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The Effects of Nucleotides on Hexamer Stability of Glutamate Dehydrogenase
Author(s) -
Lekova Virzhiniya,
Bell Ellis
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.lb220
Subject(s) - random hexamer , cofactor , chemistry , nucleotide , glutamate dehydrogenase , oxidative deamination , biochemistry , guanidine , denaturation (fissile materials) , purine , gtp' , nad+ kinase , stereochemistry , enzyme , glutamate receptor , receptor , gene , nuclear chemistry
Bovine glutamate dehydrogenase which catalyzes the oxidative deamination of L‐Glutamate using either NAD or NADP as cofactor, is extensively regulated by a variety of ligands including the purine nucleotides, ADP and GTP as well as having a regulatory site for NADH. The aims of the current study are to establish whether 1] cofactor substrates, and 2] purine nucleotide regulatory ligands affect the stability of the hexamer. Guanidine Hydrochloride denaturation‐renaturation experiments were performed using the fluorescence of the three tryptophans in each subunit. Fluorescence emission spectra and lifetime measurements were used to follow the effects of denaturation and the impact that added ligands have. Dilution experiments were used to indicate whether the presence of ligands affected refolding of the protein. The results demonstrate that both cofactors and purine nucleotides affect both unfolding and refolding processes. Interestingly in the denaturation steps NADPH which binds only to the active site has similar effects to NADH which binds to both active and regulatory sites. Supported by NSF Grant MCB 0448905 to EB