Anaerobic chlorophyll isocyclic ring formation in Rhodobacter capsulatus requires a cobalamin cofactor
Anaerobic chlorophyll isocyclic ring formation in Rhodobacter capsulatus requires a cobalamin cofactor
复制标题
DOI：
10.1073/pnas.97.12.6908
复制
发表时间：
2000-06-06
影响因子：
11.1
通讯作者：
Duus, JO
中科院分区：
文献类型：
--
作者：
Gough, SP;Petersen, BO;Duus, JO
细菌叶绿素（BChl）的异环是由Mg-原卟啉单甲酯（MPE）转化为原叶绿素（PChlide）而形成的。通过BLAST搜索与假定的厌氧MPE-环化酶BchE的相似性表明，该蛋白质也使用钴胺素辅因子。我们发现，维生素B-12（B-12）需要突变体的bluE E和blue B基因的红细菌capsulatus，生长没有B-12，积累镁卟啉。激光解吸/电离飞行时间（LDI-TOF）MS和NMR光谱鉴定为MPE及其3-乙烯基-8-乙基（mvMPE）衍生物。设计了用于将MPE转化为PClide的环化酶的体内测定，在B-12依赖性突变体中的环化酶活性需要B-12而不是蛋白质合成。对于该MPE-环化酶反应提出了以下反应机理。腺苷钴胺素形成腺苷自由基，其导致氢原子的撤回和MPE的苄基型13（1）-自由基的形成。撤回电子产生MPE的13（1）-阳离子，羟基离子攻击阳离子产生13（1）-羟基-MPE。去掉三个氢原子后，依次生成13（1）-酮基-MPE及其13（2）-自由基，然后环化为PClide。
The isocyclic ring of bacteriochlorophyll (BChl) is formed by the conversion of Mg-protoporphyrin monomethyl ester (MPE) to protochlorophyllide (PChlide), Similarities revealed by BLAST searches with the putative anaerobic MPE-cyclase BchE suggested to us that this protein also uses a cobalamin cofactor. We found that vitamin B-12 (B-12)-requiring mutants of the bluE and bluB genes of Rhodobacter capsulatus, grown without B-12, accumulated Mg-porphyrins. Laser desorption/ionization time-of-flight (LDI-TOF) MS and NMR spectroscopy identified them as MPE and its 3-vinyl-8-ethyl (mvMPE) derivative. An in vivo assay was devised for the cyclase converting MPE to PChlide, Cyclase activity in the B-12-dependent mutants required B-12 but not protein synthesis. The following reaction mechanism is proposed for this MPE-cyclase reaction. Adenosylcobalamin forms the adenosyl radical, which leads to withdrawal of a hydrogen atom and formation of the benzylic-type 13(1)-radical of MPE, Withdrawal of an electron gives the 13(1)-cation of MPE, Hydroxyl ion attack on the cation gives 13(1)-hydroxy-MPE. Withdrawal of three hydrogen atoms leads successively to 13(1)-keto-MPE, its 13(2)-radical, and cyclization to PChlide.