Overexpression, purification and characterization of a new salicylate hydroxylase from naphthalene-degrading Pseudomonas sp. strain ND6.
A new salicylate hydroxylase from naphthalene-degrading Pseudomonas sp. strain ND6, NahU, has been identified. The nahU is an isofunctional gene of the classic salicylate hydroxylase gene, nahG, and situated outside the transcriptional unit forming the naphthalene degradation lower 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 47 kDa. Both enzymes exhibit broad substrate specificities and metabolize salicylate, sulfosalicylate, aspirin, methylsalicylate, chlorosalicylate and 3,5-dinitrosalicylate. The comparison of the Km and Vmax values for NahG and NahU demonstrated that NahU possesses a higher binding ability to salicylate and cofactors and catalytic efficiency.
Zhao
Huabing
H
Department of Microbiology, Nankai University, Tianjin 300071, China.
Chen
Defu
D
Li
Yongjun
Y
Cai
Baoli
B
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001673:Biodegradation, Environmental; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D007700:Kinetics; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D009281:Naphthalenes / Q000378:metabolism*; D010957:Plasmids; D011549:Pseudomonas / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D009281:Naphthalenes; D006899:Mixed Function Oxygenases; C022619:salicylate 1-monooxygenase