Shimamoto T, Shimamoto T, Xu X J, Okazaki N, Kawakami H, Tsuchiya T
Faculty of Applied Biological Science, Hiroshima University, Kagamiyama, Higashi-Hiroshima 739-8528, Japan.
J Biochem. 2001 Apr;129(4):607-13. doi: 10.1093/oxfordjournals.jbchem.a002897.
Wild-type Citrobacter freundii cannot grow on melibiose as a sole source of carbon. The melibiose transporter gene melB was cloned from a C. freundii mutant M4 that could utilize melibiose as a sole carbon source. Although the cloned melB gene is closely similar to the melB genes of other bacteria, it is cryptic because of a frameshift mutation. Site-directed mutagenesis was used to construct a functional melB gene by deleting one nucleotide, resulting in the production of an active melibiose transporter. The active MelB transporter could utilize Na(+) and H(+) as coupling cations to melibiose transport. The amino acid sequence of the C. freundii MelB was found to be most similar to those of Salmonella typhimurium and Escherichia coli MelB. These facts are consistent with the phylogenetic relationship of bacteria and the cation coupling properties of the melibiose transporters.
野生型弗氏柠檬酸杆菌不能以蜜二糖作为唯一碳源生长。蜜二糖转运基因melB是从一株能够利用蜜二糖作为唯一碳源的弗氏柠檬酸杆菌突变体M4中克隆得到的。尽管克隆得到的melB基因与其他细菌的melB基因高度相似，但由于一个移码突变，它是无活性的。通过定点诱变删除一个核苷酸构建了一个功能性的melB基因，从而产生了一种活性蜜二糖转运蛋白。活性MelB转运蛋白可以利用Na(+)和H(+)作为与蜜二糖转运偶联的阳离子。发现弗氏柠檬酸杆菌MelB的氨基酸序列与鼠伤寒沙门氏菌和大肠杆菌MelB的氨基酸序列最为相似。这些事实与细菌的系统发育关系以及蜜二糖转运蛋白的阳离子偶联特性是一致的。
