Bringing nitrilase sequences from databases to life: the search for novel substrate specificities with a focus on dinitriles
Language English Country Germany Media print-electronic
Document type Journal Article, Research Support, Non-U.S. Gov't
PubMed
26521240
DOI
10.1007/s00253-015-7023-1
PII: 10.1007/s00253-015-7023-1
Knihovny.cz E-resources
- Keywords
- Arylacetonitrilases, Cyano acids, Dinitriles, Genome mining, Nitrilases,
- MeSH
- Aminohydrolases genetics metabolism MeSH
- Data Mining MeSH
- Escherichia coli genetics metabolism MeSH
- Gene Expression MeSH
- Cloning, Molecular MeSH
- Nitriles metabolism MeSH
- Recombinant Proteins isolation & purification metabolism MeSH
- Substrate Specificity MeSH
- Computational Biology MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Aminohydrolases MeSH
- nitrilase MeSH Browser
- Nitriles MeSH
- Recombinant Proteins MeSH
The aim of this study was to discover new nitrilases with useful activities, especially towards dinitriles that are precursors of high-value cyano acids. Genes coding for putative nitrilases of different origins (fungal, plant, or bacterial) with moderate similarities to known nitrilases were selected by mining the GenBank database, synthesized artificially and expressed in Escherichia coli. The enzymes were purified, examined for their substrate specificities, and classified into subtypes (aromatic nitrilase, arylacetonitrilase, aliphatic nitrilase, cyanide hydratase) which were largely in accordance with those predicted from bioinformatic analysis. The catalytic potential of the nitrilases for dinitriles was examined with cyanophenyl acetonitriles, phenylenediacetonitriles, and fumaronitrile. The nitrilase activities and selectivities for dinitriles and the reaction products (cyano acid, cyano amide, diacid) depended on the enzyme subtype. At a preparative scale, all the examined dinitriles were hydrolyzed into cyano acids and fumaronitrile was converted to cyano amide using E. coli cells producing arylacetonitrilases and an aromatic nitrilase, respectively.
Department of Biochemistry Charles University Prague Hlavova 8 CZ 128 40 Prague Czech Republic
Institute of Microbiology Czech Academy of Sciences Vídeňská 1083 CZ 142 20 Prague Czech Republic
References provided by Crossref.org
Genetic and Functional Diversity of Nitrilases in Agaricomycotina
