Nakai C, Kagamiyama H, Nozaki M, Nakazawa T, Inouye S, Ebina Y, Nakazawa A
J Biol Chem. 1983 Mar 10;258(5):2923-8.
Metapyrocatechase which catalyzes the oxygenative ring cleavage of catechol to form alpha-hydroxymuconic epsilon-semialdehyde is encoded by the xylE gene on the TOL plasmid of Pseudomonas putida mt-2. We have cloned the xylE region in Escherichia coli and determined the nucleotide sequence of the DNA fragment of 985 base pairs around the gene. The fragment included only one open translational frame of sufficient length to accommodate the enzyme. The predicted amino acid sequence consisted of 307 residues, and its NH2- and COOH-terminal sequences were in perfect agreement with those of the enzyme recently determined (Nakai, C., Hori, K., Kagamiyama, H., Nakazawa, T., and Nozaki, M. (1983) J. Biol. Chem. 258, 2916-2922). A mutant plasmid was isolated which did not direct the synthesis of the active enzyme. This plasmid had a DNA region corresponding to the NH2-terminal two-thirds of the polypeptide. From the deduced amino acid sequence, the secondary structure was predicted. Around 10 base pairs upstream from the initiator codon for metapyrocatechase, there was a base sequence which was complementary to the 3'-end of 16 S rRNAs from both E.coli and Pseudomonas aeruginosa. A preferential usage of C- and G-terminated codons was found in the coding region xylE, which contributed to the relatively high G + C content (57%) of this gene.
催化邻苯二酚的氧化环裂解以形成α-羟基粘康酸ε-半醛的间苯二酚酶由恶臭假单胞菌mt-2的TOL质粒上的xylE基因编码。我们已将xylE区域克隆到大肠杆菌中，并确定了该基因周围985个碱基对的DNA片段的核苷酸序列。该片段仅包含一个足够长的开放阅读框以容纳该酶。预测的氨基酸序列由307个残基组成，其NH2-末端和COOH-末端序列与最近确定的酶的序列完全一致（中井，C.，堀，K.，加美山，H.，中泽，T.和野崎，M.（1983）J.生物化学。258，2916 - 2922）。分离出一种不指导活性酶合成的突变质粒。该质粒具有与多肽NH2-末端三分之二相对应的DNA区域。根据推导的氨基酸序列预测了二级结构。在间苯二酚酶起始密码子上游约10个碱基对处，存在一个与大肠杆菌和铜绿假单胞菌16 S rRNA的3'-末端互补的碱基序列。在xylE编码区发现了C和G结尾密码子的优先使用，这导致了该基因相对较高的G + C含量（57%）。
