Irreversible Inhibition of Transglutaminases by Sulfonium Methylketones: Optimization of Specificity and Potency with w-Aminoacyl Spacers

Abstract

Sulfonium methylketones, of structure Cbz-Phe-NH(CH₂)_nCOCH₂S ⁺ (CH₃)₂, n < 2, are specific and potent inactivators of transglutaminases. The length of the -(CH₂)_n-spacer moiety, n = 1-5, is a critical determinant for both the specificity and potency of the inactivator. The dipeptidyl analog Cbz-Phe-Gly-(CH₂)_nS ⁺ (CH₃)₂, n = 1, is a more powerful inactivator of the thiol proteinase cathepsin B, k/K < 3 × 10⁵ M^?1 min^?1, than of transglutaminases, k_i(appl/K_i(appl < 1.5 × 10⁴ M^?1 min^?1. In contrast, the γ-aminobutyryl analog, n = 3, is a very potent transglutaminase inactivator with k_i(apP/ K_i(appl = 3.1 < 10⁶M^?1min^?1, but does not inactivate cathepsin B. In cell studies, the y-aminobutyryl and w-aminohexyl analogs inhibited the transglutaminase-mediated process of ionophore-induced cross-linked envelope formation by human malignant keratinocytes and the order of potency was related to that found for enzyme inhibition. The sulfonium methylketones, in equilibrium with the resonance stabilized ylides, are chemically inert towards glutathione under ambient conditions demonstrating the potential utility of this novel class of transglutaminase inhibitors for the study of enzyme inhibition in cellular environments.