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 language | English (US) |
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Pages (from-to) | 7226-7232 |
Number of pages | 7 |
Journal | Biochemistry |
Volume | 20 |
Issue number | 25 |
DOIs | |
State | Published - Dec 1981 |
ASJC Scopus subject areas
- Biochemistry