67799-13-7

Upstream product

• 5147-00-2 O-acetyl-L-serine 
• 127-07-1 N-hydroxyurea 
• 590-28-3 potassium cyanate 
• 53459-31-7 O-ureido-D-serine 

Downstream Products

• 53459-31-7 O-ureido-D-serine 
• 52-90-4 L-Cysteine 
• 1637-71-4 S-sulfo-L-cysteine 

Relevant academic research and scientific papers

PLP-independent racemization: Mechanistic and mutational studies of: O -ureidoserine racemase (DcsC)

O-Ureidoserine racemase (DcsC) is a PLP-independent enzyme in the biosynthetic route to the antibiotic d-cycloserine. Here we present the recombinant expression and characterization of a significantly more active DcsC variant featuring an N-terminal SUMO-tag. Synthesis of enantiomeric pure inhibitors in combination with site-specific mutation of active site cysteines to serines of this enzyme offers closer insights into the mechanism of this transformation. Homology modelling with a close relative (diaminopimelate epimerase, DapF) inspired C- and N-terminal truncation of DcsC to produce a more compact yet still active enzyme variant.

The structural and mutational analyses of O-ureido-L-serine synthase necessary for D-cycloserine biosynthesis

We have recently been successful in cloning a gene cluster necessary for the biosynthesis of d-cycloserine (d-CS) from d-CS-producing Streptomyces lavendulae ATCC11924. Although dcsD, one of the ORFs located on the gene cluster, encodes a protein homologous to O-acetylserine sulfhydrylase that synthesizes l-cysteine using O-acetyl-l-serine together with sulfide, it functions to form O-ureido-l-serine as a d-CS biosynthetic intermediate, using O-acetyl-l-serine together with hydroxyurea (HU). In the present study, using crystallographic and mutational studies, three amino acid residues in DcsD that are important for the substrate preference toward HU were determined. We showed that two of the three residues are important for the binding of HU into the substrate-binding pocket. The other residue contributes to the formation of a loose hydrogen-bond network during the catalytic reaction. Information regarding the amino acid residues will be very useful in the design of a new catalyst for synthesizing the β-substituted-L-alanine derivatives.

Establishment of an in vitro D-cycloserine-synthesizing system by using O-ureido-L-serine synthase and D-cycloserine synthetase found in the biosynthetic pathway

We have recently cloned a DNA fragment containing a gene cluster that is responsible for the biosynthesis of an antituberculosis antibiotic, D-cycloserine. The gene cluster is composed of 10 open reading frames, designated dcsA to dcsJ. Judging from the sequence similarity between each putative gene product and known proteins, DcsC, which displays high homology to diaminopimelate epimerase, may catalyze the racemization of O-ureidoserine. DcsD is similar to O-acetylserine sulfhydrylase, which generates L-cysteine using O-acetyl-L-serine with sulfide, and therefore, DcsD may be a synthase to generate O-ureido-L-serine using O-acetyl-L-serine and hydroxyurea. DcsG, which exhibits similarity to a family of enzymes with an ATP-grasp fold, may be an ATP-dependent synthetase converting O-ureido-D-serine into D-cycloserine. In the present study, to characterize the enzymatic functions of DcsC, DcsD, and DcsG, each protein was overexpressed in Escherichia coli and purified to near homogeneity. The biochemical function of each of the reactions catalyzed by these three proteins was verified by thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC), and, in some cases, mass spectrometry. The results from this study demonstrate that by using a mixture of the three purified enzymes and the two commercially available substrates O-acetyl-L-serine and hydroxyurea, synthesis of D-cycloserine was successfully attained. These in vitro studies yield the conclusion that DcsD and DcsG are necessary for the syntheses of O-ureido-L-serine and D-cycloserine, respectively. DcsD was also able to catalyze the synthesis of L-cysteine when sulfide was added instead of hydroxyurea. Furthermore, the present study shows that DcsG can also form other cyclic D-amino acid analogs, such as D-homocysteine thiolactone. Copyright

BIOSYNTHESIS OF β-(1,2,4-TRIAZOL-1-YL)ALANINE IN HIGHER PLANTS

β-(1,2,4-triazol-1-yl)Alanine, an important metabolite of the triazole-based fungicide Myclobutanil, was shown to be derived from O-acetyl-L-serine and 1,2,4-triazole by cysteine synthase in higher plants.Some properties of this enzyme in the biosynthesis of β-(1,2,4-triazol-1-yl)alanine are described.

ENZYMATIC SYNTHESIS OF THE NEUROEXCITATORY AMINO ACID QUISQUALIC ACID BY CYSTEINE SYNTHASE

Key Word Index - Quisqualis indica var. villosa; Combretaceae; cysteine synthase; isoenzyme; enzyme purification; biosynthesis; heterocyclic β-substituted alanines; quisqualic acid; O-acetyl-L-serine; cysteine.Purification of cysteine synthase from the leaves of Quisqualis indica var. villosa reveals the presence of two forms of this enzyme, separated by chromatography on DEAE-Sephadex A-50.Isoenzyme A was purified 10 000-fold and had a specific activity of 10.8 U/mg protein.Isoenzyme B was purified 460-fold with a specific activity of 0.49 U/mg protein.Both isoenzymes have the same Mrs (58 000) and dissociate into identical subunits (Mr 29 000).The Km value of isoenzyme A is 1.9 mM for O-acetyl-L-serine and 59 μM for sulphide, while that of isoenzyme B is 7.1 mM for O-acetyl-L-serine and 4.0 mM for 3,5-dioxo-1,2,4-oxadiazolidine.Both isoenzymes catalyse the formation of cysteine from O-acetyl-L-serine and hydrogen sulphide, but only isoenzyme B catalyses the formation of L-quisqualic acid.Other significant differences occur in the substrate specificity of the two isoenzymes.Some properties of the purified cysteine synthase isoenzymes are also described.

PURIFICATION AND CHARACTERIZATION OF β-(PYRAZOL-1-YL)-L-ALANINE SYNTHASE FROM CITRULLUS VULGARIS

From seedlings of Citrullus vulgaris the enzyme β-(pyrazol-1-yl)-L-alanine synthase was purified 200-fold, when it showed electrophoretic homogeneity (MW 58000) and could be dissociated into identical subunits (MW 32000) each containing one molecule of pyridoxal 5'-phosphate.The Km value was 2.5E-3 M for O-acetyl-L-serine and 7.4E-2 M for pyrazole.The enzyme did not catalyse the formation of related β-substituted alanines, such as mimosine and L-quisqualic acid, and significant differences were found between the β-(pyrazol-1-yl)-L-alanine synthase and β-substituted alanine synthases and cystein synthase from other sources.Key dex - Citrullus vulgaris; Cucurbitaceae; watermelon; β-(pyrazol-1-yl)-L-alanine synthase; enzyme purification; O-acetyl-L-serine; β-(pyrazol-1-yl)-L-alanine.