Nichols B P, Green J M
Department of Biological Sciences, University of Illinois, Chicago 60680.
J Bacteriol. 1992 Aug;174(16):5309-16. doi: 10.1128/jb.174.16.5309-5316.1992.
In Escherichia coli, chorismate lyase catalyzes the first step in ubiquinone biosynthesis, the conversion of chorismate to 4-hydroxybenzoate. 4-Hydroxybenzoate is converted to 3-octaprenyl-4-hydroxybenzoate by 4-hydroxybenzoate octaprenyltransferase. These two enzymes are encoded by ubiC and ubiA, respectively, and have been reported to map near one another at 92 min on the E. coli chromosome. We have cloned the ubiCA gene cluster and determined the nucleotide sequence of ubiC and a portion of ubiA. The nucleotide sequence abuts with a previously determined sequence that encodes a large portion of ubiA. ubiC was localized by subcloning, and overproducing plasmids were constructed. Overexpression of ubiC allowed the purification of chorismate lyase to homogeneity, and N-terminal sequence analysis of chorismate lyase unambiguously defined the beginning of the ubiC coding region. Although chorismate lyase showed no significant amino acid sequence similarity to 4-amino-4-deoxychorismate lyase (4-amino-4-deoxychroismate----4-aminobenzoate), the product of E. coli pabC, chorismate lyase overproduction could complement the growth requirement for 4-aminobenzoate of a pabC mutant strain. Of the several enzymes that convert chorismate to intermediates of E. coli biosynthetic pathways, chorismate lyase is the last to be isolated and characterized.
在大肠杆菌中，分支酸裂解酶催化泛醌生物合成的第一步，即将分支酸转化为4-羟基苯甲酸。4-羟基苯甲酸通过4-羟基苯甲酸辛酯转移酶转化为3-辛酯基-4-羟基苯甲酸。这两种酶分别由ubiC和ubiA编码，据报道它们在大肠杆菌染色体上92分钟处彼此相邻定位。我们克隆了ubiCA基因簇，并确定了ubiC和ubiA一部分的核苷酸序列。该核苷酸序列与先前确定的编码ubiA大部分的序列邻接。通过亚克隆对ubiC进行了定位，并构建了过量表达质粒。ubiC的过表达使得能够将分支酸裂解酶纯化至同质，并且对分支酸裂解酶的N端序列分析明确界定了ubiC编码区的起始位置。尽管分支酸裂解酶与4-氨基-4-脱氧分支酸裂解酶（4-氨基-4-脱氧分支酸----4-氨基苯甲酸）没有明显的氨基酸序列相似性，后者是大肠杆菌pabC的产物，但分支酸裂解酶的过量表达可以补充pabC突变菌株对4-氨基苯甲酸的生长需求。在将分支酸转化为大肠杆菌生物合成途径中间体的几种酶中，分支酸裂解酶是最后一个被分离和表征的。
