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首页> 外文期刊>Journal of Molecular Biology >Crystal structure of the MJ0490 gene product of the hyperthermophilic archaebacterium Methanococcus jannaschii, a novel member of the lactate/malate family of dehydrogenases
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Crystal structure of the MJ0490 gene product of the hyperthermophilic archaebacterium Methanococcus jannaschii, a novel member of the lactate/malate family of dehydrogenases

机译：嗜热古细菌甲烷甲烷球菌（脱氢酶的乳酸/苹果酸家族的新成员）的MJ0490基因产物的晶体结构

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MJ0490 gene, one of the only two genes of Methanococcus jannaschii showing sequence similarity to the lactate/malate family of dehydrogenases, was classified initially as coding for a putative L-lactate dehydrogenase (LDH). It has been re-classified as a malate dehydrogenase (MDH) gene, because it shows significant sequence similarity to MT0188, MDH II from Methanobacterium thermoautotrophicum strain DeltaH. The three-dimensional structure of its gene product has been determined in two crystal forms: a "dimeric" structure in the orthorhombic crystal at 1.9 Angstrom resolution and a "tetrameric" structure in the tetragonal crystal at 2.8 Angstrom. These structures share a similar subunit fold with other LDHs and MDHs. The tetrameric structure resembles typical tetrameric LDHs. The dimeric structure is equivalent to the P-dimer of tetrameric LDHs, unlike dimeric MDHs, which correspond to the Q-dimer. The structure reveals that the cofactor NADP(H) is bound at the active site, despite the fact that it was not intentionally added during protein purification and crystallization. The preference of NADP(H) over NAD(H) has been supported by activity assays. The cofactor preference is explained by the presence of a glycine residue in the cofactor binding pocket (Gly33), which replaces a conserved aspartate (or glutamate) residue in other NAD-dependent LDHs or MDHs. Preference for NADP(H) is contributed by hydrogen bonds between the oxygen atoms of the monophosphate group and the ribose sugar of adenosine in NADP(H) and the side-chains of Ser9, Arg34, His36, and Ser37. The MDH activity of MJ0490 is made possible by Arg86, which is conserved in MDHs but not in LDHs. The enzymatic assay showed that the MJ0490 protein possesses the fructose-1,6-bisphosphate-activated LDH activity (reduction). Thus the MJ0490 gene product appears to be a novel member of the lactate/malate dehydrogenase family, displaying an LDH scaffold and exhibiting a relaxed substrate and cofactor specificities in NADP(H) and NAD(H)-dependent malate and lactate dehydrogenase reactions. (C) 2001 Academic Press. [References: 53]
机译：MJ0490基因是詹氏甲烷球菌仅有的两个与脱氢酶的乳酸/苹果酸家族具有序列相似性的基因之一，最初被分类为编码假定的L-乳酸脱氢酶（LDH）。它已被重新分类为苹果酸脱氢酶（MDH）基因，因为它与来自嗜热甲烷化杆菌DeltaH的MT0188，MDH II具有显着的序列相似性。它的基因产物的三维结构已确定为两种晶体形式：正交晶体中1.9埃分辨率的“二聚体”结构和四方晶体中2.8埃分辨率的“四聚体”结构。这些结构与其他LDH和MDH共享相似的亚基折叠。四聚体结构类似于典型的四聚体LDH。与二聚体MDH不同，二聚体结构等同于四聚体LDH的P-二聚体，后者对应于Q-二聚体。该结构揭示了辅因子NADP（H）在活性位点结合，尽管在蛋白质纯化和结晶过程中并非故意添加辅因子。活性测定支持NADP（H）优于NAD（H）。辅助因子偏爱的解释是在辅助因子结合袋（Gly33）中存在甘氨酸残基，该残基取代了其他NAD依赖性LDH或MDH中保守的天冬氨酸（或谷氨酸）残基。 NADP（H）的优先选择是由NADP（H）中单磷酸基团的氧原子与腺苷的核糖之间的氢键以及Ser9，Arg34，His36和Ser37的侧链组成的。 MJ0490的MDH活性通过Arg86得以实现，它在MDH中是保守的，而在LDH中却不保守。酶促测定表明，MJ0490蛋白具有-1,6-双磷酸果糖激活的LDH活性（还原）。因此，MJ0490基因产物似乎是乳酸/苹果酸脱氢酶家族的新成员，在NADP（H）和NAD（H）依赖性苹果酸和乳酸脱氢酶反应中显示出LDH支架，并显示出轻松的底物和辅因子特异性。 （C）2001学术出版社。 [参考：53]

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