This enzyme is a key enzyme in the biosynthesis of mycothiol, a small molecular weight thiol found in Mycobacteria spp. and other actinomycetes. Mycothiol plays a fundamental role in these organisms by helping to provide protection from the effects of reactive oxygen species and electrophiles, including many antibiotics. The enzyme may represent a novel target for new classes of antituberculars .
the mechanism of MshC has been proposed as bi-uni-uni-bi ping pong using steady state kinetics and positional isotope exchange studies. In the first phase of the enzymatic reaction, the nucleophilic attack of carboxylate of cysteine onto the phosphate of ATP leads to the formation of the cysteine-adenylate intermediate releasing inorganic diphosphate. This cysteine-adenylate intermediate is attacked by the amine of GlcN-Ins to form Cys-GlcN-Ins and in the process releases AMP, which completes the second phase of the enzymatic reaction, overview. During this stage, the reacting substrates and coenzymes are held in place by the amino acid residues W227, H55, T46, and D251
the mechanism of MshC has been proposed as bi-uni-uni-bi ping pong using steady state kinetics and positional isotope exchange studies. In the first phase of the enzymatic reaction, the nucleophilic attack of carboxylate of cysteine onto the phosphate of ATP leads to the formation of the cysteine-adenylate intermediate releasing inorganic diphosphate. This cysteine-adenylate intermediate is attacked by the amine of GlcN-Ins to form Cys-GlcN-Ins and in the process releases AMP, which completes the second phase of the enzymatic reaction, overview. During this stage, the reacting substrates and coenzymes are held in place by the amino acid residues W227, H55, T46, and D251
This enzyme is a key enzyme in the biosynthesis of mycothiol, a small molecular weight thiol found in Mycobacteria spp. and other actinomycetes. Mycothiol plays a fundamental role in these organisms by helping to provide protection from the effects of reactive oxygen species and electrophiles, including many antibiotics. The enzyme may represent a novel target for new classes of antituberculars [2].
the ATP-dependent ligation of Cys to GlcNIns is a key step in mycothiol biosynthesis, catalyzed by MshC, which is essential for growth of Mycobacterium tuberculosis
the ATP-dependent ligation of Cys to GlcNIns is a key step in mycothiol biosynthesis, catalyzed by MshC, which is essential for growth of Mycobacterium tuberculosis
a key step in mycothiol, i.e. 1-D-myo-inosityl-2-(N-acetyl-L-cysteinyl)amido-2-deoxy-R-D-glucopyranoside or MSH or AcCys-GlcN-Ins, biosynthesis, mycothiol is produced to act against oxidative and antibiotic stress and is essential for cell growth, biosynthetic pathway, overview
the product 1D-myo-inositol 2-(L-cysteinyl)amido-2-deoxy-alpha-D-glucopyranoside is an intermediate in the biosynthetic pathway of mycothiol. Mycothiol is produced by Mycobacterium tuberculosis to maintain an intracellular reducing environment and protect against oxidative and antibiotic induced stress. The biosynthesis of mycothiol is essential for cell growth
positional isotope exchange analysis confirms that MshC catalyzes the formation of a kinetically competent cysteinyl-adenylate intermediate after the addition of ATP and cysteine
inactivation of the mshC gene in RHA1 results in the accumulation of the MSH pathway intermediate, 1D-myo-inositol 2-amino-2-deoxy-alpha-D-glucopyranoside. The mutant is deficient in the biochemical degradation of a number of xenobiotics metabolized by the parent strain, it shows increased susceptibility to a number of antibiotics and it shows unusual growth characteristics, exhibiting a long lag phase before normal exponential growth
the ATP-dependent ligation of Cys to GlcNIns is a key step in mycothiol biosynthesis, catalyzed by MshC, which is essential for growth of Mycobacterium tuberculosis
the ATP-dependent ligation of Cys to GlcNIns is a key step in mycothiol biosynthesis, catalyzed by MshC, which is essential for growth of Mycobacterium tuberculosis
a key step in mycothiol, i.e. 1-D-myo-inosityl-2-(N-acetyl-L-cysteinyl)amido-2-deoxy-R-D-glucopyranoside or MSH or AcCys-GlcN-Ins, biosynthesis, mycothiol is produced to act against oxidative and antibiotic stress and is essential for cell growth, biosynthetic pathway, overview
the product 1D-myo-inositol 2-(L-cysteinyl)amido-2-deoxy-alpha-D-glucopyranoside is an intermediate in the biosynthetic pathway of mycothiol. Mycothiol is produced by Mycobacterium tuberculosis to maintain an intracellular reducing environment and protect against oxidative and antibiotic induced stress. The biosynthesis of mycothiol is essential for cell growth
inactivation of the mshC gene in RHA1 results in the accumulation of the MSH pathway intermediate, 1D-myo-inositol 2-amino-2-deoxy-alpha-D-glucopyranoside. The mutant is deficient in the biochemical degradation of a number of xenobiotics metabolized by the parent strain, it shows increased susceptibility to a number of antibiotics and it shows unusual growth characteristics, exhibiting a long lag phase before normal exponential growth
ATP-competitive inhibitor of MshC, binds MshC with a KD of 0.22 microM, and inhibits the growth of Mycobacterium tuberculosis under aerobic and anaerobic conditions with minimum inhibitory and anaerobic bactericidal concentrations of 1.2 and 0.3 microg/ml, respectively
the enzyme structure is critical to developing potential inhibitors, synthesis of substrate analogues as potential inhibitors for the MshC enzyme. Target molecules are synthesized employing a Schmidt glycosylation strategy using an enantiomerically pure inositol acceptor and 2-deoxy trichloroacetimidate glycosyl donors with glycosylation yields greater than 70% and overall yields over 5%. The inositol acceptor is obtained via chiral resolution of myo-inositol
the enzyme structure is critical to developing potential inhibitors, synthesis of substrate analogues as potential inhibitors for the MshC enzyme. Target molecules are synthesized employing a Schmidt glycosylation strategy using an enantiomerically pure inositol acceptor and 2-deoxy trichloroacetimidate glycosyl donors with glycosylation yields greater than 70% and overall yields over 5%. The inositol acceptor is obtained via chiral resolution of myo-inositol
ATP-dependent L-cysteine:1D-myo-inosityl 2-amino-2-deoxy-alpha-D-glucopyranoside ligase, mycothiol biosynthesis enzyme MshC, is related to class I cysteinyl-tRNA synthetases.
spectrophotometric assay method development with coupling of diphosphatase to convert diphosphate to phosphate and spectrophotometric detection of the latter via the phosphomolybdate complex with malachite green, overview
mutants defective in mycothiol biosynthesis show mutations in genes coding for the glycosyltransferase (mshA) or the cysteine ligase (mshC). These mutants show low-level resistance to isoniazid but are highly resistant to ethionamide, mutations in mycothiol biosynthesis genes may contribute to isoniazid or ethionamide resistance across mycobacterial species
Mycobacterium smegmatis mutants disrupted in mscR, coding for a dual function S-nitrosomycothiol reductase and formaldehyde dehydrogenase, and mshC, coding for a mycothiol ligase, EC 6.3.1.13, and lacking mycothiol (MSH), are more susceptible to S-nitrosoglutathione (GSNO) and aldehydes than wild-type. MSH is a cofactor for MscR, and both mshC and mscR are induced by GSNO and aldehydes. The transposon mutant, S24, disrupted in mshC, is most sensitive to killing by GSNO
Mycobacterium smegmatis mutants disrupted in mscR, coding for a dual function S-nitrosomycothiol reductase and formaldehyde dehydrogenase, and mshC, coding for a mycothiol ligase, EC 6.3.1.13, and lacking mycothiol (MSH), are more susceptible to S-nitrosoglutathione (GSNO) and aldehydes than wild-type. MSH is a cofactor for MscR, and both mshC and mscR are induced by GSNO and aldehydes. The transposon mutant, S24, disrupted in mshC, is most sensitive to killing by GSNO
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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
incubation of the enzyme with the cysteinyl adenylate analogue, 5'-O-[N-(L-cysteinyl)-sulfamonyl]adenosine, followed by a 24 h limited trypsin proteolysis yields an enzyme preparation that readily crystallizes, 1.6 A crystal structure
recombinant enzyme from mshC-deficient mutant strain I64 of Mycobacterium smegmatis by ammonium sulfate fractionation, anion exchange chromatography, hydroxyapatite chromatography, and gel filtration
gene mshC, construction of three N-terminal-MshC fusion proteins where N-terminal tags include the B1 domain of 1. streptococcal protein G to give GB1-MshC, 2. glutathione-S-transferase to give GST-MshC, and 3. maltose binding protein to give MBP-MshC, for expression in M. smegmatis, expression in enzyme-deficient mutant strain mc2155, i.e. I64 L205P, optimization of recombinant enzyme expression, overview
Sareen, D.; Steffek, M.; Newton, G.L.; Fahey, R.C.
ATP-dependent L-cysteine:1D-myo-inosityl 2-amino-2-deoxy-alpha-D-glucopyranoside ligase, mycothiol biosynthesis enzyme MshC, is related to class I cysteinyl-tRNA synthetases
A coupled spectrophotometric assay for l-cysteine:1-D-myo-inosityl 2-amino-2-deoxy-alpha-D-glucopyranoside ligase and its application for inhibitor screening
Cloning, expression and rapid purification of active recombinant mycothiol ligase as B1 immunoglobulin binding domain of streptococcal protein G, glutathione-S-transferase and maltose binding protein fusion proteins in Mycobacterium smegmatis