Campus Units

Biochemistry, Biophysics and Molecular Biology

Document Type

Article

Publication Version

Published Version

Publication Date

1-2005

Journal or Book Title

Biochemistry

Volume

44

Issue

2

First Page

766

Last Page

774

DOI

10.1021/bi048191w

Abstract

The conversion of ATP, l-aspartate, and 5-aminoimidazole-4-carboxyribonucleotide (CAIR) to 5-aminoimidazole-4-(N-succinylcarboxamide) ribonucleotide (SAICAR), ADP, and phosphate by phosphoribosylaminoimidazolesuccinocarboxamide synthetase (SAICAR synthetase) represents the eighth step of de novo purine nucleotide biosynthesis. SAICAR synthetase and other enzymes of purine biosynthesis are targets of natural products that impair cell growth. Prior to this study, no kinetic mechanism was known for any SAICAR synthetase. Here, a rapid equilibrium random ter-ter kinetic mechanism is established for the synthetase from Escherichia coli by initial velocity kinetics and patterns of linear inhibition by IMP, adenosine 5‘-(β,γ-imido)triphosphate (AMP-PNP), and maleate. Substrates exhibit mutual binding antagonism, with the strongest antagonism between CAIR and either ATP or l-aspartate. CAIR binds to the free enzyme up to 200-fold more tightly than to the ternary enzyme−ATP−aspartate complex, but the latter complex may be the dominant form of SAICAR synthetase in vivo. IMP is a competitive inhibitor with respect to CAIR, suggesting the possibility of a hydrogen bond interaction between the 4-carboxyl and 5-amino groups of enzyme-bound CAIR. Of several aspartate analogues tested (hadacidin, l-malate, succinate, fumarate, and maleate), maleate was by far the best inhibitor, competitive with respect to l-aspartate. Inhibition by IMP and maleate is consistent with a chemical mechanism for SAICAR synthetase that parallels that of adenylosuccinate synthetase.

Comments

Reprinted (adapted) with permission from Biochemistry 44 (2005): 766, doi: 10.1021/bi048191w. Copyright 2005 American Chemical Society.

Copyright Owner

American Chemical Society

Language

en

File Format

application/pdf

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