Bhattacharyya Sudeepa, Habibi-Nazhad Bahram, Amegbey Godwin, Slupsky Carolyn M, Yee Adelinda, Arrowsmith Cheryl, Wishart David S
Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada.
Biochemistry. 2002 Apr 16;41(15):4760-70. doi: 10.1021/bi0115176.
As part of a high-throughput, structural proteomic project we have used NMR spectroscopy to determine the solution structure and ascertain the function of a previously unknown, conserved protein (MtH895) from the thermophilic archeon Methanobacterium thermoautotrophicum. Our findings indicate that MtH895 contains a central four-stranded beta-sheet core surrounded by two helices on one side and a third on the other. It has an overall fold superficially similar to that of a glutaredoxin. However, detailed analysis of its three-dimensional structure along with molecular docking simulations of its interaction with T7 DNA polymerase (a thioredoxin-specific substrate) and comparisons with other known members of the thioredoxin/glutaredoxin family of proteins strongly suggest that MtH895 is more akin to a thioredoxin. Furthermore, measurement of the pK(a) values of its active site thiols along with direct measurements of the thioredoxin/glutaredoxin activity has confirmed that MtH895 is, indeed, a thioredoxin and exhibits no glutaredoxin activity. We have also identified a group of previously unknown proteins from several other archaebacteria that have significant (34-44%) sequence identity with MtH895. These proteins have unusual active site -CXXC- motifs not found in any known thioredoxin or glutaredoxin. On the basis of the results presented here, we predict that these small proteins are all members of a new class of truncated thioredoxins.
作为高通量结构蛋白质组学项目的一部分，我们利用核磁共振光谱法确定了嗜热古菌嗜热自养甲烷杆菌中一种先前未知的保守蛋白（MtH895）的溶液结构，并确定了其功能。我们的研究结果表明，MtH895含有一个由四条链组成的中央β-折叠核心，一侧被两条螺旋包围，另一侧被第三条螺旋包围。它的整体折叠与谷氧还蛋白表面相似。然而，对其三维结构的详细分析以及其与T7 DNA聚合酶（硫氧还蛋白特异性底物）相互作用的分子对接模拟，以及与硫氧还蛋白/谷氧还蛋白家族其他已知成员的比较，强烈表明MtH895更类似于硫氧还蛋白。此外，对其活性位点硫醇的pK(a)值的测量以及对硫氧还蛋白/谷氧还蛋白活性的直接测量证实，MtH895确实是一种硫氧还蛋白，不表现出谷氧还蛋白活性。我们还从其他几种古细菌中鉴定出一组先前未知的蛋白质，它们与MtH895具有显著的（34-44%）序列同一性。这些蛋白质具有在任何已知硫氧还蛋白或谷氧还蛋白中未发现的不寻常的活性位点 -CXXC- 基序。基于此处给出的结果，我们预测这些小蛋白质都是一类新的截短硫氧还蛋白的成员。
