vagin, A.A., and Dodson, E.J. (1997). Refinement of macromolecular structures by the maximum-likelihood method. Acta Cryst. D53, 240–255.10.1107/S0907444996012255Suche in Google Scholar

Rauh, D., Reyda, S., Klebe, G., and Stubbs, M.T. (2002). Trypsin mutants for structure-based drug design: expression, refolding and crystallisation. Biol. Chem. 383, 1309–1314.10.1515/BC.2002.148Suche in Google Scholar

Rauh, D., Klebe, G., and Stubbs, M.T. (2004). Understanding protein-ligand interactions: the price of protein flexibility. J. Mol. Biol. 335, 1325–1341.10.1016/j.jmb.2003.11.041Suche in Google Scholar

Reichert, D., Zinkevich, T., Saalwächter, K, and Krushelnitsky A. (2012). The relation of the X-ray B-factor to protein dynamics: insights from recent dynamic solid-state NMR data. J. Biomol. Struct. Dyn. 30, 617–627.10.1080/07391102.2012.689695Suche in Google Scholar

Reyda, S., Sohn, C., Klebe, G., Rall, K., Ullmann, D., Jakubke, H.D., and Stubbs, M.T. (2003). Reconstructing the binding site of factor Xa in trypsin reveals ligand-induced structural plasticity. J. Mol. Biol. 325, 963–977.10.1016/S0022-2836(02)01337-2Suche in Google Scholar

Seemuller, U., Meier, M., Ohlsson, K., Muller, H.P., and Fritz, H. (1977). Isolation and characterisation of a low molecular weight inhibitor (of chymotrypsin and human granulocytic elastase and cathepsin G) from leeches. Hoppe-Seyler’s Z. Physiol. Chem. 358, 1105–1107.10.1515/bchm2.1977.358.2.1105Suche in Google Scholar

Seemuller, U., Eulitz, M., Fritz, H., and Strobl, A. (1980). Structure of the elastase-cathepsin G inhibitor of the leech Hirudo medicinalis. Hoppe-Seyler’s Z. Physiol. Chem. 361, 1841–1846.Suche in Google Scholar

Straub, A., Roehrig, S., and Hillisch, A. (2011). Oral, direct thrombin and factor Xa inhibitors: the replacement for warfarin, leeches, and pig intestines? Angew.Chem Int. Ed. Engl. 50, 4574–4590.Suche in Google Scholar

Tziridis, A., Rauh, D., Neumann, P., Kolenko, P., Menzel, A., Bräuer, U., Ursel, C., Steinmetz, P., Stürzebecher, J., Schweinitz, A., et al. (2014). Correlating structure and ligand affinity in drug discovery: a cautionary tale involving second shell residues. Biol. Chem. 395, 893–906.10.1515/hsz-2014-0158Suche in Google Scholar

Weiner, M.P., Costa, G.L., Schoettlin, W., Cline, J., Mathur, E., and Bauer, J.C. (1994). Site-directed mutagenesis of double-stranded DNA by the polymerase chain-reaction. Gene 151, 119–123.10.1016/0378-1119(94)90641-6Suche in Google Scholar

Winn, M.D., Ballard, C.C., Cowtan, K.D., Dodson, E.J., Emsley, P., Evans, P.R., Keegan, R.M., Krissinel, E.B., Leslie, A.G.W., McCoy, A., et al. (2011). Overview of the CCP4 suite and current developments. Acta Cryst. D67, 235–242.Suche in Google Scholar

Supplemental Material: The online version of this article (DOI 10.1515/hsz-2014-0177) offers supplementary material, available to authorized users.

Received: 2014-3-27
Accepted: 2014-5-5
Published Online: 2014-7-8
Published in Print: 2014-7-1

©2014 by Walter de Gruyter Berlin/Boston

Artikel in diesem Heft

  1. Frontmatter
  2. Guest Editorial
  3. Highlight: conformational transitions in macromolecular interactions
  4. Single-molecule spectroscopy of unfolded proteins and chaperonin action
  5. Influence of the polypeptide environment next to amyloidogenic peptides on fibril formation
  6. Structure of large dsDNA viruses
  7. Functional aspects of extracellular cyclophilins
  8. Generic tools for conditionally altering protein abundance and phenotypes on demand
  9. Structural insights into calmodulin/Munc13 interaction
  10. Interaction of linear polyamines with negatively charged phospholipids: the effect of polyamine charge distance
  11. Interaction of the human N-Ras protein with lipid raft model membranes of varying degrees of complexity
  12. Lanthanides as substitutes for calcium ions in the activation of plant α-type phospholipase D
  13. Insights from reconstitution reactions of COPII vesicle formation using pure components and low mechanical perturbation
  14. Identification of key residues in the formate channel FocA that control import and export of formate
  15. Twin-arginine translocation-arresting protein regions contact TatA and TatB
  16. Biophysical and biochemical analysis of hnRNP K: arginine methylation, reversible aggregation and combinatorial binding to nucleic acids
  17. An ancient oxidoreductase making differential use of its cofactors
  18. Biophysical characterization of polyomavirus minor capsid proteins
  19. Structural basis for PTPA interaction with the invariant C-terminal tail of PP2A
  20. Correlating structure and ligand affinity in drug discovery: a cautionary tale involving second shell residues
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https://doi.org/10.1515/hsz-2014-0177
Schlagwörter für diesen Artikel
crystal structure; enthalpy entropy compensation; isothermal titration calorimetry; protein flexibility; thermodynamics

Artikel in diesem Heft

  1. Frontmatter
  2. Guest Editorial
  3. Highlight: conformational transitions in macromolecular interactions
  4. Single-molecule spectroscopy of unfolded proteins and chaperonin action
  5. Influence of the polypeptide environment next to amyloidogenic peptides on fibril formation
  6. Structure of large dsDNA viruses
  7. Functional aspects of extracellular cyclophilins
  8. Generic tools for conditionally altering protein abundance and phenotypes on demand
  9. Structural insights into calmodulin/Munc13 interaction
  10. Interaction of linear polyamines with negatively charged phospholipids: the effect of polyamine charge distance
  11. Interaction of the human N-Ras protein with lipid raft model membranes of varying degrees of complexity
  12. Lanthanides as substitutes for calcium ions in the activation of plant α-type phospholipase D
  13. Insights from reconstitution reactions of COPII vesicle formation using pure components and low mechanical perturbation
  14. Identification of key residues in the formate channel FocA that control import and export of formate
  15. Twin-arginine translocation-arresting protein regions contact TatA and TatB
  16. Biophysical and biochemical analysis of hnRNP K: arginine methylation, reversible aggregation and combinatorial binding to nucleic acids
  17. An ancient oxidoreductase making differential use of its cofactors
  18. Biophysical characterization of polyomavirus minor capsid proteins
  19. Structural basis for PTPA interaction with the invariant C-terminal tail of PP2A
  20. Correlating structure and ligand affinity in drug discovery: a cautionary tale involving second shell residues
  21. Thermodynamic signatures in macromolecular interactions involving conformational flexibility
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