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Literature summary for 3.1.3.7 extracted from

  • Erickson, A.I.; Sarsam, R.D.; Fisher, A.J.
    Crystal structures of Mycobacterium tuberculosis CysQ, with substrate and products Bound (2015), Biochemistry, 54, 6830-6841 .
    View publication on PubMed

Application

Application Comment Organism
pharmacology the inhibition by lithium suggests the FIG superfamily of enzymes is the target of lithium therapy in manic-depressive illness Mycobacterium tuberculosis

Cloned(Commentary)

Cloned (Comment) Organism
gene cysQ, sequence comparisons, overexpression of His-tagged enzyme in Escherichia coli strain BL21(DE3) Mycobacterium tuberculosis

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recombinant Mycobacterium tuberculosis enzyme CysQ in a ligand-free structure, a lithium-inhibited state with substrate PAP bound, and a product-bound complex with AMP, phosphate, and three Mg2+ ions bound, sitting drop vapor diffusion method, mixing of 0.001 ml of 10 mg/ml protein in 20 mM Tris, pH 8.0, 100 mM NaCl, 1 mM DTT, 5% glycerol, and 1 mM AMP, with 0.001 ml of reservoir solution containing 24% PEG 1500 and 20% glycerol for the ligand-free enzyme, the calcium- and phosphate-bound structure is grown in 0.05 M CaCl2, 0.1 M Bis-Tris-HCl, pH 6.5, and 30% PEG MME 550, the magnesium- and phosphate-bound structure is grown in 0.1 M Mg acetate, 0.1 M Bis-Tris-HCl, pH 6.5, and 35% PEG MME 550, and for both the Ca2+ and Mg2+ structures, phosphate is not included in the crystallization conditions, the AMP-, Pi-, and Mg2+-bound crystals are grown in 0.1 M Mg acetate, 0.1 M Bis-Tris-HCl, pH 6.5, and 35% PEG MME 550 and soaked for 20 min in mother liquor supplemented with 3 mM AMP, 21°C, 1-5 days, X-ray diffraction structure determination and analysis at 1.45-2.05 A resolution. SIRAS (single isomorphous replacement with anomalous scattering) phasing information is collected from a ligand-free crystal grown in 0.2 M calcium acetate, 0.1 M MES-NaOH, pH 6.0, and 20% w/v PEG 8000 that is soaked overnight in mother liquor supplemented with a heavy atom cocktail: 1 mM uranyl acetate, 1 mM p-chloromercury benzoic acid sulfonate (PCMBS), and 1 mM KAu(CN)2. Before being flash-cooled, the crystal is transferred to a cryosolution containing 0.2 M Ca-acetate, 0.1 M MES-NaOH, pH 6.0, 20% w/v PEG 8000, and 20% glycerol. Molecular replacement and structure modelling Mycobacterium tuberculosis

Inhibitors

Inhibitors Comment Organism Structure
Li+ the Li+ ion displaces Mg2+ ion, specifically the Mg2+ ion at metal site 2, and is thought to stabilize the phosphate-bound product. Enzyme binding structure, overview Mycobacterium tuberculosis

Metals/Ions

Metals/Ions Comment Organism Structure
Ca2+ activates, enzyme binding structure, overview Mycobacterium tuberculosis
Mg2+ activates, three Mg2+ ions binding in the active site are required to form a productive enzyme, enzyme binding structure, overview Mycobacterium tuberculosis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
adenosine 3',5'-bisphosphate + H2O Mycobacterium tuberculosis
-
AMP + phosphate
-
?
adenosine 3',5'-bisphosphate + H2O Mycobacterium tuberculosis H37Rv
-
AMP + phosphate
-
?

Organism

Organism UniProt Comment Textmining
Mycobacterium tuberculosis P9WKJ1
-
-
Mycobacterium tuberculosis H37Rv P9WKJ1
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, dialysis, and ultrafiltration Mycobacterium tuberculosis

Reaction

Reaction Comment Organism Reaction ID
adenosine 3',5'-bisphosphate + H2O = AMP + phosphate proposed catalytic mechanism for CysQ, Asp50 deprotonates Thr96 that deprotonates the water (ligating a Mg2+), which attacks the phosphate, the pentacovalent 3'-phosphate transition state is stabilized by the three Mg2+ ions, overview Mycobacterium tuberculosis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
adenosine 3',5'-bisphosphate + H2O
-
Mycobacterium tuberculosis AMP + phosphate
-
?
adenosine 3',5'-bisphosphate + H2O i.e. PAP Mycobacterium tuberculosis AMP + phosphate
-
?
adenosine 3',5'-bisphosphate + H2O
-
Mycobacterium tuberculosis H37Rv AMP + phosphate
-
?
adenosine 3',5'-bisphosphate + H2O i.e. PAP Mycobacterium tuberculosis H37Rv AMP + phosphate
-
?
additional information Mtb CysQ can dephosphorylate myo-inositol 1-phosphate (IMP), fructose 1,6-bisphosphate (FBP), and 3'-phosphoadenosine 5'-monophosphate (PAP), but the catalytic efficiency is 1700 and 15000fold greater for PAP over FBP and IMP, respectively. AMP and phosphate enzyme binding structures, overview. The specificity of a 5'-phosphate or bis-phosphate sugars for CysQ can likely come from a conserved positively charged residue, Lys190 in CysQ, that ion pairs with the nonhydrolyzed phosphate Mycobacterium tuberculosis ?
-
?
additional information Mtb CysQ can dephosphorylate myo-inositol 1-phosphate (IMP), fructose 1,6-bisphosphate (FBP), and 3'-phosphoadenosine 5'-monophosphate (PAP), but the catalytic efficiency is 1700 and 15000fold greater for PAP over FBP and IMP, respectively. AMP and phosphate enzyme binding structures, overview. The specificity of a 5'-phosphate or bis-phosphate sugars for CysQ can likely come from a conserved positively charged residue, Lys190 in CysQ, that ion pairs with the nonhydrolyzed phosphate Mycobacterium tuberculosis H37Rv ?
-
?

Synonyms

Synonyms Comment Organism
3',5'-bisphosphate nucleotidase
-
Mycobacterium tuberculosis
CysQ
-
Mycobacterium tuberculosis
MTCY270.37
-
Mycobacterium tuberculosis
PAP phosphatase
-
Mycobacterium tuberculosis
Rv2131c
-
Mycobacterium tuberculosis

General Information

General Information Comment Organism
evolution CysQ is part of the larger FIG superfamily of phosphatases that dephosphorylates a monosaccharide-containing substrate. The FIG superfamily is comprised of family members: fructose-1,6-bisphosphatase (FBPase), inositol-monophosphatases (IMPase)/polyphosphatases (IPPase), and the glpX-encoded variant of FBPase (Class II). Many members of this superfamily display promiscuous phosphatase activity toward various monosaccharide-containing substrates but are usually more efficient in one type, which identifies their subfamily class. Mtb CysQ can dephosphorylate myo-inositol 1-phosphate (IMP), fructose 1,6-bisphosphate (FBP), and 3'-phosphoadenosine 5'-monophosphate (PAP), but the catalytic efficiency is 1700 and 15000fold greater for PAP over FBP and IMP, respectively. The FIG superfamily enzymes have demonstrated a dependence on divalent metal ions and are most active with magnesium. The superfamily also displays sensitivity to monovalent metals, and members are most strongly inhibited by lithium (Li+) Mycobacterium tuberculosis
malfunction cysQ knockout results in reduced levels of the sulfated glycolipid sulfolipid-1 and attenuation of cell growth Mycobacterium tuberculosis
physiological function because excess 3'-phosphoadenosine 5'-phosphate (PAP) alters the equilibrium of the sulfur pathway and inhibits sulfotransferases, PAP concentrations can affect the levels of sulfur-containing metabolites. PAP is removed by the phosphatase activity of CysQ, a 3',5'-bisphosphate nucleotidase, yielding AMP and phosphate, CysQ, a divalent cation metal-dependent phosphatase, is a major regulator of the sulfur activation pathway Mycobacterium tuberculosis