Skjeldal L, Westler W M, Oh B H, Krezel A M, Holden H M, Jacobson B L, Rayment I, Markley J L
National Magnetic Resonance Facility at Madison, Institute for Enzyme Research, Biochemistry Department, University of Wisconsin, Madison 53706.
Biochemistry. 1991 Jul 30;30(30):7363-8. doi: 10.1021/bi00244a002.
Hyperfine 1H NMR signals of the 2Fe-2S* vegetative ferredoxin from Anabaena 7120 have been studied by two-dimensional (2D) magnetization exchange spectroscopy. The rapid longitudinal relaxation rates of these signals required the use of very short nuclear Overhauser effect (NOE) mixing times (0.5-20 ms). The resulting pattern of NOE cross-relaxation peaks when combined with previous 1D NOE results [Dugad, L. B., La Mar, G. N., Banci, L., & Bertini, I. (1990) Biochemistry 29, 2263-2271] led to elucidation of the carbon-bound proton spin systems from each of the four cysteines ligated to the 2Fe-2S* cluster in the reduced ferredoxin. Additional NOE cross peaks were observed that provide information about other amino acid residues that interact with the iron-sulfur cluster. NOE cross peaks were assigned tentatively to Leu27, Arg42, and Ala43 on the basis of the X-ray coordinates of oxidized Anabaena 7120 ferredoxin [Rypniewski, W.R., Breiter, D.R., Benning, M.M., Wesenberg, G., Oh, B.-H., Markley, J.L., Rayment, I., & Holden, H. M. (1991) Biochemistry 30, 4126-4131]. Three chemical exchange cross peaks were detected in magnetization exchange spectra of half-reduced ferredoxin and assigned to the 1H alpha protons of Cys49 and Cys79 [both of whose sulfur atoms are ligated to Fe(III)] and Arg42 (whose amide nitrogen is hydrogen-bonded to one of the inorganic sulfurs of the 2Fe-2S* cluster). The chemical exchange cross peaks provide a means of extending assignments in the spectrum of reduced ferredoxin to assignments in the spectrum of the oxidized protein.(ABSTRACT TRUNCATED AT 250 WORDS)
利用二维（2D）磁化交换光谱研究了来自鱼腥藻7120的2Fe-2S营养型铁氧化还原蛋白的超精细1H NMR信号。这些信号的快速纵向弛豫速率要求使用非常短的核Overhauser效应（NOE）混合时间（0.5 - 20毫秒）。当结合先前的一维NOE结果[Dugad, L. B., La Mar, G. N., Banci, L., & Bertini, I. (1990) Biochemistry 29, 2263 - 2271]时，得到的NOE交叉弛豫峰模式有助于阐明还原型铁氧化还原蛋白中与2Fe-2S簇相连的四个半胱氨酸中每个半胱氨酸的碳结合质子自旋系统。还观察到了额外的NOE交叉峰，这些峰提供了与铁硫簇相互作用的其他氨基酸残基的信息。根据氧化型鱼腥藻7120铁氧化还原蛋白的X射线坐标[Rypniewski, W.R., Breiter, D.R., Benning, M.M., Wesenberg, G., Oh, B.-H., Markley, J.L., Rayment, I., & Holden, H. M. (1991) Biochemistry 30, 412,6 - 4131]，NOE交叉峰被初步指定为Leu27, Arg42和Ala43。在半还原型铁氧化还原蛋白的磁化交换光谱中检测到三个化学交换交叉峰，并指定为Cys49和Cys79的1Hα质子[它们的硫原子都与Fe(III)相连]以及Arg42（其酰胺氮与2Fe-2S*簇的一个无机硫形成氢键）。化学交换交叉峰提供了一种将还原型铁氧化还原蛋白光谱中的归属扩展到氧化型蛋白光谱归属的方法。（摘要截短于250字）
