Shapiro A B, Huber A H, McCarty R E
Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853.
J Biol Chem. 1991 Mar 5;266(7):4194-200.
We have examined the properties of the four tight nucleotide binding sites of reductively activated chloroplast coupling factor 1. Tight sites are here defined as those which retain bound nucleotides after passage of the chloroplast coupling factor 1 through Sephadex gel filtration centrifuge columns. Two of the sites, here called sites 4 and 5, have not been characterized in detail before. Site 4 has properties similar to those of site 1. It binds to ADP, ATP, and adenylyl-beta,gamma-imidodiphosphate (AMP-PNP) tightly in the presence or absence of Mg2+. Bound ADP exchanges rapidly with medium ADP, but rapid exchange with ATP or AMP-PNP requires Mg2+. Site 4 may slowly hydrolyze bound ATP in the absence of medium nucleotides. Site 5 has properties similar to those of site 2. Tight binding of ATP and AMP-PNP requires Mg2+, but Mg29+)-ADP is not tightly bound. Site 5 does not hydrolyze bound ATP in the absence of medium nucleotides. Complete filling of all four tight nucleotide binding sites requires about one millimolar nucleotide, suggesting that low affinity binding sites are converted to tight binding via a nucleotide binding-induced conformational change.
我们研究了还原激活的叶绿体偶联因子1的四个紧密核苷酸结合位点的特性。紧密位点在此定义为叶绿体偶联因子1通过葡聚糖凝胶过滤离心柱后仍保留结合核苷酸的那些位点。其中两个位点，这里称为位点4和位点5，以前尚未进行详细表征。位点4具有与位点1相似的特性。在存在或不存在Mg2+的情况下，它都能紧密结合ADP、ATP和腺苷-β,γ-亚氨基二磷酸（AMP-PNP）。结合的ADP能与介质中的ADP快速交换，但与ATP或AMP-PNP的快速交换需要Mg2+。在位点4不存在介质核苷酸的情况下，可能会缓慢水解结合的ATP。位点5具有与位点2相似的特性。ATP和AMP-PNP的紧密结合需要Mg2+，但Mg2+(9+) -ADP不能紧密结合。在位点5不存在介质核苷酸的情况下，不会水解结合的ATP。完全填满所有四个紧密核苷酸结合位点大约需要1毫摩尔核苷酸，这表明低亲和力结合位点通过核苷酸结合诱导的构象变化转化为紧密结合。
