Syntheses, Structures, and Enzymatic Evaluations of Conformationally Constrained, Analogue Inhibitors of Carnitine Acetyltransferase: (2R,6R)-, (2S,6S)-, (2R,6S)-, and (2S,6R)-6-(Carboxylatomethyl)-2-(hydroxymethyl)-2,4,4-trimethylmorpholinium

Article abstract of DOI:10.1021/jo00126a018

The syntheses and structures of the four stereoisomers of 6-(carboxylatomethyl)-2-(hydroxymethyl)-2,4,4-trimethylmorpholinium, 1, are described.The key step in the synthetic strategy involves an intramolecular Michael addition reaction.Condensation of nonracemic 3-(methylamino)-2-methylpropane-1,2-diol, 3, with methyl 4-bromo-2-butenoate followed by intramolecular Michael addition gives a mixture of two diastereomers of methyl 2-<4,6-dimethyl-6-(hydroxymethyl)morpholinyl>acetate, 5.The diastereomeric ratio of the products in this reaction changes from 6:1 to 1:1 with a change in solvent from 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 carboxylatomethyl occupying an equatorial position.All four stereoisomers inhibit pigeon breast carnitine acetyltransferase (CAT).Of this series, (2S,6R)-1 binds to CAT most strongly with a K_i of 190 +/- 20 μM and an IC 50 of 0.42 mM.The enzymatic assays of 1 confirm that CAT recognizes both configurations at C2 and C6 in the analogues.CAT has a different conformation when it binds carnitine or acetylcarnitine than when it binds 1.This latter conformation may resemble that when CAT catalyzes acetyl transfer.