Structural rearrangements of the two domains of Azotobacter vinelandii rhodanese upon sulfane sulfur release:: essential molecular dynamics, 15N NMR relaxation and deuterium exchange on the uniformly labeled protein
Structural rearrangements of the two domains of Azotobacter vinelandii rhodanese upon sulfane sulfur release:: essential molecular dynamics, 15N NMR relaxation and deuterium exchange on the uniformly labeled protein
复制标题
DOI：
10.1016/j.ijbiomac.2003.08.010
复制
发表时间：
2003-12-01
影响因子：
8.2
通讯作者：
Paci, M
中科院分区：
文献类型：
--
作者：
Cicero, DO;Melino, S;Paci, M
棕色固氮菌是一种31 kDa的硫转移酶蛋白，在氰化物解毒过程中催化硫原子从硫代硫酸盐转移到氰化物，并能在铁-硫簇中插入硫原子。用高分辨核磁共振对N-15同位素进行了均匀标记，然后进行了主干序列指认。该酶的载硫态和放硫态具有非常相似的HSQC谱，具有良好的光谱色散。很少有共振显示这两种形式之间的化学位移发生了变化。所有酰胺氮原子的弛豫参数T-1、T-2和H-1-N-15NOE以及同位素交换动力学表明，这两种形式具有相同的整体关联时间和流体力学性质。同时，基本动力学研究表明，催化型半胱氨酸过硫基的形成和释放对蛋白质的整体构象没有显著影响。这些结果结合在一起，给出了一个明确的答案，即酶的催化机制是否涉及两个结构域的构象和/或相互取向的变化。相反，这些结果清楚地表明，在催化机制下，蛋白质的两个结构域表现为一个独特的折叠。(C)2003爱思唯尔B.V.保留所有权利。
The Azotobacter vinelandii rhodanese is a 31 kDa sulfurtransferase protein that catalyzes the transfer of sulfur atom from thiosulfate to cyanide in the detoxification process from cyanide and is able to insert sulfur atom in the iron-sulfur cluster. A study of the uniformly N-15 isotopic labeling by high resolution NMR, before obtaining the backbone sequential assignment, has been carried out. The sulfur loaded and the sulfur discharged forms of the enzyme show very similar HSQC spectra with a good spectral dispersion. Few resonances show changes in chemical shift between the two forms. Relaxation parameters T-1, T-2 and H-1-N-15 NOE of all amide nitrogen atoms, as well as isotope exchange kinetics, show that the two forms exhibit the same global correlation time and hydrodynamic properties. In parallel, essential dynamics studies show that formation and discharging of catalytic cysteine persulfide group has no significant impact on the overall conformation of the protein. These results, taken together, give a clearcut answer to the question if the catalytic mechanism of the enzyme involves a change in the conformation and/or in the mutual orientation of the two domains. On the contrary these results clearly indicate that upon the catalytic mechanism the two domains of the protein behave as a unique fold. (C) 2003 Elsevier B.V. All rights reserved.