Purification and Characterization of Poly(vinyl alcohol) Dehydrogenase fromPseudomonassp. 113P3

Pseudomonas sp. 113P3, when cultured on a medium containing poly(vinyl alcohol) (PVA) in the presence of pyroroquinoline quinone (PQQ), produced a high level of a PV A dehydrogenase (PV ADH). This enzyme was purified to a homogeneous state by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) from the cell-free extract and found to be a hemoquino-protein. 1,3-Diols, especially PVA, were the most favorable substrate; primary alcohols and secondary alcohols and other diols could not act as substrates. PVADH showed an activity in the presence of PQQ. The molecular weight of the enzyme was found to be about 67,000 by SD8–PAGE and high pressure liquid chromatography using a TSK gel G-3000SW column. Our PVADH catalyzed dehydrogenation of PYA with an artificial electron acceptor, like the PQQ-dependent PVADH reported by Shimao et al. The molecular weight, PQQ dependency, requirement for metal ion, and absorption spectra of the PVADH were very similar to those of the hemoquino-protein primary alcohol dehydrogenase (ADH) isolated from Comamonas testosteroni ATCC 15667 (formerly called Pseudomonas testosteroni), but differed in the substrate spectra.