THE GENE FOR INDOLE-3-ACETYL-L-ASPARTIC ACID HYDROLASE FROM ENTEROBACTER-AGGLOMERANS - MOLECULAR-CLONING, NUCLEOTIDE-SEQUENCE, AND EXPRESSION IN ESCHERICHIA-COLI

A 5.5-kb DNA fragment containing the indole-3-acetyl-aspartic acid (IA A-asp) hydrolase gene (iaaspH) was isolated from Enterobacter agglomer ans strain GK12 using a hybridization probe based on the N-terminal am ino acid sequence of the protein. The DNA sequence of a 2.4-kb region of this fragment was determined and revealed a 1311-nucleotide ORF lar ge enough to encode the 45-kDa IAA-asp hydrolase. A 1.5-kb DNA fragmen t containing iaaspH was subcloned into the Escherichia coli expression plasmid pTTQ8 to yield plasmid pJCC2. Extracts of IPTG-induced E. col i cultures containing the pJCC2 recombinant plasmid showed IAA-asp hyd rolase levels 5 to 10-fold higher than those in E. agglomerans extract s. Homology searches revealed that the IAA-asp hydrolase was similar t o a variety of amidohydrolases. In addition, IAA-asp hydrolase showed 70% sequence identity to a putative thermostable carboxypeptidase of E . coli.