Taguchi Yuichi, Sugishima Masakazu, Fukuyama Keiichi
Department of Biology, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560-0043, Japan.
Biochemistry. 2004 Apr 13;43(14):4111-8. doi: 10.1021/bi036052t.
ATP sulfurylase (ATPS) is a ubiquitous enzyme that catalyzes the transfer of the adenylyl group from ATP to inorganic sulfate, producing adenosine 5'-phosphosulfate (APS) and pyrophosphate. The crystal structure of ATPS from Thermus thermophilus HB8 (TtATPS, 347 amino acid residues) in complex with APS was determined at 2.5 A resolution. TtATPS is composed of three domains [domain I (residues 1-134), domain II (residues 135-290), and domain III (residues 291-347)], like the Riftia pachyptila symbiont ATPS, but lacks a fourth domain present in ATPSs from the yeast Saccharomyces cerevisiae and from the fungus Penicillium chrysogenum. TtATPS forms a dimer in the crystal, and the manner of subunit association is different from that observed in dimeric R. pachyptila symbiont ATPS and in the hexameric S. cerevisiae and P. chrysogenum ATPSs. APS is located in the active site of TtATPS, which contains several motifs (QXRN, HXXH, and GRD) conserved in ATPSs. Unexpectedly, TtATPS binds one metal ion per subunit in domain III. XAFS measurement of the crystal and the Bijvoet difference Fourier map unambiguously characterized the metal ion as a zinc ion. The zinc ion is tetrahedrally coordinated by Cys294, Cys297, Cys306, and His310, and could not be removed from the protein by treatment with EDTA. The zinc ion binding site is far from the active site. Because all four residues coordinated to the zinc ion are conserved in the ATPSs from thermophilic bacteria such as Archaeoglobus fulgidus, Pyrococcus abyssi, and Sulfolobus solfataricus, zinc ion chelation may contribute to the thermal stability of these ATPSs.
ATP硫酸化酶（ATPS）是一种普遍存在的酶，它催化腺苷基团从ATP转移至无机硫酸盐，生成5'-磷酸腺苷硫酸（APS）和焦磷酸。嗜热栖热菌HB8（TtATPS，347个氨基酸残基）的ATPS与APS复合物的晶体结构在2.5埃分辨率下得以确定。TtATPS与巨型管虫共生菌的ATPS一样，由三个结构域组成[结构域I（残基1-134）、结构域II（残基135-290）和结构域III（残基291-347）]，但缺少酿酒酵母和产黄青霉的ATPS中存在的第四个结构域。TtATPS在晶体中形成二聚体，其亚基结合方式不同于在二聚体巨型管虫共生菌ATPS以及六聚体酿酒酵母和产黄青霉ATPS中观察到的方式。APS位于TtATPS的活性位点，该活性位点包含在ATPS中保守的几个基序（QXRN、HXXH和GRD）。出乎意料的是，TtATPS在结构域III中每个亚基结合一个金属离子。对晶体的XAFS测量以及比沃伊特差分傅里叶图明确将该金属离子表征为锌离子。锌离子由半胱氨酸294、半胱氨酸297、半胱氨酸306和组氨酸310以四面体方式配位，并且不能通过用EDTA处理从蛋白质中去除。锌离子结合位点远离活性位点。由于与锌离子配位的所有四个残基在嗜热栖热菌、深渊栖热球菌和嗜热栖硫叶菌等嗜热细菌的ATPS中是保守的，锌离子螯合可能有助于这些ATPS的热稳定性。
