Overexpression, purification and characterization of a new salicylate hydroxylase from naphthalene-degrading Pseudomonas sp strain ND6

A new salicytate hydroxylase from naphthalene-degrading Pseudomonas sp. strain ND6, NahU, has been identified. The nahU is an isofunctional gene of the classic saticylate hydroxylase gene, nahG, and situated outside the transcriptional unit forming the naphthalene degradation tower pathway. Both genes, nahU and nahG of Pseudomonas sp. ND6, have been cloned and overexpressed in Escherichia coli BL21 (DE3). NahU contains 429 amino acid residues and NahG contains 434 amino acid residues. SDS-PAGE analysis showed that both NahG and NahU are about 47kDa. Both enzymes exhibit broad substrate specificities and metabolize salicytate, sulfosaticylate, aspirin, methylsalicylate, chlorosalicylate and 3,5-dinitrosaticylate. The comparison of the K-m and V-max values for NahG and NahU demonstrated that NahU possesses a higher binding ability to salicylate and cofactors and catalytic efficiency. (c) 2005 Elsevier GmbH. All rights reserved.