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Purification, crystallization and preliminary X‐ray diffraction analysis of NADP‐dependent glutamate dehydrogenase from Aspergillus niger
Author(s) -
Prakash Prem,
Walvekar Adhish S.,
Punekar Narayan S.,
Bhaumik Prasenjit
Publication year - 2014
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x14021499
Subject(s) - aspergillus niger , glutamate dehydrogenase , crystallization , nad+ kinase , oxidative deamination , x ray crystallography , dehydrogenase , deamination , crystallography , molecule , crystal structure , chemistry , enzyme , stereochemistry , diffraction , biochemistry , glutamate receptor , organic chemistry , receptor , physics , optics
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 R 32, 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.