H+-Pyrophosphatase of Rhodospirillum rubrum
H+-Pyrophosphatase of Rhodospirillum rubrum
复制标题
红色红螺菌 H-焦磷酸酶
DOI：
--
复制
发表时间：
2002
影响因子：
4.8
通讯作者：
R. Lahti
中科院分区：
文献类型：
--
作者：
G. Belogurov;M. Turkina;Anni Penttinen;Saila Huopalahti;A. Baykov;R. Lahti
在大肠杆菌C43(DE3)中表达了光合细菌红色红螺菌的H+-转位焦磷酸酶(H+-PPase)。重组H+-PPase存在于内膜小泡中，它催化PPI的水解和H+的转运，并催化PPI的合成。大肠杆菌囊泡中H+-PPase的水解力是红色红曲霉发色团中H+-PPase的8倍，但对H+-PPase抑制剂氨基亚甲基二磷酸表现出相似的敏感性，对可溶性PPase抑制剂氟化物不敏感。利用该表达系统，我们发现用脂肪族残基取代Cys185、Cys222或Cys573对H+-PPase的活性没有影响，但降低了其对巯基修饰剂Mersalyl的敏感性。H+-PPase的三个半胱氨酸残基均缺失，完全抵抗肉豆蔻醇的作用。Mg2+和MgPPI保护Cys185和Cys573不受该试剂的修饰，但不保护Cys222。对23个非冗余H+-PPase序列的系统发育分析将其分为两个亚家族。一个亚家族包含Cys222，包含所有已知的K+非依赖性H+-PPase，而另一个包含保守的Cys573，但缺乏Cys222，包含所有已知的K+依赖的H+-PPase。这些数据提示，Cys在222和573位的发生率与H+-PPase的K+依赖性之间存在特定的联系。
H+-translocating pyrophosphatase (H+-PPase) of the photosynthetic bacteriumRhodospirillum rubrum was expressed in Escherichia coli C43(DE3) cells. Recombinant H+-PPase was observed in inner membrane vesicles, where it catalyzed both PPi hydrolysis coupled with H+ transport into the vesicles and PPi synthesis. The hydrolytic activity of H+-PPase in E. coli vesicles was eight times greater than that in R. rubrum chromatophores but exhibited similar sensitivity to the H+-PPase inhibitor, aminomethylenediphosphonate, and insensitivity to the soluble PPase inhibitor, fluoride. Using this expression system, we showed that substitution of Cys185, Cys222, or Cys573 with aliphatic residues had no effect on the activity of H+-PPase but decreased its sensitivity to the sulfhydryl modifying reagent, mersalyl. H+-PPase lacking all three Cys residues was completely resistant to the effects of mersalyl. Mg2+ and MgPPi protected Cys185 and Cys573 from modification by this agent but not Cys222. Phylogenetic analyses of 23 nonredundant H+-PPase sequences led to classification into two subfamilies. One subfamily invariably contains Cys222and includes all known K+-independent H+-PPases, whereas the other incorporates a conserved Cys573 but lacks Cys222 and includes all known K+-dependent H+-PPases. These data suggest a specific link between the incidence of Cys at positions 222 and 573 and the K+ dependence of H+-PPase.
影响因子：
1.5
作者：
McIntosh, MT;Drozdowicz, YM;Vaidya, AB
通讯作者：
Vaidya, AB