Neutrons describe ectoine effects on water H-bonding and hydration around a soluble protein and a cell membrane.

Giuseppe Zaccai,Yann Fichou,Victor M Galvan Josa,François-Xavier Gallat,Susanne Von Gronau,Gabriel J Cuello,Bruno Demé,Markus Neumann,Dieter Oesterhelt,Martine Moulin,Jérôme Combet,Martin Weik,Irina Bagyan,Michael Haertlein,Anne Martel

doi:10.1038/srep31434

Abstract

Understanding adaptation to extreme environments remains a challenge of high biotechnological potential for fundamental molecular biology. The cytosol of many microorganisms, isolated from saline environments, reversibly accumulates molar concentrations of the osmolyte ectoine to counterbalance fluctuating external salt concentrations. Although they have been studied extensively by thermodynamic and spectroscopic methods, direct experimental structural data have, so far, been lacking on ectoine-water-protein interactions. In this paper, in vivo deuterium labeling, small angle neutron scattering, neutron membrane diffraction and inelastic scattering are combined with neutron liquids diffraction to characterize the extreme ectoine-containing solvent and its effects on purple membrane of H. salinarum and E. coli maltose binding protein. The data reveal that ectoine is excluded from the hydration layer at the membrane surface and does not affect membrane molecular dynamics, and prove a previous hypothesis that ectoine is excluded from a monolayer of dense hydration water around the soluble protein. Neutron liquids diffraction to atomic resolution shows how ectoine enhances the remarkable properties of H-bonds in water—properties that are essential for the proper organization, stabilization and dynamics of biological structures.

Highlights

1 ns molecular dynamics simulations[13]
E.coli maltose binding protein (MBP)), which can be obtained with various levels of deuterium labeling, has been used extensively as a model for biophysical studies[35], while the hydration dependence of structure and dynamics of purple membranes of Halobacterium salinarum (PM) is currently the best characterized for a natural membrane36–38. (i) The hydration shell around MBP in solution with ectoine was measured by small angle neutron scattering (SANS), by using natural abundance and deuterated protein and H2O/D2O contrast variation[29]; (ii) PM occur naturally as highly ordered two-dimensional crystalline patches of bacteriorhodopsin and lipids
Energy-resolved neutron scattering data show that membrane molecular dynamics is not affected by the presence of ectoine in the solvent environment

Summary

Introduction

1 ns molecular dynamics simulations[13]. An early small angle neutron scattering (SANS) experiment provided experimental evidence for such a structure around ribonuclease in aqueous solvent containing glycerol[14]. Hahn et al.[9] combined surface plasmon resonance, confocal Raman spectroscopy, molecular dynamics simulations, and density functional theory calculations to study the local hydration shell around ectoine and its influence on the binding of a gene-5-protein to a single-stranded DNA. E.coli maltose binding protein (MBP) (calculated pI 5.47)), which can be obtained with various levels of deuterium labeling, has been used extensively as a model for biophysical studies[35], while the hydration dependence of structure and dynamics of purple membranes of Halobacterium salinarum (PM) is currently the best characterized for a natural membrane[36,37,38]. (i) The hydration shell around MBP in solution with ectoine was measured by SANS, by using natural abundance and deuterated protein and H2O/D2O contrast variation[29];

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