An NMR-derived model for the solution structure of oxidized putidaredoxin, a 2-Fe, 2-S ferredoxin from Pseudomonas.
An NMR-derived model for the solution structure of oxidized putidaredoxin, a 2-Fe, 2-S ferredoxin from Pseudomonas.
复制标题
氧化腐臭还蛋白（一种来自假单胞菌的 2-Fe, 2-S 铁氧还蛋白）溶液结构的 NMR 衍生模型。
DOI：
10.1021/bi00187a006
复制
发表时间：
1994
期刊：
影响因子：
2.9
通讯作者：
Lyons,TA
中科院分区：
文献类型：
--
作者：
Pochapsky,TC;Ye,XM;Ratnaswamy,G;Lyons,TA
修订稿于 1994 年 3 月 23 日收到®摘要：已使用同核 NMR 方法确定了氧化腐臭氧还蛋白 (Pdx) 的溶液结构模型，Pdx 是一种含有 Fe2S2 簇的 106 个残基球状蛋白。 Pdx 是 Fe2S2Cys4 铁氧还蛋白类中的第一个，可作为待进行结构表征的 P-450 单加氧酶的电子转移伴侣，但尚未确定 Pdx 或任何紧密同源蛋白质的晶体结构。 Pdx是细胞色素P-450cam的生理氧化还原伴侣——在模拟退火结构精修中使用了总共878个NOE距离约束、66个源自NH-CaH耦合常数的角度约束和五个顺磁展宽约束，以获得主链原子的成对均方根偏差为1.14A和所有非氢原子的成对均方根偏差为1.80A的结构族。约内共振的顺磁展宽。 8-金属簇的半径阻止在蛋白质的该区域使用核磁共振衍生的约束；用于模拟金属簇环境的结构约束是通过定点诱变和模型化合物并通过与已知的铁氧还蛋白结构进行比较而获得的。 Pdx 保留了与其他具有结构特征的 Fe2S2Cys4 铁氧还蛋白类似的折叠拓扑，但与其他铁氧还蛋白的不同之处在于，在蛋白质的 C 端一半包含明显更紧凑的结构。活生物体中的电子转移必须在空间和时间上受到严格调控，以防止“短路”，即跨化学势梯度的快速平衡。这种监管所涉及的机制尚未得到很好的理解，并且是当前争论的主题（McLendon，1988；Thomson，1991）。动力学和机制研究提供了有关生物电子转移的大量信息（Moser 等，1992；Beratan 等，1992），但如果要了解分子水平上的电子转移机制，所涉及蛋白质的结构数据至关重要。
Revised Manuscript Received March 23, 1994® abstract: A model for the solution structure of oxidized putidaredoxin (Pdx), a 106-residue globular protein containing a Fe2S2 cluster, has been determined using homonuclear NMR methods. Pdx is the First of the class of Fe2S2Cys4 ferredoxins which act as electron-transfer partners for P-450monooxygenases to be structurally characterized, and no crystal structure has been determined for Pdx or for any closely homologous protein. Pdx is the physiological redox partner of cytochrome P-450cam-A total of 878 NOE distance constraints, 66< j> angular constraints derived from NH-CaH coupling constants, and five paramagnetic broadening constraints were used in simulated annealing structural refinements to obtain a family of structures with pairwise rms deviations of 1.14 A for backbone atoms and 1.80 A for all non-hydrogen atoms. Paramagnetic broadening of resonances within a ca. 8-A radius of the metal cluster prevents the use of NMR-derived constraints in this region of the protein; structural constraints used to model the environment of the metal cluster were obtained from site-directed mutagenesis and model compounds and by comparison with known ferredoxin structures. Pdx retains a similar folding topologyto other structurally characterized Fe2S2Cys4 ferredoxins but differs from the other ferredoxins in containing a significantly more compact structure in the C-terminal half of the protein.Electron transfer in living organisms must be tightly regulated in both space and time in order to prevent “short circuits”, that is, rapid equilibration across chemical potential gradients. The mechanisms involved in such regulation are not well understood and are the subject of much current debate (McLendon, 1988; Thomson, 1991). Kinetic and mechanistic studies have provided much information on biological electron transfer (Moser et al., 1992; Beratan et al., 1992), but structural data on the proteins involved are critical if an understanding of the mechanism (s) of electron transfer at the molecular level is to be gained.