Molecular aspects of the hydration process in a globular protein

Molecular Aspects of the Hydration Process in a Globular Protein G. CARER1 A N D A. GIANSANTI Istituto di Fisica dell’Unioersitri degli Studi di R o m a , P.le A.Moro 5- R o m a , Italy E. GRATTON Department of Physics, University of Illinois, Urbana Champaign, Illinois, U.S.A. In past years we have been interested in some general problems of enzyme dynamics [ I ] , and we have considered the possibility that random changes in the hydration of the globular protein may induce conformational fluctuations relevant toward catalysis. We have also obtained experimental evidence [2] of a critical hydration value (at about 20% mg water/mg dry weight) for the onset of the enzymatic activity in lysozyme powder. We have also been able to follow in detail the hydration dependent events of this macromolecule by gravimetric and infrared techniques [ 3 ] . However, there are still some existing problems at the molecular level, particularly in the region of low coverage (below 10% hydration) which we believe are appropriate for discussion here. More exactly, we would like to understand the nature of the rather sharp transition that occurs in this range, and that is displayed by several physical properties, notably by the profile of the I R stretching band of the adsorbed D20. Let us try to discuss the nature of this low hydration transition in terms of the current order-disorder and nucleation theories of embryonic droplets [4]. Ac- cording to nucleation theory it is impossible to condense water on an insoluble particle with a radius smaller than about 100 A, unless the pressure is higher than the equilibrium vapor pressure of liquid water. Therefore, a globular protein dry powder could not become hydrated under normal laboratory conditions if its surface is made of amide backbone and of nonionizable residues. Actually it has just occurred for “soluble particles” like the ionizable residues that water vapor condenses on a protein macromolecule, much in the same way as in the clouds, namely, the condensation of water droplets is induced by small salt particles. Of course this is because the solution process decreases the free energy to compensate for the free energy increase due to the surface tension of the droplet. As a matter of fact in our experiments [2], the first detectable event-when the dry specimen of protein gets hydrated-is correct for the ionization of the acidic side chains. It is conceivable that these strong hydration sites will continue to grow even connecting together similar side chains of dif- ferent close macromolecules. However, while the hydration continues to increase, the surface backbone and the nonionizable side chains finally get involved in a process dominated by the surface tension of these hydrogen bonded species. From an empirical study of the surface entropy of a large number of liquids, International Journal of Quantum Chemistry, Val. XIX, 1133 - 1 135 (1981) ‘c: 1981 by John Wiley & Sons. Inc. CCC 0161-3642/8 I /061133-03$0l .OO