Cloning and analysis of the leuB gene of Leptospira interrogans serovar pomona.
Cloning and analysis of the leuB gene of Leptospira interrogans serovar pomona.
复制标题
问号钩端螺旋体波莫纳血清leuB基因的克隆与分析
DOI：
10.1099/00221287-139-5-1093
复制
发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Yelton,DB
中科院分区：
文献类型：
--
作者：
Ding,M;Yelton,DB
通过与大肠杆菌leuB突变体的互补，将问号钩端螺旋体血清变种pomonastrein kenniwicki的leuB基因克隆到9·5 kb的质粒pWVL 1上。亚克隆和Tn 5突变表明，互补所需的区域长度约为1.2kb。酶活性测定结果表明，该基因的产物为β-异丙基苹果酸脱氢酶。E. pWVL 1不与大肠杆菌互补。已经确定了theleuB互补区域和周围DNA的核苷酸序列。发现了3个开放阅读框，分别编码40·9、38·8和15 kDa蛋白。对含有不同大小的核苷酸缺失的亚克隆的分析表明，只有38·8 kDa的蛋白质是获得E. colileuB突变。检索PIR数据库，发现来自6种不同微生物的3-异丙基苹果酸脱氢酶与38·8 kDaL具有显著的氨基酸序列相似性（43-57%）。问号亮氨酸基因产物。L.亮氨酸生物合成基因的组织。疑问句与E.大肠杆菌、鼠伤寒沙门氏菌和枯草芽孢杆菌。
TheleuBgene ofLeptospira interrogansserovarpomonastrain kenniwicki has been cloned on a 9·5 kb plasmid, pWVL1, by complementation ofEscherichia coli leuBmutants. Subcloning and Tn5mutagenesis showed that the region required for complementation was approximately 1·2 kb in length. Enzyme assays showed that the product of the cloned gene was aβ-isopropylmalate dehydrogenase. Defects in theleuA, leuCandleuDgenes ofE. coliwere not complemented by pWVL1. The nucleotide sequence of theleuB-complementing region and surrounding DNA has been determined. Three open reading frames were found which encode proteins of 40·9, 38·8 and 15 kDa. Analysis of subclones containing nucleotide deletions of varying sizes showed that only the 38·8 kDa protein was necessary to obtain complementation ofE. coli leuBmutations. The PIR data base was searched and the enzyme 3-isopropylmalate dehydrogenase from six different micro-organisms was found to share significant amino acid sequence similarity (43–57%) with the 38·8 kDaL. interrogans leuBgene product. The organization of the leucine biosynthetic genes inL. interrogansdiffers from that found inE. coli, Salmonella typhimuriumandBacillus subtilis.