A three-component dicamba O-demethylase from Pseudomonas maltophilia, strain DI-6: Purification and characterization

Chakraborty, Sarbani; Behrens, Mark; Herman, Patricia L.; Arendsen, A.F.; Hagen, Wilfred R.; Carlson, Deborah; Wang, Xiao Zhuo; Weeks, Donald P.

doi:10.1016/j.abb.2005.02.024

Cited by 28 publications

(36 citation statements)

References 28 publications

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“…Cloning of the Reductase Genes-The first 23 residues of the Nterminal sequence was determined for the purified reductase DIC protein (Table I) as described previously (10). A comparison of this sequence to the GenBank TM data base showed that it was 90% identical in a 20-amino acid overlap to the cytochrome P450-type reductase component of dioxin dioxygenase, a three-component enzyme previously isolated from Sphingomonas sp.…”

Section: Methodsmentioning

confidence: 99%

“…Cloning of the Ferredoxin Gene-The first 29 residues of the Nterminal amino acid sequence was determined for the purified ferredoxin DIC protein (Table I) as described previously (10). A comparison of this sequence to the GenBank data base showed that it was 35% identical in a 26-amino acid overlap to a terpredoxin from a Pseudomonas species, a [2Fe-2S] ferredoxin in the adrenodoxin family (14).…”

Section: Methodsmentioning

confidence: 99%

“…Amino Acid Sequencing-The purification of the reductase DIC , ferredoxin DIC , and oxygenase DIC components of dicamba O-demethylase as well as the N-terminal amino acid sequences of the purified proteins have been described (10). To obtain internal amino acid sequence information, the purified ferredoxin DIC and reductase DIC proteins were digested with trypsin and the isolated peptide fragments were sequenced by automated Edman degradation in the Protein Core Facility (Center for Biotechnology, University of Nebraska-Lincoln).…”

Section: Methodsmentioning

confidence: 99%

“…Cloning of the Oxygenase Gene-The first 27 residues of the Nterminal amino acid sequence was determined for the purified oxygenase DIC protein (Table I) as described previously (10). To clone the oxygenase gene, a degenerate oligonucleotide (17-mer, 32 variants) based on the sequence NAWYVA (Table I) was designed and synthesized.…”

Section: Methodsmentioning

confidence: 99%

“…We also partially purified the enzyme and found that at least three separate components are required for activity (9). Recently, we provided a detailed description of the purification and characterization of the reductase DIC , ferredoxin DIC , and oxygenase DIC components of dicamba O-demethylase (10). Oxygenase DIC is a homotrimer (␣) 3 with a subunit molecular mass of ϳ40 kDa and contains a single Rieske [2Fe-2S] cluster.…”

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confidence: 99%

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A Three-component Dicamba O-Demethylase from Pseudomonas maltophilia, Strain DI-6

Herman¹,

Behrens

Chakraborty

et al. 2005

Journal of Biological Chemistry

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118

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Dicamba O-demethylase is a multicomponent enzyme from Pseudomonas maltophilia, strain DI-6, that catalyzes the conversion of the widely used herbicide dicamba (2-methoxy-3,6-dichlorobenzoic acid) to DCSA (3,6-dichlorosalicylic acid). We recently described the biochemical characteristics of the three components of this enzyme (i.e. reductase DIC , ferredoxin DIC , and oxygenase DIC ) and classified the oxygenase component of dicamba O-demethylase as a member of the Rieske nonheme iron family of oxygenases. In the current study, we used N-terminal and internal amino acid sequence information from the purified proteins to clone the genes that encode dicamba O-demethylase. Two reductase genes (ddmA1 and ddmA2) with predicted amino acid sequences of 408 and 409 residues were identified. The open reading frames encode 43.7-and 43.9-kDa proteins that are 99.3% identical to each other and homologous to members of the FAD-dependent pyridine nucleotide reductase family. The ferredoxin coding sequence (ddmB) specifies an 11.4-kDa protein composed of 105 residues with similarity to the adrenodoxin family of [2Fe-2S] bacterial ferredoxins. The oxygenase gene (ddmC) encodes a 37.3-kDa protein composed of 339 amino acids that is homologous to members of the Phthalate family of Rieske non-heme iron oxygenases that function as monooxygenases. Southern analysis localized the oxygenase gene to a megaplasmid in cells of P. maltophilia. Mixtures of the three highly purified recombinant dicamba O-demethylase components overexpressed in Escherichia coli converted dicamba to DCSA with an efficiency similar to that of the native enzyme, suggesting that all of the components required for optimal enzymatic activity have been identified. Computer modeling suggests that oxygenase DIC has strong similarities with the core ␣ subunits of naphthalene 1,2-dioxygenase. Nonetheless, the present studies point to dicamba O-demethylase as an enzyme system with its own unique combination of characteristics.The herbicide dicamba (2-methoxy-3,6-dichlorobenzoic acid) has been used to effectively control broadleaf weeds in crops such as corn and wheat for almost 40 years. Like a number of other chlorinated organic compounds, dicamba does not persist in the soil because it is efficiently metabolized by a consortium of soil bacteria under both aerobic and anaerobic conditions (1-4). Studies with different soil types treated with dicamba have demonstrated that 3,6-dichlorosalicylic acid (DCSA), 1 a compound without herbicidal activity, is a major product of the microbial degradation process (2, 3, 5). Soil samples taken from a single site exposed to dicamba for several years yielded a number of bacterial species capable of utilizing dicamba as a sole carbon source (6). These soil microorganisms could completely mineralize dicamba to carbon dioxide, water, and chloride ion (7). Studies on the metabolism of dicamba in the cells of one of these bacteria, the DI-6 strain of Pseudomonas maltophilia, showed that DCSA is a major degradation product (7,8).We have be...

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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A Three-component Dicamba O-Demethylase from Pseudomonas maltophilia, Strain DI-6

Herman¹,

Behrens

Chakraborty

et al. 2005

Journal of Biological Chemistry

Self Cite

118

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Glyphosate‐Resistant Crops: Developing the Next Generation Products

Feng

CaJacob

Martino-Catt

et al. 2010

Glyphosate Resistance in Crops and Weeds

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Rieske Oxygenases and Other Ferredoxin‐Dependent Enzymes: Electron Transfer Principles and Catalytic Capabilities

2023

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Enzymes that depend on sophisticated electron transfer via ferredoxins (Fds) exhibit outstanding catalytic capabilities, but despite decades of research, many of them are still not well understood or exploited for synthetic applications. This review aims to provide a general overview of the most important Fddependent enzymes and the electron transfer processes involved. While several examples are discussed, we focus in particular on the family of Rieske non-heme iron-dependent oxygenases (ROs). In addition to illustrating their electron transfer principles and catalytic potential, the current state of knowledge on structure-function relationships and the mode of interaction between the redox partner proteins is reviewed. Moreover, we highlight several key catalyzed transformations, but also take a deeper dive into their engineerability for biocatalytic applications. The overall findings from these case studies highlight the catalytic capabilities of these biocatalysts and could stimulate future interest in developing additional Fddependent enzyme classes for synthetic applications.

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A three-component dicamba O-demethylase from Pseudomonas maltophilia, strain DI-6: Purification and characterization

Cited by 28 publications

References 28 publications

A Three-component Dicamba O-Demethylase from Pseudomonas maltophilia, Strain DI-6

A Three-component Dicamba O-Demethylase from Pseudomonas maltophilia, Strain DI-6

Glyphosate‐Resistant Crops: Developing the Next Generation Products

Rieske Oxygenases and Other Ferredoxin‐Dependent Enzymes: Electron Transfer Principles and Catalytic Capabilities

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