Cloning and sequence analysis of a plasmid-encoded 2-haloacid dehalogenase gene from Pseudomonas putida No. 109

The 2-haloacid dehalogenase of Pseudomonas putida No. 109 was mediated by a 74-kb conjugative plasmid, which was transferred by mating into Pseudomonas and Escherichia coli and there expressed the dehalogenase. A 2.8-kb EcoRI-fragment generated from the plasmid was cloned and sequenced. The dehalogenase gene (dehH109) was identified by comparison with the N-terminal amino acid sequence and the molecular weight of the enzyme protein. The gene dehH109 coded for a 224-amino acid protein of M(r) 25,231, which showed significant homology to the other four L-specific 2-haloacid dehalogenases from Pseudomonas sp. CBS3, P. putida AJ1, and Xanthobacter autotrophicus GJ10 and also to the haloacetate dehalogenase H-2 from Moraxella sp. strain B, but no homology with another haloacetate dehalogenase H-1 and the D-specific 2-haloacid dehalogenase from P. putida AJ1.