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The structural basis of proteolytic activation of bovine glutamate dehydrogenase
Author(s) -
Carrigan John B.,
Engel Paul C.
Publication year - 2008
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.034785.108
Subject(s) - glutamate dehydrogenase , chymotrypsin , proteolysis , biochemistry , trypsin , glutamate receptor , chemistry , cleavage (geology) , allosteric regulation , dehydrogenase , enzyme , biology , receptor , paleontology , fracture (geology)
Abstract In this work, we re‐examine the previously reported phenomenon of the creation of a superactive glutamate dehydrogenase by proteolytic modification by chymotrypsin and explore the various discrepancies that came to light during those studies. We find that superactivation is caused by cleavage at the N terminus of the protein and not the C‐terminal allosteric site, as has previously been suggested. N‐terminal sequencing reveals that TLCK‐treated chymotrypsin cleaves bovine glutamate dehydrogenase at phenylalanine 10. We suggest that trypsin contamination in nontreated chymotrypsin may have led to the production of the larger 4–5 kDa digestion product, previously misinterpreted as having caused the activation. In line with some previous studies, we can confirm that GTP inhibition is attenuated to some extent by the proteolysis, while ADP activation is almost abolished. Utilizing the recently solved structures of bovine glutamate dehydrogenase, we illustrate the cleavage points.