Improved expression, purification and crystallization of a putative N-acetyl-gamma-glutamyl-phosphate reductase from rice (Oryza sativa).
N-Acetyl-gamma-glutamyl-phosphate reductase (AGPR) catalyzes the third step in an eight-step arginine-biosynthetic pathway that starts with glutamate. This enzyme converts N-acetyl-gamma-glutamyl phosphate to N-acetylglutamate-gamma-semialdehyde by an NADPH-dependent reductive dephosphorylation. AGPR from Oryza sativa (OsAGPR) was expressed in Escherichia coli at 291 K as a soluble fusion protein with an upstream thioredoxin-hexahistidine [Trx-(His)6] extension. OsAGPR(Ala50-Pro366) was purified and crystals were obtained using the sitting-drop vapour-diffusion method at 293 K and diffract X-rays to at least 1.8 A resolution. They belong to the hexagonal space group P6(1), with unit-cell parameters a = 86.11, c = 316.3 A.
Miura-Ohnuma
Jun
J
National Institute of Agrobiological Sciences, Japan.
Nonaka
Tsuyoshi
T
Katoh
Shizue
S
Murata
Katsuyoshi
K
Kita
Akiko
A
Miki
Kunio
K
Katoh
Etsuko
E
D000445:Aldehyde Oxidoreductases / Q000737:chemistry*; D000596:Amino Acids / Q000737:chemistry; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D004058:Diffusion; D004926:Escherichia coli / Q000378:metabolism; D005971:Glutamates / Q000737:chemistry; D006639:Histidine / Q000737:chemistry; D006801:Humans; D009249:NADP / Q000737:chemistry; D009842:Oligopeptides / Q000737:chemistry; D012275:Oryza / Q000201:enzymology*; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013696:Temperature; D013879:Thioredoxins / Q000737:chemistry; D014961:X-Ray Diffraction
D000596:Amino Acids; D005971:Glutamates; C471213:His-His-His-His-His-His; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D006639:Histidine; D013879:Thioredoxins; D009249:NADP; D000445:Aldehyde Oxidoreductases; C018722:N-acetyl-gamma-glutamyl-phosphate reductase