Pitterle D M, Rajagopalan K V
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710.
J Bacteriol. 1989 Jun;171(6):3373-8. doi: 10.1128/jb.171.6.3373-3378.1989.
Molybdopterin (MPT) is not produced by the Escherichia coli mutants chlA1, chlM, or chlN or by the Neurospora crassa mutant nit-1. Extracts of E. coli chlA1 contain an activity, the converting factor, which is functionally defined by its ability to convert a low-molecular-weight precursor present in crude extracts of N. crassa nit-1 into molybdopterin in vitro. In this study, it has been shown that the converting factor consists of two associative proteins (10 and 25 kilodaltons [kDa]) which can be separated by using either anion-exchange or gel filtration chromatography. Neither protein is able to complement extracts of nit-1 by itself. Analysis of chlA Mu insertion mutants has shown that the two proteins are distinct gene products encoded at the chlA locus. Twelve chlA Mu insertion strains which lacked converting factor activity were deficient in one or both of the proteins. Converting factor activity could be generated by mixing extracts from strains having the 25-kDa protein with those having the 10-kDa protein but not those lacking both proteins. Finally, it was shown that the chlM mutant lacks the 10-kDa protein while the chlN mutant, which contains both the 10- and 25-kDa proteins, lacks a function required to activate the 10-kDa protein.
钼蝶呤（MPT）不是由大肠杆菌突变体chlA1、chlM或chlN产生的，也不是由粗糙脉孢菌突变体nit-1产生的。大肠杆菌chlA1提取物含有一种活性物质，即转化因子，其功能定义为能够在体外将粗糙脉孢菌nit-1粗提取物中存在的低分子量前体转化为钼蝶呤。在本研究中，已表明转化因子由两种缔合蛋白（10和25千道尔顿[kDa]）组成，可通过阴离子交换或凝胶过滤色谱法分离。这两种蛋白单独都不能补充nit-1的提取物。对chlA Mu插入突变体的分析表明，这两种蛋白是在chlA基因座编码的不同基因产物。十二个缺乏转化因子活性的chlA Mu插入菌株在一种或两种蛋白上存在缺陷。通过将具有25 kDa蛋白的菌株提取物与具有10 kDa蛋白的菌株提取物混合可产生转化因子活性，但与两种蛋白都缺乏的菌株提取物混合则不能产生。最后，表明chlM突变体缺乏10 kDa蛋白，而同时含有10 kDa和25 kDa蛋白的chlN突变体缺乏激活10 kDa蛋白所需的功能。
