A conserved cysteine near the P-site is accessible to cysteine modifications and increases ROS stability in P-type plasma membrane H+-ATPase.
A conserved cysteine near the P-site is accessible to cysteine modifications and increases ROS stability in P-type plasma membrane H+-ATPase.
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P位点附近的保守半胱氨酸可进行半胱氨酸修饰，并增加P型质膜H-ATP酶中ROS的稳定性
DOI：
10.1042/bcj20200559
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Palmgren
中科院分区：
文献类型：
--
作者：
Pedersen;Ravnsborg;Hayashi;Becher;Jensen;Stöhr;Palmgren
含硫氨基酸残基在抗氧化反应中起作用，这可以由过量的铜和过氧化氢产生的活性氧引起。在所有的Na+/K+、Ca 2+和H+泵P型ATP酶中，半胱氨酸残基存在于必需天冬氨酸残基上游的两个残基，其在每个催化循环中被强制磷酸化。尽管它的保守性，这个半胱氨酸残基的功能是迄今为止未知的。本研究通过突变拟南芥质膜H ~+-ATP酶同工型2（AHA 2），并在酵母中异源表达，分析了AHA 2中相应的半胱氨酸残基（Cys-327）的功能。丙氨酸、丝氨酸和亮氨酸取代的酶动力学与野生型泵相同，但突变泵对铜和过氧化氢的敏感性增加。肽鉴定和质谱测序表明Cys-327易于氧化。这些数据表明Cys-327在质膜H+-ATP酶中起保护性残基的作用，并且可能在其他P-型ATP酶中也起保护性残基的作用。
Sulfur-containing amino acid residues function in antioxidative responses, which can be induced by the reactive oxygen species generated by excessive copper and hydrogen peroxide. In all Na+/K+, Ca2+, and H+pumping P-type ATPases, a cysteine residue is present two residues upstream of the essential aspartate residue, which is obligatorily phosphorylated in each catalytic cycle. Despite its conservation, the function of this cysteine residue was hitherto unknown. In this study, we analyzed the function of the corresponding cysteine residue (Cys-327) in the autoinhibited plasma membrane H+-ATPase isoform 2 (AHA2) fromArabidopsis thalianaby mutagenesis and heterologous expression in a yeast host. Enzyme kinetics of alanine, serine, and leucine substitutions were identical with those of the wild-type pump but the sensitivity of the mutant pumps was increased towards copper and hydrogen peroxide. Peptide identification and sequencing by mass spectrometry demonstrated that Cys-327 was prone to oxidation. These data suggest that Cys-327 functions as a protective residue in the plasma membrane H+-ATPase, and possibly in other P-type ATPases as well.
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