Allen R M, Chatterjee R, Ludden P W, Shah V K
Department of Biochemistry, College of Agricultural and Life Sciences, University of Wisconsin, Madison 53706, USA.
J Biol Chem. 1995 Nov 10;270(45):26890-6. doi: 10.1074/jbc.270.45.26890.
NifB-co is an iron- and sulfur-containing precursor to the iron-molybdenum cofactor (FeMo-co) of dinitrogenase. The synthesis of NifB-co requires at least the product of the nifB gene. Incorporation of 55Fe and 35S from NifB-co into FeMo-co was observed only when all components of the in vitro FeMo-co synthesis system were present. Incorporation of iron and sulfur from NifB-co into dinitrogenase was not observed in control experiments in which the apodinitrogenase (lacking FeMo-co) was initially activated with purified, unlabeled FeMo-co or in assays where NifB-co was oxygen-inactivated prior to addition to the synthesis system. These data clearly demonstrate that iron and sulfur from active NifB-co are specifically incorporated into FeMo-co of dinitrogenase and provide direct biochemical identification of an iron-sulfur precursor of FeMo-co. Under different in vitro FeMo-co synthesis conditions, iron and sulfur from NifB-co were associated with at least two other proteins (NIFNE and gamma) that are involved in the formation of active dinitrogenase. The results presented here suggest that multiple FeMo-co processing steps might occur on NIFNE and that FeMo-co synthesis is most likely completed prior to the association of FeMo-co with gamma.
NifB-辅酶是固氮酶铁钼辅因子（FeMo-辅因子）含硫和铁的前体。NifB-辅酶的合成至少需要nifB基因的产物。只有当体外FeMo-辅因子合成系统的所有组分都存在时，才能观察到NifB-辅酶中的55Fe和35S掺入FeMo-辅因子。在对照实验中未观察到NifB-辅酶中的铁和硫掺入固氮酶，在这些对照实验中，脱辅基固氮酶（缺乏FeMo-辅因子）最初用纯化的、未标记的FeMo-辅因子激活，或者在将NifB-辅酶加入合成系统之前先进行氧灭活的测定中。这些数据清楚地表明，活性NifB-辅酶中的铁和硫被特异性地掺入固氮酶的FeMo-辅因子中，并提供了FeMo-辅因子的铁硫前体的直接生化鉴定。在不同的体外FeMo-辅因子合成条件下，NifB-辅酶中的铁和硫与至少另外两种参与活性固氮酶形成的蛋白质（NIFNE和γ）相关。此处给出的结果表明，多个FeMo-辅因子加工步骤可能发生在NIFNE上，并且FeMo-辅因子的合成很可能在FeMo-辅因子与γ结合之前完成。
