The KdpF subunit is part of the K+-translocating Kdp complex of Escherichia coli and is responsible for stabilization of the complex in vitro
The KdpF subunit is part of the K+-translocating Kdp complex of Escherichia coli and is responsible for stabilization of the complex in vitro
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DOI：
10.1074/jbc.274.53.37901
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发表时间：
1999-12-31
影响因子：
4.8
通讯作者：
Altendorf, K
中科院分区：
文献类型：
--
作者：
Gassel, M;Möllenkamp, T;Altendorf, K
hdpABC操纵子编码大肠杆菌的高亲和力K+转运Kdp复合物（P型ATP酶）。当该操纵子在微细胞中表达时，除了KdpA、KdpB和KdpC亚基之外，还可以在高分辨率SDS-聚丙烯酰胺凝胶上鉴定到迄今为止未被识别的小疏水多肽KdpF。如通过质谱法测定的，该肽以其甲酰化形式存在，并且具有3100 Da的分子量。KdpF对于在低K+（0.1 mM）培养基上生长不是必需的，如通过KdpF的缺失分析所示，但是证明对于体外功能性酶复合物是必需的。在没有KdpF的情况下，膜结合Hdp复合物的ATP酶活性几乎与野生型的没有区别。相反，纯化的洗涤剂溶解的酶复合物显示出酶活性的急剧下降。然而，向KdpABC复合物中加入纯化的KdpF使活性恢复到野生型水平。大肠杆菌脂质具有类似的效果。虽然KdpF对于Hdp复合物在体内的功能不是必需的，但它是复合物的一部分，并在体外作为稳定元件发挥作用。
The hdpABC operon codes for the high affinity K+-translocating Kdp complex (P-type ATPase) of Escherichia coli, Upon expression of this operon in minicells, a so far unrecognized small hydrophobic polypeptide, KdpF, could be identified on high resolution SDS-polyacrylamide gels in addition to the subunits KdpA, KdpB, and KdpC, Furthermore, it could be demonstrated that KdpF remains associated with the purified complex. As determined by mass spectrometry, this peptide is present in its formylated form and has a molecular mass of 3100 Da, KdpF is not essential for growth on low K+ (0.1 mM) medium, as shown by deletion analysis of KdpF, but proved to be indispensable for a functional enzyme complex in vitro. In the absence of KdpF, the ATPase activity of the membrane-bound Hdp complex was almost indistinguishable from that of the wild type. In contrast, the purified detergent-solubilized enzyme complex showed a dramatic decrease in enzymatic activity. However, addition of purified KdpF to the KdpABC complex restored the activity up to wild type level, It is interesting to note that the addition of high amounts of E. coli lipids had a similar effect. Although KdpF is not essential for the function of the Hdp complex in vivo, it is part of the complex and functions as a stabilizing element in vitro, The corresponding operon should now be referred to as kdpFABC.