Microbial metabolism of aromatic nitriles. Enzymology of C–N cleavage by Nocardia sp. (Rhodochrous group) N.C.I.B. 11216
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TLDR
Cell-free extracts of benzonitrile-grown cells contain an enzyme that catalyses the conversion of benzoneitrile directly into benzoic acid without intermediate formation of benzamide, and studies with different substrates indicate that the nitrilase is specific for nitrile groups directly attached to the benzene ring.Abstract:
1. An organism utilizing benzonitrile as sole carbon and nitrogen source was isolated by the enrichment-culture technique and identified as a Nocardia sp. of the rhodochrous group. 2. Respiration studies indicate that nitrile degradation proceeds through benzoic acid and catechol. 3. Cell-free extracts of benzonitrile-grown cells contain an enzyme that catalyses the conversion of benzonitrile directly into benzoic acid without intermediate formation of benzamide. 4. This nitrilase enzyme was purified by DEAE-cellulose chromatography and gel filtration on Sephadex G-100 in the presence and absence of substrate. The purity of the enzyme was confirmed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and isoelectric focusing on polyacrylamide gel. 5. The enzyme shows a time-dependent substrate-activation process in which the substrate catalyses the association of inactive subunits of mol.wt. 45000 to form the polymeric 12-unit active enzyme of mol.wt. 560000. The time required for complete association is highly dependent on the concentration of the enzyme, temperature and pH. 6. The associated enzyme has a pH optimum of 8.0 and Km with benzonitrile as substrate of 4mm. The activation energy of the reaction as deduced from the Arrhenius plot is 51.8kJ/mol. 7. Enzyme activity is inhibited by thiol-specific reagents and several metal ions. 8. Studies with different substrates indicate that the nitrilase is specific for nitrile groups directly attached to the benzene ring. Various substituents in the ring are compatible with activity, though ortho-substitution, except by fluorine, renders the nitrile invulnerable to attack. 9. The environmental implications of these findings and the possible significance of the enzyme in the regulation of metabolism are discussed.read more
Citations
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The nitrilase superfamily: classification, structure and function
Helen C. Pace,Charles Brenner +1 more
TL;DR: The nitrilase superfamily consists of thiol enzymes involved in natural product biosynthesis and post-translational modification in plants, animals, fungi and certain prokaryotes and genetic and biochemical analysis of the family members and their associated domains assists in predicting the localization, specificity and cell biology of hundreds of uncharacterized protein sequences.
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The nitrilase family of CN hydrolysing enzymes - a comparative study.
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Nitrile hydratases (NHases): At the interface of academia and industry
TL;DR: The present review critically describes the available information on natural sources (based on activity and phylogenetic analysis), biochemical properties, catalysis-structure relationship, molecular expression and potential applications of this enzyme.
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Nitrilases in nitrile biocatalysis: recent progress and forthcoming research.
TL;DR: This critical review summarizes the current status of nitrilase research, and discusses a number of challenges and significant attempts in its further development.
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The superiority of the third-generation catalyst, Rhodococcus rhodochrous J1 nitrile hydratase, for industrial production of acrylamide
TL;DR: The superiority of Rhodococcus rhodochrous J1 nitrile hydratase was demonstrated in comparison with other acrylamide-producing bacteria and the high stability, high catalytic efficiency and other outstanding features of the J1 enzyme are analysed and discussed.
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