BIOCHEMICAL-CHARACTERIZATION OF NFSA, THE ESCHERICHIA-COLI MAJOR NITROREDUCTASE EXHIBITING A HIGH AMINO-ACID-SEQUENCE HOMOLOGY TO FRP, A VIBRIO-HARVEYI FLAVIN OXIDOREDUCTASE

We identified the nfsA gene, encoding the major oxygen-insensitive nit roreductase in Escherichia coli, and determined its position on the E. coli map to be 19 min. We also purified its gene product, NfsA, to ho mogeneity. It was suggested that NfsA is a nonglobular protein with a molecular weight of 26,799 and is associated tightly with a flavin mon onucleotide. Its amino acid sequence is highly similar to that of Frp, a flavin oxidoreductase from Vibrio harveyi (B. Lei, M. Liu, S. Huang , and S.-C. Tu, J. Bacteriol. 176:3552-3558, 1994), an observation sup porting the notion that E. coli nitroreductase and luminescent-bacteri um flavin reductase families are intimately related in evolution. Alth ough no appreciable sequence similarity was detected between two E. co li nitroreductases, NfsA and NfsB, NfsA exhibited a low level of the f lavin reductase activity and a broad electron acceptor specificity sim ilar to those of NfsB. NfsA reduced nitrofurazone by a ping-pong Bi-Bi mechanism possibly to generate a two-electron transfer product.