Kinetic mechanism of NADH-enoyl-ACP reductase from Brassica napus

Enoyl-ACP reductase, a component of fatty acid synthase, is a target for an ti-microbial agents and herbicides. Here we demonstrate the kinetic mechani sm to be a compulsory-order ternary complex with NADH binding before the ac yl substrate. Matrix-assisted laser desorption ionisation mass spectrometry analysis of enzymatically and synthesised crotonyl-ACP substrate showed th e former to contain a single acyl group, whereas the latter contained up to four additional crotonylations. The use of authentic crotonyl-ACP will be important in future kinetic and crystallographic studies. (C) 2000 Federati on of European Biochemical Societies. Published by Elsevier Science B.V. Al l rights reserved.