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A stable carbocyclic analog of 5-phosphoribosyl-1-pyrophosphate to probe the mechanism of catalysis and regulation of glutamine phosphoribosylpyrophosphate amidotransferase.

Authors
  • Kim, J H
  • Wolle, D
  • Haridas, K
  • Parry, R J
  • Smith, J L
  • Zalkin, H
Type
Published Article
Journal
The Journal of biological chemistry
Publication Date
Jul 21, 1995
Volume
270
Issue
29
Pages
17394–17399
Identifiers
PMID: 7542237
Source
Medline
License
Unknown

Abstract

Glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase catalysis and regulation were studied using a new stable carbocyclic analog of PRPP, 1-alpha-pyrophosphoryl-2-alpha, 3-alpha-dihydroxy-4-beta-cyclopentane-methanol-5-phosphate (cPRPP). Although cPRPP competes with PRPP for binding to the catalytic C site of the Escherichia coli enzyme, two lines of evidence demonstrate that cPRPP, unlike PRPP, does not promote an active enzyme conformation. First, cPRPP was not able to "activate" Cys1 for reaction with glutamine or a glutamine affinity analog. The ring oxygen of PRPP may thus be necessary for the conformation change that activates Cys1 for catalysis. Second, binding of cPRPP to the C site blocks binding of AMP and GMP, nucleotide end product inhibitors, to this site. However, the binding of nucleotide to the allosteric site was essentially unaffected by cPRPP in the C site. Since it is expected that nucleotide inhibitors would bind with low affinity to the active enzyme conformation, the nucleotide binding data support the conclusion that cPRPP does not activate the enzyme.

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