Burgess B K, Stiefel E I, Newton W E
J Biol Chem. 1980 Jan 25;255(2):353-6.
The interactions of the iron-molybdenum cofactor, FeMoco, isolated from acid-treated Azotobacter vinelandii molybdenum-iron protein (Av1) with EDTA and thiophenol in N-methylformamide solution have been reinvestigated. Our studies show that EDTA alone is sufficient to eliminate the EPR signal of dithionite-reduced FeMoco. Neither light/5-deazaflavin nor carbon monoxide are required, which implies that this EPR-silent form of FeMoco does not correspond to the EPR-silent, substrate-reducing state of Av1. As EDTA-treated FeMoco does not regain EPR activity on addition of sodium dithionite or thiophenol, it is apparently distinct from the EPR-silent form of either dye-oxidized FeMoco or dye-oxidized Av1. Thiophenol sharpens the EPR signal of dithionite-reduced FeMoco and shifts the g = 3.3 feature to g = 3.6. This shift is complete at 1:1 ratio of thiophenol/Mo atom, while the EDTA effect requires about 40 molecules/Mo atom. Thiophenol and EDTA probably affect different sites of FeMoco. The binding of either reactant does not affect the activity of FeMoco as measured by its ability to reconstitute extracts of A. vinelandii mutant UW45.
对从酸处理的棕色固氮菌钼铁蛋白（Av1）中分离出的铁钼辅因子（FeMoco）在N-甲基甲酰胺溶液中与乙二胺四乙酸（EDTA）和苯硫酚的相互作用进行了重新研究。我们的研究表明，单独的EDTA就足以消除连二亚硫酸盐还原的FeMoco的电子顺磁共振（EPR）信号。既不需要光/5-脱氮黄素，也不需要一氧化碳，这意味着这种EPR沉默形式的FeMoco与Av1的EPR沉默、底物还原状态不对应。由于经EDTA处理的FeMoco在添加连二亚硫酸钠或苯硫酚后不会恢复EPR活性，它显然与染料氧化的FeMoco或染料氧化的Av1的EPR沉默形式不同。苯硫酚会锐化连二亚硫酸盐还原的FeMoco的EPR信号，并将g = 3.3的特征峰移至g = 3.6。当苯硫酚与钼原子的比例为1:1时，这种位移完成，而EDTA的作用则需要约40个分子/钼原子。苯硫酚和EDTA可能影响FeMoco的不同位点。通过其重组棕色固氮菌突变体UW45提取物的能力来衡量，两种反应物的结合均不影响FeMoco的活性。
