A three-component dicamba O-demethylase from Pseudomonas maltophilia, strain DI-6: purification and characterization.
Dicamba O-demethylase is a multicomponent enzyme that catalyzes the conversion of the herbicide 2-methoxy-3,6-dichlorobenzoic acid (dicamba) to 3,6-dichlorosalicylic acid (DCSA). The three components of the enzyme were purified and characterized. Oxygenase(DIC) is a homotrimer (alpha)3 with a subunit molecular mass of approximately 40 kDa. FerredoxinDIC and reductaseDIC are monomers with molecular weights of approximately 14 and 45 kDa, respectively. EPR spectroscopic analysis suggested the presence of a single [2Fe-2S](2+/1+) cluster in ferredoxinDIC and a single Rieske [2Fe-2S](2+; 1+) cluster within oxygenaseDIC. Consistent with the presence of a Rieske iron-sulfur cluster, oxygenaseDIC displayed a high reduction potential of E(m,7.0) = -21 mV whereas ferredoxinDIC exhibited a reduction potential of approximately E(m,7.0) = -171 mV. Optimal oxygenaseDIC activity in vitro depended on the addition of Fe2+. The identification of formaldehyde and DCSA as reaction products demonstrated that dicamba O-demethylase acts as a monooxygenase. Taken together, these data suggest that oxygenaseDIC is an important new member of the Rieske non-heme iron family of oxygenases.
Chakraborty
Sarbani
S
Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE 68588-0664, USA.
Behrens
Mark
M
Herman
Patricia L
PL
Arendsen
Alexander F
AF
Hagen
Wilfred R
WR
Carlson
Deborah L
DL
Wang
Xiao-Zhuo
XZ
Weeks
Donald P
DP
D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification; D002723:Chlorobenzoates; D003996:Dicamba / Q000737:chemistry; D005288:Ferredoxins / Q000737:chemistry / Q000302:isolation & purification; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000302:isolation & purification; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000302:isolation & purification; D010084:Oxidation-Reduction; D010090:Oxidoreductases, O-Demethylating / Q000737:chemistry* / Q000302:isolation & purification; D020836:Protein Structure, Quaternary; D021122:Protein Subunits / Q000737:chemistry / Q000302:isolation & purification; D011549:Pseudomonas / Q000201:enzymology*; D012459:Salicylates / Q000737:chemistry
D001426:Bacterial Proteins; D002723:Chlorobenzoates; D005288:Ferredoxins; D009097:Multienzyme Complexes; D021122:Protein Subunits; D012459:Salicylates; C487853:3,6-dichlorosalicylic acid; D006899:Mixed Function Oxygenases; D010090:Oxidoreductases, O-Demethylating; D003996:Dicamba