INHIBITION OF THE INTRINSIC NAD(+) GLYCOHYDROLASE ACTIVITY OF CD38 BYCARBOCYCLIC NAD ANALOGS

Carba-NAD and pseudocarba-NAD are carbocyclic analogues of NAD(+) in w hich a 2,3-dihydroxycyclopentane methanol replaces the beta-D-ribonucl eotide ring of the nicotinamide riboside moiety of NAD(+) [Slama and S immons (1988) Biochemistry 27, 183-193]. These carbocyclic NAD(+) anal ogues, related to each other as diastereomers, have been tested as inh ibitors of the intrinsic NAD(+) glycohydrolase activity of human CD38, dog spleen NAD(+) glycohydrolase, mouse CD38 and Aplysia californica cADP-ribose synthetase. Pseudocarba-NAD, the carbocyclic dinucleotide in which L-2,3-dihydroxycyclopentane methanol replaces the D-ribose of the nicotinamide riboside moiety of NAD(+), was found to be the more potent inhibitor. Pseudocarba-NAD was shown to inhibit the intrinsic N AD(+) glycohydrolase activity of human CD38 competitively, with K-i= 1 48 mu M determined for the recombinant extracellular protein domain an d K-1 = 180 mu M determined for the native protein expressed as a cell -surface enzyme on cultured Jurkat cells. Pseudocarba-NAD was shown to be a non-competitive inhibitor of the purified dog spleen NAD(+) glyc ohydrolase, with K-is = 47 mu M and K-ii =198 mu M. Neither pseudocarb a-NAD nor carba-NAD inhibited mouse CD38 or Aplysia californica cADP-r ibose synthetase significantly at concentrations up to 1 mM. The resul ts underscore significant species differences in the sensitivity of th ese enzymes to inhibition, and indicate that pseudocarba-NAD will be u seful as an inhibitor of the enzymic activity of human but not mouse C D38 in studies using cultured cells.