Expression, purification and preliminary crystallographic studies of NahF, a salicylaldehyde dehydrogenase from Pseudomonas putida G7 involved in naphthalene degradation.
Pseudomonas putida G7 is one of the most studied naphthalene-degrading species. The nah operon in P. putida, which is present on the 83 kb metabolic plasmid NAH7, codes for enzymes involved in the conversion of naphthalene to salicylate. The enzyme NahF (salicylaldehyde dehydrogenase) catalyzes the last reaction in this pathway. The nahF gene was subcloned into the pET28a(TEV) vector and the recombinant protein was overexpressed in Escherichia coli Arctic Express at 285 K. The soluble protein was purified by affinity chromatography followed by gel filtration. Crystals of recombinant NahF (6×His-NahF) were obtained at 291 K and diffracted to 2.42 Å resolution. They belonged to the hexagonal space group P6(4)22, with unit-cell parameters a = b = 169.47, c = 157.94 Å. The asymmetric unit contained a monomer and a crystallographic twofold axis generated the dimeric biological unit.
Departamento de Bioquímica e Imunologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Avenida Antônio Carlos 6627, 31270-901 Belo Horizonte-MG, Brazil.
Juliana Barbosa
JB
Coitinho
Débora Maria Abrantes
DM
Costa
Samuel Leite
SL
Guimarães
Alfredo Miranda
AM
de Góes
Ronaldo Alves Pinto
RA
Nagem
D000445:Aldehyde Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D018360:Crystallography, X-Ray; D015870:Gene Expression; D009281:Naphthalenes / Q000378:metabolism; D016958:Pseudomonas putida / Q000201:enzymology*
D009281:Naphthalenes; C031721:naphthalene; D000445:Aldehyde Oxidoreductases; C024246:salicylaldehyde dehydrogenase