Solution conformation of the Met 61 to His 61 mutant of Pseudomonas stutzeri ZoBell ferrocytochrome c-551.
Solution conformation of the Met 61 to His 61 mutant of Pseudomonas stutzeri ZoBell ferrocytochrome c-551.
复制标题
施氏假单胞菌 ZoBell 铁细胞色素 c-551 的 Met 61 到 His 61 突变体的溶液构象。
DOI：
10.1016/s0006-3495(01)76258-3
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发表时间：
2001
影响因子：
3.4
通讯作者：
Timkovich,R
中科院分区：
文献类型：
--
作者：
Miller,GT;Hardman,JK;Timkovich,R
对来自施氏假单胞菌ZoBell亚株的编码细菌细胞色素c-551的基因进行突变，将不变的第六配体蛋氨酸残基转化为组氨酸，产生位点特异性突变体M61 H。质子NMR共振分配的所有主链和大多数侧链质子的抗磁性，还原形式在pH值为9.2和333 K的二维NMR技术。距离约束（1074）由核奥弗豪泽增强确定，主链扭转角约束（72）由标量耦合估计确定。通过模拟退火方法计算蛋白质的溶液构象。对于28个计算结构，除了末端残基1、2、81和82之外，骨架原子的均方根位移为0.52 π（σ=0.096），所有重原子的均方根位移为0.90 π（σ=0.122）。突变蛋白的整体折叠与野生型相同。最大的变化定位在残基60-65的肽跨度中。突变体蛋白质最引人注目的行为是，在室温和中性pH下，它以类似于在温和变性条件下描述的几种蛋白质的熔融球状状态的状态存在，但突变体在高pH和温度下转化为更有序的状态。
The gene encoding for bacterial cytochrome c-551 fromPseudomonas stutzerisubstrain ZoBell has been mutated to convert the invariant sixth ligand methionine residue into histidine, creating the site-specific mutant M61H. Proton NMR resonance assignments were made for all main-chain and most-side chain protons in the diamagnetic, reduced form at pH 9.2 and 333K by two-dimensional NMR techniques. Distance constraints (1074) were determined from nuclear Overhauser enhancements and main-chain torsion-angle constraints (72) from scalar coupling estimates. Solution conformations for the protein were computed by the simulated annealing approach. For 28 computed structures, the root mean squared displacement from the average structure excluding the terminal residues 1, 2, 81, and 82 was 0.52Å (σ=0.096) for backbone atoms and 0.90Å (σ=0.122) for all heavy atoms. The global folding of the mutant protein is the same as for wild type. The biggest changes are localized in a peptide span over residues 60–65. The most striking behavior of the mutant protein is that at room temperature and neutral pH it exists in a state similar to the molten globular state that has been described for several proteins under mild denaturing conditions, but the mutant converts to a more ordered state at high pH and temperature.