AZOTOBACTER-VINELANDII NADPH-FERREDOXIN REDUCTASE CLONING, SEQUENCING, AND OVEREXPRESSION

Azotobacter vinelandii ferredoxin I (AvFdI) controls the expression of another protein that was originally designated Protein X. Recently we reported that Protein X is a NADPH-specific flavoprotein that binds s pecifically to FdI (Isas, J. M., and Burgess, B. K. (1994) J. Biol. Ch em, 269, 19404-19409), The gene encoding this protein has now been clo ned and sequenced, Protein X is 33% identical and has an overall 53% s imilarity with the fpr gene product from Escherichia coli that encodes NADPH:ferredoxin reductase, On the basis of this similarity and the s imilarity of the physical properties of the two proteins, we now desig nate Protein X as A. vinelandii NADPH:ferredoxin reductase and its gen e as the fpr gene, The protein has been overexpressed in its native ba ckground in A. vinelandii by using the broad host range multicopy plas mid, pKT230. In addition to being regulated by FdI, the fpr gene produ ct is overexpressed when A. vinelandii is grown under N-2-fixing condi tions even though the fpr gene is not preceded by a nif specific promo ter, By analogy to what is known about fpr expression in E. coli, we p ropose that FdI may exert its regulatory effect on fpr by interacting with the SoxRS regulon.