Visible light-sensitive mutants of Escherichia coli

我们分离到了对可见光敏感的E.ColiK-12突变体。这种现象是由编码铁络合酶的VISA(HemH)基因缺陷引起的。该基因的缺陷会导致原卟啉IX的积累，这是一种光敏剂，会产生一种对细胞有害的活性氧物种。我们从一株VISA缺失的菌株中分离出耐光性逆转株，发现第二个突变位于参与血红素生物合成途径的其他基因中，在铁络合酶催化反应之前的步骤中。利用这些突变体，得到以下结果：1)突变株VSR751在hemG基因产物原卟啉原氧化酶(PPO)中存在缺陷。通过互补分析，我们分离和鉴定了大肠杆菌的PPO基因。2)我们在hemA下游的一个新基因中发现了一个突变，命名为heme K。3)我们之前已经证明，E.colihemH突变体对蓝光的敏感度比其野生型亲本高100倍。在此，我们发现，血红素突变体AW823对蓝光的敏感度至少与hemH突变体一样高。4)通过筛选E.coli的hemG突变体的互补性，从构建于lambda载体中的拟南芥cDNA文库中获得了编码原卟啉原氧化酶的cDNA。5)我们发现，对血红素的饥饿引起了严格的反应，这种饥饿促进了卟啉的合成，而对HEMA-LacZ融合基因的表达没有任何影响。我们提出了一个卟啉合成的调控模型，在该模型中，卟啉的合成与蛋白质的合成是协调的。

We isolated the mutants of E.coli K-12 that were sensitive to visible light. This phenomenon was brought about by a defect in the visA (hemH) gene that encodes ferrochelatase. A defect in this gene causes the accumulation of protoporphyrin IX,which is a photosensitizer and produces an active species of oxygen that is harmful to the cell. We isolated photoresistant revertants from a visA-deleted strain and found that the second mutations were located in other genes involved in the heme biosynthetic pathway at steps prior to the reaction catalyzed by ferrochelatase. Utilizing those mutants, the following results were obtained :1) The mutant strain VSR751must be defective in protoporphyrinogen oxidase (PPO), the product of the hemG gene. By complementation analysis using this atrain we isolated and identified this PPO gene of E.coli.2) We found a mutation in a new gene downstream of hemA.This new gene, designated heme K,was analyzed.3) We showed previously that an E.coli hemH mutant is somewhat better than 100-fold more sensitive than its wild-type parent in tumbling to blue light. Here we found that the hemG mutant AW823 tumbles to blue light at least as sensitively as the hemH mutant.4) A cDNA encoding protoporphyrinogen oxidase, was obtained from a library of Arabidopsis thaliana cDNA constructed in a lambda vector by screening for complementation of a hemG mutant of E.coli.5) We found that starvation for heme caused the stringent response, and such starvation promote the synthesis of porphyrins without having any effect on the expression of the hemA-lacZ fusion gene. We suggested a model for the regulation of porphyrin synthesis wherein the synthesis of porphyrins is coordinated with that of proteins.