Weston M F, Kotake S, Davis L C
Arch Biochem Biophys. 1983 Sep;225(2):809-17. doi: 10.1016/0003-9861(83)90093-0.
The interaction of a large number of ATP and ADP analogs with nitrogenase from Azotobacter vinelandii, Klebsiella pneumoniae, and Clostridium pasteurianum has been examined. Only 1,N6-etheno-ATP and 2'-deoxy-ATP served as substrates for acetylene reduction. Other triphosphates including GTP, ITP, 8-Br-ATP, alpha,beta-methylene ATP, beta,gamma-methylene ATP, 6-chloropurine riboside triphosphate, and AMP-PNP were inert, showing less than 50% inhibition at levels up to two- to fivefold greater than ATP. Xanthosine triphosphate behaved simply as a chelator of magnesium, activating the enzyme at low levels but strongly inhibiting at high levels. When nucleotide diphosphates were tested as inhibitors with enzyme from A. vinelandii, GDP, dGDP, and 6-chloropurine riboside diphosphate were ineffective, XDP was three- to fivefold less effective, and dADP and 1,N6-etheno-ADP were about equally as effective as ADP. With enzyme from C. pasteurianum, dADP was twofold less effective than ADP, XDP was fivefold less effective, and IDP and 1,N6-etheno-ADP appeared to be ineffective. Results with enzyme from K. pneumoniae were very similar to those obtained with A. vinelandii. Different metal ions were tested in the presence of both ATP and ADP to determine whether preferential binding to one nucleotide or the other might alter the ADP/ATP ratio needed for 50% inhibition of activity. Magnesium and manganese gave the same ratio, while with Fe and Co, slightly less ADP was required for equivalent inhibition. Nickel appeared to reduce the sensitivity of A. vinelandii nitrogenase to ADP inhibition while increasing that of C. pasteurianum, but both effects were less than twofold. Calcium, strontium, and aluminum ions were inert with enzymes from these organisms. Cd and Zn were also ineffective with K. pneumoniae. Two isomers of ATP beta S were prepared by enzymatic synthesis from ADP beta S. The A form was a more potent inhibitor of A. vinelandii nitrogenase.
研究了大量ATP和ADP类似物与棕色固氮菌、肺炎克雷伯菌和巴氏梭菌的固氮酶之间的相互作用。只有1,N6-乙烯基-ATP和2'-脱氧-ATP可作为乙炔还原的底物。其他三磷酸酯，包括GTP、ITP、8-溴-ATP、α,β-亚甲基ATP、β,γ-亚甲基ATP、6-氯嘌呤核糖三磷酸酯和AMP-PNP均无活性，在浓度比ATP高两到五倍时，抑制作用小于50%。黄嘌呤三磷酸酯仅表现为镁的螯合剂，在低浓度时激活酶，但在高浓度时强烈抑制酶。当测试核苷酸二磷酸酯对棕色固氮菌酶的抑制作用时，GDP、dGDP和6-氯嘌呤核糖二磷酸酯无效，XDP的效力低三到五倍，dADP和1,N6-乙烯基-ADP的效力与ADP大致相同。对于巴氏梭菌的酶，dADP的效力比ADP低两倍，XDP的效力低五倍，IDP和1,N6-乙烯基-ADP似乎无效。肺炎克雷伯菌酶的结果与棕色固氮菌的非常相似。在ATP和ADP存在的情况下测试了不同的金属离子，以确定与一种核苷酸或另一种核苷酸的优先结合是否会改变抑制50%活性所需的ADP/ATP比率。镁和锰给出相同的比率，而对于铁和钴，等效抑制所需的ADP略少。镍似乎降低了棕色固氮菌固氮酶对ADP抑制的敏感性，同时增加了巴氏梭菌的敏感性，但两种效应均小于两倍。钙、锶和铝离子对这些生物体的酶无活性。镉和锌对肺炎克雷伯菌也无效。通过从ADP-β-S进行酶促合成制备了ATP-β-S的两种异构体。A形式是棕色固氮菌固氮酶更有效的抑制剂。
