Compound 800, a natural product isolated from genetically engineered Pseudomonas: proposed structure, reactivity, and putative relation to heme d1.
Compound 800, a natural product isolated from genetically engineered Pseudomonas: proposed structure, reactivity, and putative relation to heme d1.
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化合物 800，一种从基因工程假单胞菌中分离出来的天然产物：提出的结构、反应性以及与血红素 d1 的假定关系。
DOI：
10.1021/bi0491954
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Timkovich,Russell
中科院分区：
文献类型：
--
作者：
Youn,Hyung-Sun;Liang,Qiaoli;Cha,JinK;Cai,Mengli;Timkovich,Russell
将斯氏假单胞菌Zobell亚株的FDLGH基因簇转移到高拷贝质粒上，制备了大肠杆菌和铜绿假单胞菌的基因工程菌株。这些基因以前被认为对血红素D1的生物合成是必不可少的，血红素D1是厌氧反硝化细菌中异化亚硝酸还原酶的辅助组。从这两种生物中鉴定出在稳态水平上可检测到的四氢吡咯，并使用每种生物的无细胞提取物在体外转化尿卟啉原。大肠埃希菌不能自然产生d1，工程菌也不能产生d1或任何四吡咯的异物脚趾。Coli.因此，虽然irFDLGH可能是d1生物合成所必需的，但这是不够的。在反硝化炉中。铜绿假单胞菌阳性率较高。该质粒的存在导致了D1表达水平的增加。此外，还检测到了一种以前未鉴定的四吡咯。用可见光吸收光谱、红外光谱、X-射线光电子能谱、质谱学、核磁共振等方法对该化合物进行了表征，并提出了该化合物的初步结构。四吡咯的结构特征类似于西罗盐酸盐(作为前科林-2或西罗四氯化物，已知的d1的中间体)和d1本身。最不常见的取代基是环氧化物和亚砜基团。当用强无机酸和热处理该四吡咯时，它被转化为天然的D_1。
Genetically engineered strains ofEscherichia coliandPseudomonas aeruginosawere prepared harboring the gene clusternirFDLGHfromPseudomonas stutzerisubstrain ZoBell on a high copy plasmid. These genes have been previously implicated as being essential for the biosynthesis of heme d1, the prosthetic group of dissimilatory nitrite reductases in anaerobic, denitryfying bacteria. Tetrapyrroles detectable at steady-state levels were identified from both organisms, and cell-free extracts from each were also used to transform uroporphyrinogen in vitro.E. colidoes not naturally produce d1, and the engineered strain failed to produce d1or any tetrapyrrole foreign toE. coli. Therefore, whilenirFDLGHmay be necessary for d1biosynthesis, it is not sufficient. In the denitrifierP. aeruginosa, the results were more positive. The presence of the plasmid led to increased levels of d1. In addition, a previously unidentified tetrapyrrole was detected. This compound was characterized by visible absorption spectroscopy, infrared spectroscopy, X-ray photoelectron spectroscopy, mass spectrometry, and NMR, and a tentative structure was proposed for this compound. The tetrapyrrole has structural features similar to sirohydrochlorin (as precorrin-2 or sirotetrahydrochlorin, a known intermediate of d1) and d1itself. The most unusual substituents are epoxide and sulfoxide moieties. When this tetrapyrrole was treated with strong mineral acid and heat, it was converted into natural d1.