Reactivation of methionine synthase from Thermotoga maritima (TM0268) requires the downstream gene product TM0269
Reactivation of methionine synthase from Thermotoga maritima (TM0268) requires the downstream gene product TM0269
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DOI：
10.1110/ps.072936307
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发表时间：
2007-08-01
期刊：
影响因子：
8
通讯作者：
Ludwig, Martha
中科院分区：
文献类型：
--
作者：
Huang, Sha;Romanchuk, Gail;Ludwig, Martha
海栖热袍菌基因产物TM 0269的晶体结构，作为T. maritima的序列，揭示了与来自大肠杆菌的钴胺素依赖性甲硫氨酸合酶（MetH）的第四模块的结构同源性，尽管缺乏显著的序列同源性。指定TM 0269的基因与另一个基因TM 0268非常接近，TM 0268显示出与E. coli MetH.第四模块E.大肠杆菌MetH是辅因子的cob（II）丙氨酸形式的还原性再甲基化所必需的，并结合甲基供体进行这种再活化，S-腺苷甲硫氨酸（S-腺苷甲硫氨酸）。在存在和不存在TM 0269和蛋氨酸的情况下，对蛋氨酸形成速率的测量表明，TM 0269和蛋氨酸都是TM 0268的无活性cob（II）丙氨酸形式再活化所必需的。这些活性测量结果证实了TM 0269基因产物功能的基于结构的归属。在TM 0269、蛋氨酸和还原剂存在下，T.在80 ℃附近，纯化蛋白的比活性最高，约为E. coli蛋氨酸合成酶（MetH）。比较了TM 0269与E. coliMetH的结果表明，同源蛋白质与C57 Met的结合可能略有不同。然而，在E.大肠杆菌MetH，它构成了一个基本的结构元件，被保留在T. maritima再激活蛋白，尽管序列存在显著差异。
The crystal structure of the Thermotoga maritima gene product TM0269, determined as part of genome- wide structural coverage of T. maritima by the Joint Center for Structural Genomics, revealed structural homology with the fourth module of the cobalamin-dependent methionine synthase (MetH) from Escherichia coli, despite the lack of significant sequence homology. The gene specifying TM0269 lies in close proximity to another gene, TM0268, which shows sequence homology with the first three modules of E. coli MetH. The fourth module of E. coli MetH is required for reductive remethylation of the cob(II) alamin form of the cofactor and binds the methyl donor for this reactivation, S-adenosylmethionine (AdoMet). Measurements of the rates of methionine formation in the presence and absence of TM0269 and AdoMet demonstrate that both TM0269 and AdoMet are required for reactivation of the inactive cob(II) alamin form of TM0268. These activity measurements confirm the structure-based assignment of the function of the TM0269 gene product. In the presence of TM0269, AdoMet, and reductants, the measured activity of T. maritima MetH is maximal near 80 degrees C, where the specific activity of the purified protein is similar to 15% of that of E. coli methionine synthase ( MetH) at 37 degrees C. Comparisons of the structures and sequences of TM0269 and the reactivation domain of E. coli MetH suggest that AdoMet may be bound somewhat differently by the homologous proteins. However, the conformation of a hairpin that is critical for cobalamin binding in E. coli MetH, which constitutes an essential structural element, is retained in the T. maritima reactivation protein despite striking divergence of the sequences.