Crystallization of Arthrobacter sp. strain 1C N-(1-D-carboxyethyl)-L-norvaline dehydrogenase and its complex with NAD+

The novel NAD+-linked opine dehydrogenase from a soil isolate Arthrobacter sp. strain 1C belongs to an enzyme superfamily whose members exhibit quite diverse substrate specificites. Crystals of this opine dehydrogenase, obtained in the presence or absence of co-factor and substrates, have been shown to diffract to beyond 1.8 ? resolution. X-ray precession photographs have established that the crystals belong to space group P21212, with cell parameters a = 104.9, b = 80.0, c = 45.5 ? and a single subunit in the asymmetric unit. The elucidation of the three-dimensional structure of this enzyme will provide a structural framework for this novel class of dehydrogenases to enable a comparison to be made with other enzyme families and also as the basis for mutagenesis experiments directed towards the production of natural and synthetic opine-type compounds containing two chiral centres.     

Desolvation of BSA-ligand complexes measured using the quartz crystal microbalance and dual polarization interferometer

By taking advantage of their unique difference in hydration sensitivity, we have shown that dual polarization interferometer (DPI) and quartz-crystal microbalance with dissipation monitoring (QCM-D) measurements can be used together to explore the degree of desolvation involved in the binding of small drug molecules to an immobilized bovine serum albumin film in real time. Results with DPI and QCM-D show significantly different mass values for three ligands of varying hydrophobicities that may be attributed to changes in the degree of hydration of the ligand-protein complexes in accordance with the physicochemical properties of the ligands. Furthermore, our data suggest that masses measured by QCM-D can be overwhelmed by changes in water content of ligand-protein, binary complexes, which has important consequences for future studies using mechanical resonators to study protein-binding events.