Raman spectra of interchanging β-lactamase inhibitor intermediates on the millisecond time scale.

Rapid mix-rapid freeze is a powerful method to study the mechanisms of enzyme-substrate reactions in solution. Here we report a protocol that combines this method with normal (non-resonance) Raman microscopy to enable us to define molecular details of intermediates at early time points. With this combined method, SHV-1, a class A β-lactamase, and tazobactam, a commercially available β-lactamase inhibitor, were rapidly mixed on the millisecond time scale and then were flash-frozen by injection into an isopentane solution surrounded by liquid nitrogen.

In-situ confocal Raman observation of structural changes of insulin crystals in sequential dehydration process.

In-situ confocal Raman spectroscopy combined with relative humidity (RH) control technique was used to study the sequential dehydration process of insulin crystals. By gradually decreasing the ambient RH of the insulin crystal, the content of the hydration water in the crystal was quantitatively controlled. Tyrosine (Tyr) residues were very sensitive to the micro-environmental changes, and four Raman features 828cm(-1), 852cm(-1), 1174cm(-1) and 1206cm(-1) of Tyr were employed to monitor the dehydration process.

Adsorption and desorption kinetics of water in lysozyme crystal investigated by confocal Raman spectroscopy.

Adsorption and desorption are critical to crystal engineering for the protein crystal applications. In this paper, we present a new method to study the adsorption and desorption kinetics of waters in lysozyme crystals. H(2)O and D(2)O were used as ideal indicators for the purpose.

Micro-Raman analysis of association equilibria in supersaturated NaClO4 droplets.

We report an in situ strategy for the quantitative analysis of association equilibria in a single NaClO(4) droplet of nanogram mass deposited on a quartz substrate. In the new approach, the single droplet was forced to enter into a supersaturated state by decreasing the relative humidity (RH) of the environment, allowing accurate control over the concentration of the solute within the droplet. An analysis of the solvated structure of the ClO(4)(-) anion with change in molar water-to-solute ratio (WSR) was performed by micro-Raman spectroscopy within the confines of a single droplet.

Micro-Raman spectroscopic observation of water expulsion induced destruction of hydrophobic clusters in crystalline lysozyme.

In this paper, three kinds of solid lysozyme samples with different water contents were investigated by confocal Raman spectroscopy. For the rod-like lysozyme crystal with highest water content, a sudden decrease of the intensity ratio of the doublet at 1338 and 1360 cm(-1) was observed when the ambient relative humidity (RH) was lower than 86%, indicating the destruction of hydrophobic clusters of lysozyme induced by the expulsion of the hydration water from the crystal. In contrast to the rod-like crystal, tetragonal crystal and floor-like precipitate with a smaller amount of water showed no change of the structures of the hydrophobic clusters when the relative humidity was decreasing.

Micro-Raman study on the conformation behavior of succinate in supersaturated sodium succinate aerosols.

Micro-Raman spectra of supersaturated aerosols of sodium succinate were obtained. The conformation behavior of the succinate dianion as a function of relative humidity (RH) was investigated by combining micro-Raman spectroscopy with theoretical calculations. A shoulder at 968 cm(-1) of the v(C-CO(2)(-)) band on the rise in more concentrated droplets was believed indicative of conformation transformations.