活跃的站点赖氨酸orotate phosphoribosyltransferase。
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多尔夫曼Segura Grubmeyer C, E, R
活跃的站点赖氨酸orotate phosphoribosyltransferase。
生物化学杂志。1993年9月25日,268 (27):20299 - 304。
- PubMed ID
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8376388 (在PubMed]
- 文摘
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Orotate phosphoribosyltransferase (OPRTase;EC 2.4.2.10)催化的形成核苷酸orotidine-5一磷酸从orotate 5-phosphoribosyl-1-pyrophosphate (PRPP)。细菌酶,与哺乳动物的相同器官,是单功能的23000 (r)亚基的二聚体。大量的可用性高纯度结晶鼠伤寒沙门氏菌OPRTase使我们研究酶的结构/功能。像其他phosphoribosyltransferases OPRTase结合高度带电MgPRPP复杂,以及阴离子orotate,暗示一个活跃的站点包含基本的残留物。美国培养的序列(Scapin G。Sacchettini, j . C。在全世界,。Bhatia, M。Grubmeyer, c (1993) j·摩尔。杂志。230年,1304 - 1308)包含13 12赖氨酸和精氨酸残基,其中Lys-26 arg - 99,赖氨酸- 100,赖氨酸- 103和参数- 156是守恒的身份OPRTases序列中从其他生物,Lys-19和参数- 161取而代之的是另一种基本残留在一个或多个序列。赖氨酸修饰符2 4 6-trinitrobenzene磺酸盐灭活美国沙门氏菌感染OPRTase伪一阶过程,和OMP PRPP对失活提供了良好的保护。光谱定量trinitrophenyl (TNP)集团公司表明,失活与公司每单元一组对照组。令人惊讶的是,胰蛋白酶的蛋白水解作用其次是高效液相色谱法和氨基酸序列分析显示,四肽,包含三个不同的赖氨酸,已经修改。 Peptides modified at Lys-26, Lys-100 and Lys-103, as well as a doubly modified peptide containing TNP groups at Lys-100 and Lys-103, were identified. Inactivation kinetics showed that the 3 lysine residues were modified at equal rates. Protection studies demonstrated that Lys-26, and to a lesser extent Lys-100 and Lys-103, were protected against modification by OMP, whereas PRPP protected Lys-26, Lys-100 and Lys-103. Pyrophosphate protected Lys-100 and Lys-103. The results suggest active site locations for the sequence-conserved and TNP-modified lysine residues, with Lys-26 interacting with the ribose-5-phosphate moiety of OMP and PRPP, and Lys-100 and Lys-103 interacting with the pyrophosphate moiety of PRPP.