SYNTHESES, STRUCTURES, AND ENZYMATIC EVALUATIONS OF CONFORMATIONALLY CONSTRAINED, ANALOG INHIBITORS OF CARNITINE ACETYLTRANSFERASE - YL)-2-(HYDROXYMETHYL)-2,4,4-TRIMETHYLMORPHOLINIUM, YL)-2-(HYDROXYMETHYL)-2,4,4-TRIMETHYLMORPHOLINIUM, YL)-2-(HYDROXYMETHYL)-2,4,4-TRIMETHYLMORPHOLINIUM, AND HYL)-2-(HYDROXYMETHYL)-2,4,4-TRIMETHYLMORPHOLINIUM

The syntheses and structures of the four stereoisomers of 6-(carboxyla tomethyl)-2-(hydroxymethyl)- 2,4,4-trimethylmorpholinium, 1, are descr ibed. The key step in the synthetic strategy involves an intramolecula r Michael addition reaction. Condensation of nonracemic 3-(methylamino )-2-methylpropane-1,2-diol, 3, with methyl 4-bromo-2-butenoate followe d by intramolecular Michael addition gives a mixture of two diastereom ers of methyl ,6-dimethyl-6-(hydroxymethyl)morpholinyl]-acetate, 5. Th e diastereomeric ratio ofthe products in this reaction changes from 6: 1 to 1:1 with a change in solvent fi om diethyl ether:methanol (35:1, v:v) to methanol. The structures and absolute configurations of 1 were determined by single crystal X-ray analyses. In crystals and solution , the morpholinium rings adopt a chair conformation with carboxylatome thyl occupying an equatorial position. All four stereoisomers inhibit pigeon breast carnitine acetyltransferase (CAT). Of this series, (2S,6 R)-1 binds to CAT most strongly with a Ki of 190 +/- 20 mu M and an IC 50 of 0.42 mM. The enzymatic assays of 1 confirm that CAT recognizes b oth configurations at C2 and C6 in the analogues. CAT has a different conformation when it binds carnitine or acetylcarnitine than when it b inds 1. This latter conformation may resemble that when CAT catalyzes acetyl transfer.