人类组胺N-methyltransferase药物基因学:肾脏cDNA的克隆和表达。
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吉拉德B, Otterness DM、木材TC、Honchel R, Wieben ED,温什波姆RM
人类组胺N-methyltransferase药物基因学:肾脏cDNA的克隆和表达。
摩尔杂志。1994年3月,45 (3):461 - 8。
- PubMed ID
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8145732 (在PubMed]
- 文摘
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组胺N-methyltransferase (HNMT)催化NT-methylation组胺。HNMT活动水平在人类红细胞是由一个共同的遗传多态性。我们克隆和表达cDNA HNMT从人体组织作为第一步确定这种基因多态性的分子基础。克隆策略是基于老鼠和人类肾脏HNMT之间可能的序列同源性。人类肾脏cDNA库筛选885 -核苷酸鼠肾脏HNMT cDNA开放阅读框。1.4千碱基cDNA克隆是孤立的,包含两个潜在的翻译起始密码子,在相同的阅读框。最长的人类肾脏cDNA克隆的开放阅读框包含876个核苷酸和蛋白质编码292个氨基酸。这种蛋白质的氨基酸序列与大鼠肾脏HNMT 84%。人类肾脏cDNA克隆在兔网织红细胞体外转录和翻译lystate系统产生蛋白质的表观分子量33 kDa,所估计的钠十二烷基sulfate-polyacrylamide凝胶电泳。人类肾脏cDNA也是subcloned到真核表达载体p91023 (B)。 Partially purified HNMT isolated from the cytosol of GOS-1 cells transfected with this expression construct had biochemical properties similar to those of human kidney HNMT. Human renal cortical HNMT, partially purified human renal cortical HNMT, and partially purified transfected COS-1 cell HNMT had Km values for histamine and S-adenosyl-L-methionine, the two cosubstrates for the enzyme reaction, of 20, 13, and 14 microM and 2.0, 3.0, and 6.2 microM, respectively. IC50 values for the HNMT inhibitor amodiaquine were 0.50, 0.48, and 0.40 microM, respectively, for enzyme from these same three sources. Northern blot analyses performed with poly(A)+ RNA from a series of human tissues including kidney demonstrated three transcripts, approximately 1.3, 3.8, and 4.0 kilobases in length. Cloning of a cDNA for HNMT may now make it possible to determine the molecular basis for the HNMT genetic polymorphism in humans.