Open AccessPurification and characterisation of xylitol dehydrogenase from Neurospora crassa NCL communication No. 6347.
Author(s) -
Phadtare S.U,
Rawat U.B,
Rao M.B
Publication year - 1997
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1997.tb10174.x
Subject(s) - neurospora crassa , xylitol , sorbitol , ammonium sulfate precipitation , biochemistry , xylose , molecular mass , nad+ kinase , enzyme , chemistry , chromatography , size exclusion chromatography , sorbitol dehydrogenase , agarose , dehydrogenase , biology , fermentation , mutant , gene
The purification and characterisation of one of the key enzymes in the conversion of xylose to ethanol, xylitol dehydrogenase (XD), from Neurospora crassa is reported. This organism has the unique ability to directly convert biomass to ethanol. The enzyme was purified 81‐fold by fractional ammonium sulfate precipitation, column chromatography on DEAE‐cellulose, Sephacryl S‐200 and β‐NAD agarose. The purified enzyme had a native molecular mass of 87 kDa (gel filtration) and was composed of two identical subunits of molecular mass 43.6 kDa (SDS‐PAGE). The enzyme was most active at pH 8.4 and at 28°C. It was most stable at pH 7 and at 4°C. It was activated by Mg 2+ and stabilised by Ca 2+ , Mg 2+ and Mn 2+ . It showed activity towards xylitol and sorbitol, the respective K m s being 28.5 and 100 mM and the K m for NAD was 0.7 mM.