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Cyclohexane-1,2-Dione Hydrolase from Denitrifying Azoarcus sp. Strain 22Lin, a Novel Member of the Thiamine Diphosphate Enzyme Family
- Source :
- Journal of Bacteriology
- Publication Year :
- 2011
- Publisher :
- American Society for Microbiology, 2011.
-
Abstract
- Alicyclic compounds with hydroxyl groups represent common structures in numerous natural compounds, such as terpenes and steroids. Their degradation by microorganisms in the absence of dioxygen may involve a C—C bond ring cleavage to form an aliphatic intermediate that can be further oxidized. The cyclohexane-1,2-dione hydrolase (CDH) (EC 3.7.1.11) from denitrifying Azoarcus sp. strain 22Lin, grown on cyclohexane-1,2-diol as a sole electron donor and carbon source, is the first thiamine diphosphate (ThDP)-dependent enzyme characterized to date that cleaves a cyclic aliphatic compound. The degradation of cyclohexane-1,2-dione (CDO) to 6-oxohexanoate comprises the cleavage of a C—C bond adjacent to a carbonyl group, a typical feature of reactions catalyzed by ThDP-dependent enzymes. In the subsequent NAD + -dependent reaction, 6-oxohexanoate is oxidized to adipate. CDH has been purified to homogeneity by the criteria of gel electrophoresis (a single band at ∼59 kDa; calculated molecular mass, 64.5 kDa); in solution, the enzyme is a homodimer (∼105 kDa; gel filtration). As isolated, CDH contains 0.8 ± 0.05 ThDP, 1.0 ± 0.02 Mg 2+ , and 1.0 ± 0.015 flavin adenine dinucleotide (FAD) per monomer as a second organic cofactor, the role of which remains unclear. Strong reductants, Ti(III)-citrate, Na + -dithionite, and the photochemical 5-deazaflavin/oxalate system, led to a partial reduction of the FAD chromophore. The cleavage product of CDO, 6-oxohexanoate, was also a substrate; the corresponding cyclic 1,3- and 1,4-diones did not react with CDH, nor did the cis - and trans -cyclohexane diols. The enzymes acetohydroxyacid synthase (AHAS) from Saccharomyces cerevisiae , pyruvate oxidase (POX) from Lactobacillus plantarum , benzoylformate decarboxylase from Pseudomonas putida , and pyruvate decarboxylase from Zymomonas mobilis were identified as the closest relatives of CDH by comparative amino acid sequence analysis, and a ThDP binding motif and a 2-fold Rossmann fold for FAD binding could be localized at the C-terminal end and central region of CDH, respectively. A first mechanism for the ring cleavage of CDO is presented, and it is suggested that the FAD cofactor in CDH is an evolutionary relict.
- Subjects :
- Rossmann fold
Stereochemistry
Molecular Sequence Data
Azoarcus
Microbiology
Cofactor
Benzoylformate decarboxylase
Substrate Specificity
chemistry.chemical_compound
Bacterial Proteins
Cytochrome P-450 Enzyme System
ddc:570
Hydrolase
Pyruvate oxidase
Amino Acid Sequence
Molecular Biology
Flavin adenine dinucleotide
biology
Cyclohexanones
Enzymes and Proteins
Biochemistry
chemistry
FAD binding
Denitrification
biology.protein
Thiamine Pyrophosphate
Sequence Alignment
Pyruvate decarboxylase
Subjects
Details
- ISSN :
- 10985530 and 00219193
- Volume :
- 193
- Database :
- OpenAIRE
- Journal :
- Journal of Bacteriology
- Accession number :
- edsair.doi.dedup.....822d639d84162d0da9ca8ddc031ed7d6