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Comaparative analyses of a thermostable alpha-amylase and its mesophilic homologue show maximal catalytic activity at different temperatures with the former requiring a higher temperature. The temperatures for protein unfolding are same in both cases with the thermostable alpha-amylase exhibiting higher structural flexibility.

Neutron scattering technique has been used to study structural and dynamical properties of hydration water and its influence on the behavior of halobacteria purple membranes. Results demonstrate the effect of hydration process on the membrane properties.

The dynamics of proton binding to the extracellular and the cytoplasmic surfaces of the purple membrane were measured by laser-induced proton pulses. Purple membranes, selectively labeled by fluorescein at Lys-129 of bacteriorhodopsin, were pulsed by protons released in the aqueous bulk from excited pyranine (8-hydroxy-1,3,6-pyrenetrisulfonate) and the reaction of protons with the indicators was measured. Kinetic analysis of the data imply that the two faces of the membrane differ in their buffer capacities and in their rates of interaction with bulk protons. The extracellular surface of the purple membrane contains one anionic proton binding site per protein molecule with pK = 5.1. This site is within a Coulomb cage radius ([approximately equal to]15 [Angstrom]) from Lys-129. The cytoplasmic surface of the purple membrane bears 4-5 protonable moieties (pK = 5.1) that, due to close proximity, function as a common proton binding site. The reaction of the proton with this cluster is at a very fast rate (3[center dot][10.sup.10] [M.sup.-1][center dot][s-.sup.1]). The proximity between the elements is sufficiently high that even in 100 mM NaCl they still function as a cluster. Extraction of the chromophore retinal from the protein has a marked effect on the carboxylates of the cytoplasmic surface, and two to three of them assume positions that almost bar their reaction with bulk protons. The protonation dynamics determined at the surface of the purple membrane is of relevance both for the vectorial proton transport mechanism of bacteriorhodopsin and for energy coupling, not only in halobacteria, but also in complex chemiosmotic systems such as mitochondrial and thylakoid membranes.

Quasielastic incoherent neutron scattering from hydrogen atoms, which are distributed nearly homogeneously in biological molecules, allows the investigation of diffusive motions occurring on the pico- to nanosecond time scale. A quasielastic incoherent neutron scattering study was performed on the integral membrane protein bacteriorhodopsin (BR), which is a light-driven proton pump in Halobacterium salinarium. BR is embedded in lipids, forming patches in the cell membrane of the organism, which are the so called purple membranes (PMs). Measurements were carried out at room temperature on oriented PM-stacks hydrated at two different levels (low hydration, h = 0.03 g of [D.sub.2]O per g of PM; high hydration, h = 0.28 g of [D.sub.2]O per g of PM) using time-of-flight spectrometers. From the measured spectra, different diffusive components were identified and analyzed with respect to the influence of hydration. This study supports the idea that a decrease in hydration results in an appreciable decrease in internal molecular flexibility of the protein structure. Because it is known from studies on the function of BR that the pump activity is reduced if the hydration level of the protein is insufficient, we conclude that the observed diffusive motions are essential for the function of this protein. A detailed analysis and classification of the different kinds of diffusive motions, predominantly occurring in PMs under physiological conditions, is presented.

Neutron diffraction studies of the positional oscillation of the deuterium labels of bacteriorhodopsin (BR) in the beta-ionone ring (D11) and adjacent to the Schiff base (D5), between the BR568 ground state and the M412 photocycle intermediate, reveals that the transition from BR568 to M412 does not affect the D11 ring position in the projected structure. The D5 label moves closer to the D11 ring by 1.4 plus or minus 0.9 angstroms. The reduction in interlabel distance indicates that the C-5 to C-13 portion of the polyene chain gets out of the membrane plane by 11 degrees plus or minus 6 degrees toward the cytoplasm during the BR568-M412 transition due to the all-trans/13-cis isomerization.