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4.2.1.84: nitrile hydratase

This is an abbreviated version!
For detailed information about nitrile hydratase, go to the full flat file.

Word Map on EC 4.2.1.84

Reaction

an aliphatic amide
=
A nitrile
 +
H2O

Synonyms

3-cyanopyridine hydratase, acrylonitrile hydratase, aliphatic nitrile hydratase, ANHase, Co-type NHase, Co-type nitrile hydratase, cobalt-containing nitrile hydratase, CoIII-NHase, CtNHase, Fe-NHase, H-NHase, H-nitrilase, high-molecular mass nitrile hydratase, high-molecular weight nitrile hydratase, hydratase, nitrile, iron-type nitrile hydratase, L-Nhase, L-nitrilase, low-molecular mass nitrile hydratase, low-molecular weight nitrile hydratase, MbNHase, NHase, NHaseK, NI1 NHase, NilCo, NilFe, nitrilase, nitrile hydratase, NthAB, PaNit, ppNHase, ReNHase, TNHase, toyocamycin nitrile hydratase

ECTree

     4 Lyases
         4.2 Carbon-oxygen lyases
             4.2.1 Hydro-lyases
                4.2.1.84 nitrile hydratase

Cloned

Cloned on EC 4.2.1.84 - nitrile hydratase

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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
chaperone GroEL2 is co-expressed with nitrile hydratase (NHase) in Escherichia coli in three ways: (a) monocistronic expression with one T7 promoter, (b) bicistronic expression with double T7 promoters, and (c) fusion expression with one T7 promoter driving the NHase-GroEL2 chimera. The NHase-GroEL2 chimera is the most successful expression strategy. Maximal nitrile hydratase activity is enhanced by 63.6% compared with the single NHase control
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co-expression of recombinant strains YHJ-1 to YHJ-4 with plasmids encoding the chaperons pG-KJE8, pGro7, pKJE7, pG-Tf2 and pTf16
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Co-substituted NHase produced in Escherichia coli grown in Co supplemented medium
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expressed in Escherichia coli BL21(DE3) cells
expression in Escherichia coli
expression in Escherichia coli BL21(DE3)
expression in Escherichia coli rosetta
expression in Escherichia coli, NovaBlue, series of recombinant plasmids with genes encoding NHase (nha1: alpha subunit and nha2: beta subunit) and P44k protein (nha3), different positions and combinations
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expression in Escherichia coli, strain BL21(DE3)
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expression of His-tagged wild-type enzyme and peptide fused enzyme variants in Escherichia coli strain BL21(DE3)
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expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
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expression of wild-type and mutant enzymes in Escherichia coli strain JM109
gene nit-30, DNA and amino acid sequence determination and analysis, overexpression in Escherichia coli strain Bl21(DE3)
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gene nthAB, DNA and amino acid sequence determination, analysis, and comparison, phylogenetic analysis and tree, expression of C-terminally His6-tagged enzyme in Escherichia coli strain BL21(DE3)
gene Pput_2728, gene cluster and phylogenetic analysis, functional expression of His6-tagged subunits in Escherichia coli strain BL21(DE3) by assisting of a putative activator gene adjacent to beta-subunit region. Maximal enzyme activity is obtained when the structural and activator genes are transcribed as one unit in plasmid pCDFDuet-1 at 18°C. The expressed product does not show any NHase activity when the downstream activator gene is ignored, and the product completely exists in insoluble inclusion body
gene Pput_2729, gene cluster and phylogenetic analysis, functional expression of His6-tagged subunits in Escherichia coli strain BL21(DE3) by assisting of a putative activator gene adjacent to beta-subunit region. Maximal enzyme activity is obtained when the structural and activator genes are transcribed as one unit in plasmid pCDFDuet-1 at 18°C. The expressed product does not show any NHase activity when the downstream activator gene is ignored, and the product completely exists in insoluble inclusion body
genes nha1 and nha2 encoding the subunits of nitrile hydratase, DNA and amino acid sequence determination and analysis
genes nha1 and nha2 encoding the subunits of nitrile hydratase, DNA and amino acid sequence determination and analysis. Genes ami (encoding an amidase), nha1 and nha2 of Rhodococcus erythropolis A4 form an operon transcribed from the Pami promoter and an internal Pnha promoter
genes nhpA and nhpB encoding the alpha and beta subunits of NHase, transcriptional regulation of the nitrile hydratase gene cluster, the NHase gene cluster comprises seven genes: oxdA, amiA, nhpA, nhpB, nhpC, nhpS, and acsA, nhpR codes for a positive transcriptional regulator in the NHase gene cluster, overview
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genes toyJKL and toyJ, recombinant expression of N-terminally His6-tagged trimeric and monomeric subunits in Escherichia coli
genetic organization of wild-type and chimeric mutant enzymes, expression of wild-type and recombinant chimeric mutants in Escherichia coli strain JM109
genomic organization and phylogenetic analysis, sequence comparison, overview
heterologously expressed in Escherichia coli. The resulting enzyme expressed as a single polypeptide with fused alpha- and beta-subunits linked by a seventeen-histidine region. The enzyme contains its full complement of Co(III) and is fully functional without the coexpression of an activator protein or Escherichia coli GroES/EL molecular chaperones
high molecular mass-NHase and low molecular mass-NHase genes cloned into Escherichia coli
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mutant enzymes are expressed in Escherichia coli HMS174(DE3)pLysS cells
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nitrile hydratase from Comamonas testosteroni is expressed in Escherichia coli, strain TG1, overexpressed chaperonin system
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Nocardia sp.(strain YS-2002) NHase gene with modified start codon in alpha subunit is overexpressed in Escherichia coli, strain BL21(DE3)
overexpressed in Escherichia coli by codon-optimization, engineering of Ribosome Binding Site and spacer sequences
recombinant expression in Escherichia coli strain HB101 from vector pUC18. The recombinant enzyme shows almost the same specific activity and other properties as the native enzyme
recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)
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separate expression of L-NHase, NhlAE, an the NhhA-NhlE hybrid mediator complex in Rhodococcus rhodochrous strain ATCC12674 and Rhodococcus fascians DSM43985. Necessity of gene nhhG for functional H-NHase expression
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the nhhBAG gene of Rhodococcus rhodochrous M33 that encodes nitrile hydratase is cloned and expressed in Corynebacterium glutamicum under the control of an ilvC promoter. To overexpress the nitrile hydratase, five types of plasmid variants are constructed by introducing mutations into 80 nucleotides near the translational initiation region of nhhB. Of them, pNBM4 with seven mutations shows the highest NHase activity, exhibiting higher expression levels of NhhB and NhhA than wild-type pNBW33, mainly owing to decreased secondary-structure stability and an introduction of a conserved Shine-Dalgarno sequence in the translational initiation region. In a fed-batch culture of recombinant Corynebacterium cells harboring pNBM4, the cell density reaches 53.4 g dry cell weight/l within 18 h, and the specific and total enzyme activities are estimated to be 0.0373 mM/min/mg dry cell weight and 1.992 mM/min/ml, respectively
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the nitrile hydratase (NHaseK, consisting of alpha- and beta-subunits) genes and the putative activator (17K) gene adjacent to the beta subunit region are cloned from Klebsiella oxytoca KCTC 1686. 17K is essential for the functional expression of recombinant NHaseK in Escherichia coli. The expression level of 17K is very low when the 17K gene and NHaseK structural genes are expressed as a gene cluster in Escherichia coli BL21(DE3). To improve the 17K expression level and NHaseK activity, the expression cassette is redesigned by placing the 17K gene and NHaseK structural genes under the control of different promoters in the pETDuet-1 expression vector, co-expressing the 17K gene with the gene cluster in a double plasmid or a single plasmid with a double promoter, and introducing an efficient Shine-Dalgarno sequence 5' to the 17K gene. The specific activity of NHaseK is improved when 17K is co-expressed with the gene cluster, whereas the production of NHaseK protein decreases. The maximum activity is achieved when an efficient Shine-Dalgarno sequence is introduced 5' to the 17K gene. The expression level of 17K is significantly improved and the expression level of NHaseK does not decrease significantly
YHJ-1, Escherichia coli strain BL21(DE3) with plasmid pYHJ1 (alphabeta+P44k)
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YHJ-2, Escherichia coli strain BL21(DE3) with plasmid pYHJ2 (alpha+betaP44k)
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YHJ-3, Escherichia coli strain BL21(DE3) with plasmid pYHJ3 (alphabeta)
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YHJ-4, Escherichia coli strain BL21(DE3) with plasmid pYHJ4 (alpha+beta)
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YHJ-5, Escherichia coli strain BL21(DE3) co-expressing plasmids pYHJ3/pYHJ5 (P44k)
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YHJ-6, Escherichia coli strain BL21(DE3) co-expressing plasmids pYHJ4/pYHJ5
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