Substrate specificity of S-Adenosylhomocysteinase : Custeine is a substrate of the plant and mammalian enzymes

Abstract

Substrate specificity of S-adenosylhomocysteinases ( S- adenosyl- l- homocysteine hydrolase, EC 3.3.1.1) with respect to amino acid has been studied using homogeneous preparations of the enzymes from yellow lupin ( Lupinus luteus) seeds and bovine liver. Both enzymes use cysteine, in addition to homocysteine, as a substrate. Homoserine, serine, pinicillamine, reduced glutathione and 2-mercaptoethanol are not substrates. In the presence of cysteine, the reaction of S-adenosylthio-amino acid synthesis is characterized by 20–40-fold lower k cat values ( k cat = 0.23 s −1 or 0.11 s −1 in the presence of cysteine and either bovine or lupin enzyme) and 270-250-fold higher K m values ( K m for cysteine is 15 mM and 35 mM with bovine and lupin enzyme, respectively) than the reaction in the presence of the normal substrate, homocysteine. In the reverse reaction, S-adenosylcysteine is hydrolyzed by the mammalian enzyme much faster than by the plant one. Specificity ( k cat/ K m) towards S-adenosylcysteine and S-adenosyihomocysteine is 0.9 M −1 · s −1 and 60 000 M −1 · s −1, respectively, with the plant enzyme and 15.3 M −1 · s −1 and 70 000 M −1 · s −1, respectively, with the mammalian enzyme. With plant enzyme, the reactions with cysteine and homocysteine are not competitive, i.e., cysteine does not inhibit the synthesis of S-adenosylhomocysteine, and homocysteine does not inhibit the synthesis of S-adenosylcysteine. This is consistent with independent binding of cysteine and homocysteine to both enzyme subunits. Using adenosine analogs and the mammalian S-adenosylhomocysteinase we were able to synthesize a number of novel S-adenosylcysteine analogs. These included: S-N 6- hydroxyadenosyl- l- cysteine , S-2- aminoadenosyl- l- cysteine , S- nebularyl- l- cysteine , S-3- deazaadenosyl- l- cysteine , S- formycyl- l- cysteine , S-N 6- methyladenosyl- l- cysteine and S-N 1- oxideadenosyl- ll- cysteine .