Characterization of three different flavodoxins from Azotobacter vinelandii

Abstract

The flavodoxins from Azotobacter vinelandii cells grown N₂-fixing and from cells grown NH₄OAc have been purified and characterized. The purified flavodoxins from these cells are a mixture of three different flavodoxins (Fld I, II, III) with different primary structures. The three proteins were separated by fast protein liquid chromatography; Fld I eluted at 0.38 M K Cl, Fld II at 0.43 M K Cl and Fld III at 0.45 M K Cl. The most striking difference between the three flavodoxins was the midpoint potential (pH 7.0, 25°C) of the semiquinone/hydroquinone couple, which was −320 mV for Fld I and −500 mV for the other two flavodoxins (Fld II and Fld III). All three flavodoxins were present in cells grown on NH₄OAc. In cells grown on N₂ as N source only Fld I and Fld II were found. The concentration of Fld II was 10-fold higher in N₂-fixing cells than in cells grown on NH₄OAc. Evidence has been obtained that Fld II is involved in electron transport to nitrogenase. As will be discussed, our observation that preparations of Azotobacter flavodoxin are heterogeneous, has consequences for the published data.