Allosteric regulation of the NAD malic enzyme from cauliflower: activation by fumarate and coenzyme A

Abstract

Activation of the NAD malic enzyme is shown to be caused by free, uncomplexed fumarate2-. Mg-fumarate has no detectable effect on the enzyme. Fumarate2- isotherms are biphasic in that they consist of an activating as well as a deactivating region. Activation is shown to result from an increase in the affinity of the enzyme for malate2- while deactivation results from a reduction in Vmax. Phosphate does not affect the response of the enzyme to fumarate2-, while Cl- inhibits the enzyme in a manner that cannot be overcome by fumarate2-. SO42-, another activator of the malic enzyme, reduces the Ka for fumarate2- from 3.9 to 2.1 mM. Activation of the enzyme by coenzyme A (CoA) is hyperbolic with a Ka for CoA of 2.1 microM. Fumarate2- reduces this value to 1.2 microM. CoA, like SO42-, is able to increase the affinity of the enzyme for fumarate2-, decreasing its Ka by 56%. An additional effect of fumarate2- is to cause the interconversion of different catalytic forms of the enzyme which exist when Mg2- is limiting. On the basis of these results, a model of the number and types of allosteric sites present on the NAD malic enzyme is proposed.