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Literature summary extracted from

  • Takahashi, R.; Nakamura, S.; Yoshida, T.; Kobayashi, Y.; Ohkubo, T.
    Crystallization of human nicotinamide phosphoribosyltransferase (2007), Acta Crystallogr. Sect. F, 63, 375-377.
    View publication on PubMedView publication on EuropePMC

Application

EC Number Application Comment Organism
2.4.2.12 drug development inhibitor-soaking experiments Homo sapiens

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.4.2.12 cDNA into pGEX6p1 for expression in Escherichia coli BL21 (DE) as glutathione S-transferase (GST) tagged fusion protein Homo sapiens

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
2.4.2.12 apo enzyme, diffraction of third crystals at 2 A resolution at -174°C, crystals belong to space group P2(1) with unit-cell parameters a = 60.56 A, b = 106.40 A, c =82.78 A, beta = 96.50°, technique: 1 microlitre of 10 mg/ml protein solution to 1 microlitre reservoir solution (38% pentaerytritol propoxylate (5/4 PO/OH) and 200 mM KCl pH 9) one week at 4°C, generation of first crystals by hanging-drop vapour diffusion (Index HT screening kit), generation of second crystals by microseeding: reservoir solution contained crushed first crystals, generation of third crystals by Repeated Seeding Technique: reservoir solution contained crushed second crystals Homo sapiens

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.4.2.12 nicotinamide + alpha-D-5-phosphoribosyl-1-diphosphate Homo sapiens
-
nicotinamide mononucleotide + diphosphate
-
r

Organism

EC Number Organism UniProt Comment Textmining
2.4.2.12 Homo sapiens P43490
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.4.2.12 lysis of bacteria by ultrasonification in presence of 1 mM phenylmethylsulfonylfluorid (PMSF) and 1 mM dithiothreitol (DTT), ultracentrifugation, supernatant applied to glutathione Sepharose 4B beads and washed, cleavage of glutathione S-transferase (GST) tag by PreScission Protease, purification by gel-filtration chromatography in Tris-HCl pH 8.5, 150 mM NaCl, 1 mM DTT, concentration to 10 mg/ml by ultrafiltration on Microcon YM-10, stored at 4°C Homo sapiens

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.4.2.12 nicotinamide + alpha-D-5-phosphoribosyl-1-diphosphate
-
Homo sapiens nicotinamide mononucleotide + diphosphate
-
r

Subunits

EC Number Subunits Comment Organism
2.4.2.12 dimer according to sedimentation-equilibrium measurements following ultracentrifugation of the protein solution Homo sapiens
2.4.2.12 homodimer two molecules in the asymmetric unit of the crystal according to Matthews coefficient Vm = 2.4 A/Da, corresponding to a solvent content of 48.5% Homo sapiens

Synonyms

EC Number Synonyms Comment Organism
2.4.2.12 nicotinamide phosphoribosyltransferase
-
Homo sapiens
2.4.2.12 NMPRTase
-
Homo sapiens