Inhibition of serine and proline racemases by substrate-product analogues.

Abstract d -Amino acids can play important roles as specific biosynthetic building blocks required by organisms or act as regulatory molecules. Consequently, amino acid racemases that catalyze the formation of d -amino acids are potential therapeutic targets. Serine racemase catalyzes the reversible formation of d -serine (a modulator of neurotransmission) from l -serine, while proline racemase (an essential enzymatic and mitogenic protein in trypanosomes) catalyzes the reversible conversion of l -proline to d -proline. We show the substrate-product analogue α-(hydroxymethyl)serine is a modest, linear mixed-type inhibitor of serine racemase from Schizosaccharomyces pombe ( K i  = 167 ± 21 mM, K i ′ = 661 ± 81 mM, cf. K m  = 19 ± 2 mM). The bicyclic substrate-product analogue of proline, 7-azabicyclo[2.2.1]heptan-7-ium-1-carboxylate is a weak inhibitor of proline racemase from Clostridium sticklandii , giving only 29% inhibition at 142.5 mM. However, the more flexible bicyclic substrate-product analogue tetrahydro-1 H -pyrrolizine-7a(5 H )-carboxylate is a noncompetitive inhibitor of proline racemase from C. sticklandii ( K i  = 111 ± 15 mM, cf. K m  = 5.7 ± 0.5 mM). These results suggest that substrate-product analogue inhibitors of racemases may only be effective when the active site is capacious and/or plastic, or when the inhibitor is sufficiently flexible.