Proteomic analysis of the benzoate degradation pathway in Acinetobacter sp. KS-1.
The purpose of this study was to perform proteome analysis of Acinetobacter sp. KS-1, a bacterium capable of degrading benzoate as a sole carbon source. In order to understand the benzoate degradation pathway used by strain KS-1, proteomes of benzoate-cultured and succinate-cultured KS-1 were comparatively analyzed by two dimensional gel electrophoresis (2-DE). Eighteen protein spots proteins were exclusively induced from the benzoate-cultured strain KS-1. Of these 18 spots, two benzoate-degrading enzymes (catechol 1,2-dioxygenase and beta-ketoadipate succinyl-CoA transferase) were identified by MS/MS analysis by MALDI-TOF/TOF mass spectrometry, which suggests that strain KS-1 degrades benzoate by the beta-ketoadipate pathway. DEAE-chromatography suggested that strain KS-1 induced only one type of catechol 1,2-dioxygenase during benzoate degradation. The catechol 1,2-dioxygenase was purified using three steps of ammonium sulfate precipitation, DEAE-sepharose, and Mono-Q chromatography. The purified catechol 1,2-dioxygenase of strain KS-1 had strong dioxygenase activity for 4-methylcatechol as well as catechol. Sequencing analysis using N-terminal and internal amino acid sequences showed that this catechol 1,2-dioxygenase is highly homologous with catechol 1,2-dioxygenase of Acinetobacter radioresistens. These results suggest that comparative proteomic analysis of biodegrading bacteria cultured under different conditions may be a useful initial step toward the elucidation of the aromatic compound degradation pathway.
Kim
Seung Il
SI
Proteome Analysis Team, Korea Basic Science Institute, 305-333, Daejon, South Korea.
Song
Seung Youl
SY
Kim
Kyung Wook
KW
Ho
Eun Mi
EM
Oh
Kye Heon
KH
D000150:Acinetobacter / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001565:Benzoates / Q000378:metabolism*; D001673:Biodegradation, Environmental; D050548:Catechol 1,2-Dioxygenase; D049308:Dioxygenases; D015180:Electrophoresis, Gel, Two-Dimensional; D008969:Molecular Sequence Data; D010105:Oxygenases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D020543:Proteome; D017422:Sequence Analysis, DNA; D019802:Succinic Acid / Q000378:metabolism
D001426:Bacterial Proteins; D001565:Benzoates; D010455:Peptides; D020543:Proteome; D019802:Succinic Acid; D010105:Oxygenases; D049308:Dioxygenases; D050548:Catechol 1,2-Dioxygenase