A flavoprotein encoded in Selenomonas ruminantium is characterized after expression in Escherichia coli.
Selenomonas ruminantium is an obligate anaerobe that is very important for the provision of vitamin B12 to ruminants, which are particularly dependent upon this cofactor. One important use for vitamin B12 in anaerobic bacteria is for the utilization of glycerol as carbon source. A new flavoprotein has been found expressed by Escherichia coli from a plasmid created as part of a gene library of S. ruminantium. The 2.5-kb fragment of chromosomal DNA responsible for protein expression contains parts of two operons. Only one polypeptide (the flavoprotein) encoded by the S. ruminantium DNA is produced in E. coli in large amounts. The gene for the flavoprotein has been identified and is probably transcribed as part of an operon involved in glycerol metabolism in S. ruminantium. The flavoprotein has been purified and its molecular properties have been examined. Sequence analysis showed that this protein is a divergent member of the family of nitroreductases. Pure protein is a homodimer with a molecular weight of 44,500, containing one molecule of FMN per dimer. Like other nitroreductases, this protein forms a complex with pyridine nucleotide (NADPH), but unlike other nitroreductases, it fails to be reduced in this complex at a biologically significant rate. It has none of the common catalytic properties of other members of the nitroreductase family.
Anderson
Peter J
PJ
Molecular and Cellular Biology, Armidale, New South Wales 2351, Australia.
Cole
Lindsay J
LJ
McKay
David B
DB
Entsch
Barrie
B
D000595:Amino Acid Sequence; D015139:Blotting, Southern; D002852:Chromatography, Ion Exchange; D004926:Escherichia coli; D005420:Flavoproteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D016680:Genome, Bacterial; D008969:Molecular Sequence Data; D009601:Nitroreductases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D020357:Selenomonas / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment
D005420:Flavoproteins; D011994:Recombinant Proteins; D009601:Nitroreductases