Substrate Stereochemistry of 3‐Hydroxy‐3‐Methylglutaryl‐Coenzyme A Synthase

1 Synthetic specimens of R ‐[ 2 H 1 , 3 H 1 ]acetate and of S ‐[ 2 H 1 , 3 H 1 ]acetate were converted into coenzyme A thiolesters. 2 Each of these thiolesters was condensed with acetoacetyl‐CoA on 3‐hydroxy‐3‐methyl‐glutaryl‐CoA synthase. 3 The products, which were found to be hydroxymethylglutaryl‐phosphopantetheines owing to a secondary cleavage of the coenzyme A moieties, were reduced chemically to 3 R mevalonates. 4 Each mevalonate specimen, after mixing with [2‐ 14 C]mevalonate, was converted into squalene and cholesterol by a rat liver preparation. 5 Each cholesterol specimen was converted into androst‐1,4‐diene‐3,17‐dione by incubation with a strain of Mycobacterium phlei in the presence of an inhibitor. 6 When the procedures 4 and 5 were carried out with authentic specimens of 2 R ‐ and 2 S ‐[2‐ 3 H 1 ]mevalonate, it was found that the androstadienedione had lost nearly all tritium derived from the 2 S ‐mevalonate but had retained nearly all tritium (relative to 14 C) derived from the 2 R ‐mevalonate. 7 The androstadienedione derived, by procedures 1–5, from R ‐acetate showed a loss of more than half of its tritium (relative to 14 C). The androstadienedione derived by the same procedure from S ‐acetate lost less than half of its tritium. 8 It is concluded that the reaction on hydroxymethylglutaryl‐CoA synthase is stereospecific and is associated with an intramolecular hydrogen isotope effect. If this effect is normal ( k H / k D >1) then the condensation proceeds with inversion of configuration at the methyl group.