Rangaraj P, Ruttimann-Johnson C, Shah V K, Ludden P W
Department of Biochemistry and Center for the Study of Nitrogen Fixation, College of Agricultural and Life Sciences, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
J Biol Chem. 2001 May 11;276(19):15968-74. doi: 10.1074/jbc.M100907200. Epub 2001 Feb 16.
Iron-molybdenum cofactor (FeMo-co) biosynthesis involves the participation of several proteins. We have used (55)Fe-labeled NifB-co, the specific iron and sulfur donor to FeMo-co, to investigate the accumulation of protein-bound precursors of FeMo-co. The (55)Fe label from radiolabeled NifB-co became associated with two major protein bands when the in vitro FeMo-co synthesis reaction was carried out with the extract of an Azotobacter vinelandii mutant lacking apodinitrogenase. One of the bands, termed (55)Fe-labeled upper band, was purified and shown to be NifH by immunoblot analysis. The (55)Fe-labeled lower band was identified as NifX by N-terminal sequencing. NifX purified from an A. vinelandii nifB strain showed a different electrophoretic mobility on anoxic native gels than did NifX with the FeMo-co precursor bound.
铁钼辅因子（FeMo-co）的生物合成涉及多种蛋白质的参与。我们使用了（55）Fe标记的NifB-co，即FeMo-co的特定铁和硫供体，来研究FeMo-co蛋白质结合前体的积累。当用缺乏脱辅基固氮酶的棕色固氮菌突变体提取物进行体外FeMo-co合成反应时，放射性标记的NifB-co中的（55）Fe标记与两条主要蛋白质条带相关联。其中一条带，称为（55）Fe标记的上带，经纯化并通过免疫印迹分析显示为NifH。通过N端测序将（55）Fe标记的下带鉴定为NifX。从棕色固氮菌nifB菌株中纯化的NifX在无氧天然凝胶上的电泳迁移率与结合了FeMo-co前体的NifX不同。
