Structure of a human S-adenosylmethionine decarboxylase self-processing ester intermediate and mechanism of putrescine stimulation of processing as revealed by the H243A mutant
Structure of a human S-adenosylmethionine decarboxylase self-processing ester intermediate and mechanism of putrescine stimulation of processing as revealed by the H243A mutant
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DOI：
10.1021/bi010736o
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发表时间：
2001-08-14
期刊：
影响因子：
2.9
通讯作者：
Ealick, SE
中科院分区：
文献类型：
--
作者：
Ekstrom, JL;Tolbert, WD;Ealick, SE
S-腺苷甲硫氨酸脱羧酶（S-Adenosylmethionine decarboxylase，S-MetDC）作为酶原合成，其在腐胺刺激的反应中通过N ->O酰基转移和β-消除来切割自身，以产生在新的N-末端具有催化必需的异戊酰残基的活性酶。N -->O酰基转移启动了其他蛋白质如内含肽和酰胺水解酶的自我加工，但它们在这些蛋白质中的机制还不清楚。我们已经解析了H243 A突变体的晶体结构，分辨率为1.5埃。突变蛋白被困在酯的形式，提供了明确的证据，酯中间体的结构在处理的异戊酰酶。此外，腐胺分子结合在β-三明治内的带电区域中，并通过氢键将两个β-片层交联到几个酸性残基和有序的水分子上。高分辨率的结构提供了洞察的自加工反应的机制，并提供证据的腐胺模拟的自加工反应的机制。腐胺或4-氨基丁醇对突变体CIMetDC酶原加工的影响的研究与一个模型一致，在该模型中，单个激活剂分子与掩埋的Asp 174，Glu 178和Glu 256相互作用，导致Glu 11位置的改变，从而刺激自我加工。
S-Adenosylmethionine decarboxylase (AdoMetDC) is synthesized as a proenzyme that cleaves itself in a putrescine-stimulated reaction via an N -->O acyl shift and beta -elimination to produce an active enzyme with a catalytically essential pyruvoyl residue at the new N-terminus. N -->O acyl shifts initiate the self-processing of other proteins such as inteins and amidohydrolases, but their mechanisms in such proteins are not well understood. We have solved the crystal structure of the H243A mutant of AdoMetDC to 1.5 Angstrom resolution. The mutant protein is trapped in the ester form, providing clear evidence for the structure of the ester intermediate in the processing of pyruvoyl enzymes. In addition, a putrescine molecule is bound in a charged region within the beta -sandwich, and cross-links the two beta -sheets through hydrogen bonds to several acidic residues and ordered water molecules. The high-resolution structure provides insight into the mechanism for the self-processing reaction and provides evidence for the mechanism for simulation of the self-processing reaction by putrescine. Studies of the effects of putrescine or 4-aminobutanol on the processing of mutant AdoMetDC proenzymes are consistent with a model in which a single activator molecule interacts with buried Asp 174, Glu178, and Glu256, leading to an alteration in the position of Glu11, resulting in stimulation of self-processing.