Novel mechanism of inhibition of rat kidney-type glutaminase by bis-2-(5-phenylacetamido-1,2,4-thiadiazol-2-yl)ethyl sulfide (BPTES)

Mary M. Robinson, Steven J. McBryant, Takashi Tsukamoto, Camilo Rojas, Dana V. Ferraris, Sean K. Hamilton, Jeffrey C. Hansen, Norman P. Curthoys

Research output: Contribution to journalArticle

Abstract

The release of GA (mitochondrial glutaminase) from neurons following acute ischaemia or during chronic neurodegenerative diseases may contribute to the propagation of glutamate excitotoxicity. Thus an inhibitor that selectively inactivates the released GA may limit the accumulation of excess glutamate and minimize the loss of neurological function that accompanies brain injury. The present study examines the mechanism of inactivation of rat KGA (kidney GA isoform) by the small-molecule inhibitor BPTES [bis-2-(5-phenylacetamido-1,2,4- thiadiazol-2-yl)ethyl sulfide]. BPTES is a potent inhibitor of KGA, but not of the liver GA isoform, glutamate dehydrogenase or γ-glutamyl transpeptidase. Kinetic studies indicate that, with respect to glutamine, BPTES has a Ki of approx. 3 μM. Moreover, these studies suggest that BPTES inhibits the allosteric activation caused by phosphate binding and promotes the formation of an inactive complex. Gel-filtration chromatography and sedimentation-velocity analysis were used to examine the effect of BPTES on the phosphate-dependent oligomerization of KGA. This established that BPTES prevents the formation of large phosphate-induced oligomers and instead promotes the formation of a single oligomeric species with distinct physical properties. Sedimentation-equilibrium studies determined that the oligomer produced by BPTES is a stable tetramer. Taken together, the present work indicates that BPTES is a unique and potent inhibitor of rat KGA and elucidates a novel mechanism of inactivation.

Original languageEnglish (US)
Pages (from-to)407-414
Number of pages8
JournalBiochemical Journal
Volume406
Issue number3
DOIs
StatePublished - Sep 15 2007
Externally publishedYes

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Keywords

  • Bis-2-(5-phenylacetamido-1,2,4-thiadiazol-2-yl)ethyl sulfide (BPTES)
  • Glutamate excitotoxicity
  • Glutaminase inhibitor

ASJC Scopus subject areas

  • Biochemistry

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