Kinetic Mechanism of Beef Pancreatic L-Asparagine Synthetase

Rodney S. Markin, Craig A. Luehr, Sheldon M. Schuster

Research output: Contribution to journalArticlepeer-review

14 Scopus citations

Abstract

The kinetic mechanism of bovine pancreatic asparagine synthetase was deduced from initial velocity studies and product inhibition studies of both the glutamine-dependent and ammonia-dependent reactions. For the glutamine-dependent pathway, parallel lines were observed in the double reciprocal plot of 1 / V vs. 1 / [glutamine] at varied aspartate concentrations, and in the plot of 1 / V vs. 1 / [ATP] at varied aspartate concentrations. Intersecting lines were found for the plot of 1 / V vs. 1 / [ATP] at varied glutamine concentrations. Product inhibition patterns, including dual inhibitor studies for measuring the synergistic effects of multiproduct inhibition, were used to support an ordered bi-uni-uni-ter ping-pong mechanism. Glutamine and ATP sequentially bind, followed by the release of glutamate and the addition of aspartate. Pyrophosphate, AMP, and asparagine are then sequentially released. When the ammonia-dependent reaction was studied, it was found that the mechanism was significantly different. NH3 bound first followed by a random addition of ATP and aspartate. Pyrophosphate, AMP, and asparagine were then sequentially released as in the glutamine-utilizing mechanism. From these studies, a comprehensive mechanism has been proposed through which either glutamine or NH3 can provide nitrogen for asparagine production from aspartate.

Original languageEnglish (US)
Pages (from-to)7226-7232
Number of pages7
JournalBiochemistry
Volume20
Issue number25
DOIs
StatePublished - Dec 1981

ASJC Scopus subject areas

  • Biochemistry

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