Purification, crystallization and preliminary X-ray diffraction analysis of NADP-dependent glutamate dehydrogenase from Aspergillus niger.
Glutamate dehydrogenase (GDH) catalyzes the NAD-dependent or NADP-dependent oxidative deamination of L-glutamate to 2-oxoglutarate and ammonia. This important reversible reaction establishes the link between carbon and nitrogen metabolism. In this study, Aspergillus niger NADP-GDH (AnGDH) has been overexpressed and purified. Purified AnGDH, with a high specific activity of 631.1 units per milligram of protein, was crystallized and the crystal diffracted to 2.9 Å resolution using a home X-ray source. Preliminary analysis of the X-ray diffraction data showed that the crystal belonged to space group R32, with unit-cell parameters a=b=173.8, c=241.5 Å, α=β=90, γ=120°. The crystals exhibited an unusually high solvent content (83.0%) and had only one molecule in the asymmetric unit. Initial phases were obtained by molecular replacement, and model building and structure refinement of AnGDH are in progress.
Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai 400 076, India.
Prem
P
Prakash
Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai 400 076, India.
Adhish S
AS
Walvekar
Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai 400 076, India.
Narayan S
NS
Punekar
Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai 400 076, India.
Prasenjit
P
Bhaumik
D001234:Aspergillus niger / Q000201:enzymology*; D003460:Crystallization; D018086:Glutamate Dehydrogenase (NADP+) / Q000737:chemistry* / Q000302:isolation & purification*; D014961:X-Ray Diffraction
D018086:Glutamate Dehydrogenase (NADP+)
Aspergillus niger; glutamate dehydrogenase