Soft Report - Dipartimento di Fisica - Sapienza

Effect of Solvent and of Confinement on the Dynamics ofHydrated ProteinsThe studies on protein dynamics mostly focused onthe effect of the chemical and physical environmentson the dynamics and the ‘flexibility’ of thebiomolecules. They have involved the use of nonfully aqueous solvents, of confining media (poroussilica hydrogels and saccharide ‘coatings’) and ofmedium to high pressures. The molecular flexibilityof enzymes may be controlled by properly choosingthe physical and chemical properties of theenvironment around the protein. This perspectivehas a number of practical consequences on thebiological, food science and pharmaceutical fields. Anelastic scattering study of the mean square atomicfluctuations of lysozyme in water-glycerol mixturesshowed that the environment rules the proteindynamical transition. Above a certain threshold valuelocated approximately at 0.1 hydration, the thermalstability of lysozyme in glycerol progressivelydecreases. At the same time, the protein internaldynamics as measured by the atomic mean squaredisplacement shows a marked onset. Such a resultseems to confirm that protein flexibility is inverselycorrelated to thermal stability.Several studies in the recent literature have beenaddressed to the relevance of the interactionsbetween the external medium and the proteinsurface in determining the internal protein dynamics.In this respect the study of the dynamic properties ofproteins embedded in saccharide glasses has provento be a most useful experimental approach. Indeed,this is a strategy exploited, in nature, by someorganisms which replace water by a glass formingsolvent. This enables these organisms to affordadverse conditions, like high temperatures orextreme drought, in a state of suspended animationcalled anhydrobiosis. Elastic neutron scatteringmeasurements were performed in the temperaturerange 20-340 K on samples obtained by dryingeither MbCO-trehalose-H 2O or MbCO-trehalose-D 2Osolution, and then by re-hydrating in a 75% relativehumidity D 2O atmosphere. The hydrogen atom meansquare displacements thus obtained indicate that inthe trehalose-water solid glasses or amorphousplasticized systems each component looses itsindividuality becoming a collective system.In view of forthcoming studies on the dynamics ofproteins embedded in porous media, the dynamics ofthe water solvent has been investigated underconfining conditions in silica hydrogels performingelastic temperature scans. The obtained meansquare hydrogen displacements (MSD) point out thatwater in cryogenic conditions is in an amorphousstate and a dynamic glass-like transition around 200K can be evidenced. Test experiments were alsocarried out to investigate the pressure dependenceof the atomic fluctuations in small globular proteins(lysozyme and trypsin). The investigation wascarried out in a pressure range (1 – 2000 bar) belowthe cold denaturation limit having thus enzymes thatwere still biologically active. The MSD data show onlysmall variations with pressure that are howevercoherent with the results of QENS experiments thatshow a small reduction of the volume accessed bythe protons with increasing pressure.Fig. 1. Mean square fluctuation for the two systems,at the high concentration, in the range oftemperature 275 K – 310 K tot/61.61.41.21.00.80.60.40.20.00 50 100 150 200 250 300T (K)Fig. 2. Temperature dependence of the MSDobtained using the gaussian model (eq. 1). ● =Encapsulated Mb; = Mb powder. Continuous line:harmonic contribution of the MSD.AuthorsA. Gliozzi, R. Rolandi, Università di Genova, S.Magazù, U. Wanderlingh, Università di Messina, M.Corti, L. Cantù, Univ. Milano, R. Cordone, A.Cupane,Università di Palermo, A. Deriu Università di Parma,G. Onori, A. Paciaroni Università di Perugia, CRS-SOFT, A. Congiu Castellano Università di Roma LaSapienza, A. Filabozzi, G. Paradossi Università diRoma Tor Vergata.105SOFT Scientific Report 2004-06