Negoro S, Kakudo S, Urabe I, Okada H
Department of Biotechnology, Osaka University, Japan.
J Bacteriol. 1992 Dec;174(24):7948-53. doi: 10.1128/jb.174.24.7948-7953.1992.
Flavobacterium sp. strain KI725 harbors plasmid pOAD21, a derivative of nylon oligomer-degradative plasmid pOAD2, in which all of nylA (the gene for 6-aminohexanoate cyclic dimer hydrolase [EI]) was deleted but nylB (the gene for 6-aminohexanoate dimer hydrolase [EII]) was retained. KI725 showed no growth on unfractionated nylon oligomers (Nom1) obtained from a nylon factory as a sole carbon and nitrogen source (Nom1 minimum plate). Extracts of KI725 cells possessed hydrolytic activity for Nom1 (approximately 5% of the activity of KI72), but pOAD2-cured strains (KI722 and KI723) showed no activity. KI725R strains which grew on the Nom1 minimum plate were spontaneously isolated from KI725 at a frequency of 10(-7) per cell. Activity toward Nom1 was enhanced in KI725R strains (10 to 30% of the activity of KI72). This new Nom1 degrading enzyme (EIII, the nylC gene product) hydrolyzed not only Nom1 but also the N-carbobenzoxy-6-aminohexanoate trimer, a substrate which was not hydrolyzed by either EI or EII. Cloning and sequence analysis showed that the nylC gene is located close to nylB on pOAD21 and is a 1,065-bp open reading frame corresponding to 355 amino acid residues. The nucleotide sequence of the nylC gene and the deduced amino acid sequence of EIII had no detectable homology with the sequences of nylA (EI) and nylB (EII).
黄杆菌属菌株KI725含有质粒pOAD21，它是尼龙低聚物降解性质粒pOAD2的衍生物，其中所有的nylA（6-氨基己酸环二聚体水解酶[EI]的基因）被删除，但nylB（6-氨基己酸二聚体水解酶[EII]的基因）被保留。KI725在从尼龙工厂获得的未分级尼龙低聚物（Nom1）作为唯一碳源和氮源的情况下（Nom1基本平板）无法生长。KI725细胞提取物对Nom1具有水解活性（约为KI72活性的5%），但pOAD2消除菌株（KI722和KI723）没有活性。能在Nom1基本平板上生长的KI725R菌株以每细胞10^(-7)的频率从KI725中自发分离得到。KI725R菌株对Nom1的活性增强（为KI72活性的10%至30%）。这种新的Nom1降解酶（EIII，nylC基因产物）不仅能水解Nom1，还能水解N-苄氧羰基-6-氨基己酸三聚体，这是一种EI和EII都不能水解的底物。克隆和序列分析表明，nylC基因位于pOAD21上靠近nylB的位置，是一个1065 bp的开放阅读框，对应355个氨基酸残基。nylC基因的核苷酸序列和EIII推导的氨基酸序列与nylA（EI）和nylB（EII）的序列没有可检测到的同源性。
