Cloning and expression of the gene encoding the thermophilic NAD(P)H-FMN oxidoreductase coupling with the desulfurization enzymes from Paenibacillus sp A11-2

The gene encoding the NAI)(P)H-flavin oxidoreductase (flavin reductase) whi ch couples with the thermophilic dibenzothiophene (DBT)-desulfurizing monoo xygenases of Paenibacillus sp. All-2 was cloned in Escherichia coli and des ignated tdsD, Nucleotide sequence analysis suggested that the gene product consisted of 200 amino acids and showed about 30%, 27% and 26% amino acid s equence similarity to the major flavin reductase of Vibriofischeri, the NAD H dehydrogenase of Thermus thermophilus and several oxygen-insensitive NAD( P)H nitroreductases in the Enterobacteriaceae family, respectively. Both th e growing and resting recombinant E, coli, in which tdsD was coexpressed wi th a set of desulfurizing genes, showed a rate of DBT removal about 5 times higher than the recombinants lacking tdsD. Maximal desulfurization was obs erved close to 45 degreesC and 55 degreesC in the resting cells and in the cell-free extraction reaction with the tdsD-coexpressing recombinants, resp ectively. In an organic/aqueous biphasic system, the coexpression of tdsD a lso markedly enhanced the rate of DBT removal.