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Journal of Biomolecular NMR

, Volume 45, Issue 1–2, pp 157–169 | Cite as

Conformational entropy changes upon lactose binding to the carbohydrate recognition domain of galectin-3

  • Carl Diehl
  • Samuel Genheden
  • Kristofer Modig
  • Ulf Ryde
  • Mikael Akke
Article

Abstract

The conformational entropy of proteins can make significant contributions to the free energy of ligand binding. NMR spin relaxation enables site-specific investigation of conformational entropy, via order parameters that parameterize local reorientational fluctuations of rank-2 tensors. Here we have probed the conformational entropy of lactose binding to the carbohydrate recognition domain of galectin-3 (Gal3), a protein that plays an important role in cell growth, cell differentiation, cell cycle regulation, and apoptosis, making it a potential target for therapeutic intervention in inflammation and cancer. We used 15N spin relaxation experiments and molecular dynamics simulations to monitor the backbone amides and secondary amines of the tryptophan and arginine side chains in the ligand-free and lactose-bound states of Gal3. Overall, we observe good agreement between the experimental and computed order parameters of the ligand-free and lactose-bound states. Thus, the 15N spin relaxation data indicate that the molecular dynamics simulations provide reliable information on the conformational entropy of the binding process. The molecular dynamics simulations reveal a correlation between the simulated order parameters and residue-specific backbone entropy, re-emphasizing that order parameters provide useful estimates of local conformational entropy. The present results show that the protein backbone exhibits an increase in conformational entropy upon binding lactose, without any accompanying structural changes.

Keywords

Spin relaxation Order parameters Molecular dynamics simulations Ligand binding Entropy 

Notes

Acknowledgments

This work was supported by the Swedish Research Council (MA, UR), The Göran Gustafsson Foundation for Research in Natural Sciences and Medicine (MA), and the FLÄK Research School for Pharmaceutical Sciences at Lund University (MA, UR). Computer resources were provided by Lunarc at Lund University and HPC2N at Umeå University. We thank Hakon Leffler for the plasmid harboring the Gal3-thioredoxin fusion construct, and HL, Ulf Nilsson, and Gunnar Karlström for discussions.

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© Springer Science+Business Media B.V. 2009

Authors and Affiliations

  1. 1.Center for Molecular Protein Science, Biophysical ChemistryLund UniversityLundSweden
  2. 2.Theoretical ChemistryLund UniversityLundSweden

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