Galvão Teca Calcagno, Mencía Mario, de Lorenzo Víctor
Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Darwin 3, Cantoblanco, 28049 Madrid, Spain.
Mol Microbiol. 2007 Aug;65(4):907-19. doi: 10.1111/j.1365-2958.2007.05832.x. Epub 2007 Jul 21.
Evolutionary expansion of metabolic networks entails the emergence of regulatory factors that become sensitive to new chemical species. A dedicated genetic system was developed for the soil bacterium Pseudomonas putida aimed at deciphering the steps involved in the gain of responsiveness of the toluene-activated prokaryotic regulator XylR to the xenobiotic chemical 2,4 dinitrotoluene (DNT). A mutant library of the A domain of XylR was screened in vivo for those variants activated by DNT through coupling the cognate promoter Pu to the P. putida yeast URA3 homologue, pyrF. All DNT-responsive clones maintained their sensitivity to ordinary effectors of XylR and broadened the range of inducers to unrelated aromatics. Yet, none of the altered amino acids lay in the recognizable effector binding pocket of the polypeptide. Instead, mutations appeared in protein surfaces believed to engage in the conformational shifts that follow effector binding and modulate signal transmission between XylR domains. It thus seems that transcriptional factors are likely to regress into functionally multipotent forms (i.e. stem protein types) as a first step towards the divergence of a new specificity.
代谢网络的进化扩张需要出现对新化学物质敏感的调控因子。针对土壤细菌恶臭假单胞菌开发了一个专门的遗传系统，旨在解析甲苯激活的原核调节因子XylR对异源化学物质2,4-二硝基甲苯（DNT）产生响应的过程中所涉及的步骤。通过将同源启动子Pu与恶臭假单胞菌酵母URA3同源物pyrF偶联，在体内筛选XylR A结构域的突变文库，以寻找被DNT激活的变体。所有对DNT有响应的克隆都保持了对XylR普通效应物的敏感性，并将诱导剂范围扩大到不相关的芳烃。然而，没有一个氨基酸的改变位于多肽可识别的效应物结合口袋中。相反，突变出现在被认为参与效应物结合后构象变化并调节XylR结构域之间信号传递的蛋白质表面。因此，转录因子似乎可能会退化为功能多能形式（即主干蛋白类型），作为迈向新特异性分化的第一步。
