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Literature summary for 4.4.1.9 extracted from

  • Yi, H.; Juergens, M.; Jez, J.M.
    Structure of soybean beta-cyanoalanine synthase and the molecular basis for cyanide detoxification in plants (2012), Plant Cell, 24, 2696-2706.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
recombinant enzyme expression as N-terminally His-tagged, functional dimeric type and mutant enzyme proteins lacking the mitochondrial localization sequence Glycine max

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recombinant wild-type and K95A mutant enzymes, hanging drops vapor diffusion method , 0004 ml of 10-12 mg/ml protein solution is mixed with an equal volume of crystallization solution containing 10% PEG 8000, 0.2 M NaCl, and 0.1 sodium/potassium phosphate, pH 7.0, and 0.2 M MgSO4, at 4°C over a 0.7 mL reservoir. The K95A mutant is crystallized in similar drops from a solution containing 20% PEG 3350, 0.2 M potassium citrate, pH 7.5, X-ray diffraction structure determination and analysis at 2.50 A and 1.77 A resolution, respectively, molecular replacement Glycine max

Protein Variants

Protein Variants Comment Organism
K95A site-directed mutagenesis, inactive mutant Glycine max

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.26
-
hydrogen cyanide recombinant enzyme, pH 9.0, 25°C Glycine max
0.81
-
L-cysteine recombinant enzyme, pH 9.0, 25°C Glycine max

Localization

Localization Comment Organism GeneOntology No. Textmining
mitochondrion the enzyme has a mitochondrial localization sequence Glycine max 5739
-

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
L-cysteine + hydrogen cyanide Glycine max
-
hydrogen sulfide + L-3-cyanoalanine
-
?

Organism

Organism UniProt Comment Textmining
Glycine max I1L6I6
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant functional N-terminally His-tagged wild-type and mutant enzymes lacking the mitochondrial localization sequence by nickel affinity chromatography and gel filtration Glycine max

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
L-cysteine + hydrogen cyanide
-
Glycine max hydrogen sulfide + L-3-cyanoalanine
-
?
L-cysteine + hydrogen cyanide the enzyme forms an alpha-aminoacrylate reaction intermediate from L-Cys Glycine max hydrogen sulfide + L-3-cyanoalanine
-
?
additional information the enzyme displays a 230fold higher catalytic efficiency with Cys than O-acetylserine, cf. EC 2.5.1.47 Glycine max ?
-
?

Subunits

Subunits Comment Organism
dimer The N-terminal domain (residues 94 to 197) is formed by a central beta-sheet (beta3 to beta5) surrounded by four alpha-helices (alpha1 to alpha4). In between the two domains, alpha5 (residues 195 to 2019) spans both halves of the structure. The C-terminal domain consists of a short beta-sheet (beta1 to beta2, residues 51 to 93) and a larger beta-sheet (beta6 to beta8) surrounded by three alpha-helices (alpha6 to alpha8). An additional alpha-helix (alpha9) forms part of the dimer interface by packing against alpha2 and alpha3 of the adjoining monomer Glycine max

Synonyms

Synonyms Comment Organism
CAS
-
Glycine max

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Glycine max

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
38.9
-
L-cysteine recombinant enzyme, pH 9.0, 25°C Glycine max
39.2
-
hydrogen cyanide recombinant enzyme, pH 9.0, 25°C Glycine max

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
9
-
assay at Glycine max

Cofactor

Cofactor Comment Organism Structure
pyridoxal 5'-phosphate binding site determination, each monomer consists of two mixed alpha/eta domains with a pyridoxal 5'-phosphate cofactor at the active site separating the two halves Glycine max

General Information

General Information Comment Organism
evolution the enzyme is a member of the beta-substituted alanine synthase (BSAS) family, structure and substrate specificity comparisons with another family member, O-acetylserine sulfhydrylase, EC 2.5.1.47, from Glycine max. This enzyme prefers O-acetylserine versus Cys by 23fold and sulfide by 270fold over CN- Glycine max
additional information comparison of the open active site wild-type enzyme and the closed active site K95A mutant monomer structures, overview Glycine max

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
48.02
-
L-cysteine recombinant enzyme, pH 9.0, 25°C Glycine max
150.8
-
hydrogen cyanide recombinant enzyme, pH 9.0, 25°C Glycine max