Peptidyl (acyloxy)methyl ketones and the quiescent affinity label concept: the departing group as a variable structural element in the design of inactivators of cysteine proteinases.
(Acyloxy)methyl ketones, of general structure Z-[AA2]-[AA1]-CH2OCOAr, are potent inactivators of the cysteine proteinase cathepsin B. These reagents have been designed as affinity labels in which the dipeptidyl moiety serves as an affinity group (complementary to the S1 and S2 sites of the enzyme), while the (acyloxy)methyl ketone unit (-COCH2OCOR), containing a weak leaving group in the form of a carboxylate nucleofuge, functions as the potentially reactive entity that labels the enzyme. The inhibition is time dependent, active site directed, and irreversible. The apparent second-order rate constant kinact/Kinact, which characterizes the inhibition of cathepsin B by this series, spans several orders of magnitude and in certain cases exceeds 10(6) M-1 s-1. The activity of this series of inhibitors was found to be exquisitely sensitive to the nature of the carboxylate leaving group as well as the affinity group. A strong dependence of second-order inactivation rate on leaving group pKa was uncovered for Z-Phe-Ala (acyloxy)methyl ketones [log(k/K) = 1.1 (+/- 0.1) X pKa + 7.2 (+/- 0.4); r2 = 0.82, n = 26]. Heretofore in constructing affinity labels the choice of leaving group was quite restricted. The aryl carboxylate group thus offers considerable variation as a design element in that both its binding affinity and reactivity can be controlled by substituent effects. Specific peptidyl (acyloxy)methyl ketones thus represent prime examples of highly potent, chemically stable enzyme inhibitors with variable structural elements in both the affinity and departing groups.
Krantz
A
A
Syntex Research, Mississauga, Ontario, Canada.
Copp
L J
LJ
Coles
P J
PJ
Smith
R A
RA
Heard
S B
SB
D000345:Affinity Labels / Q000737:chemistry*; D015853:Cysteine Proteinase Inhibitors; D004151:Dipeptides / Q000737:chemistry*; D011480:Protease Inhibitors / Q000138:chemical synthesis / Q000737:chemistry*
D000345:Affinity Labels; D015853:Cysteine Proteinase Inhibitors; D004151:Dipeptides; D011480:Protease Inhibitors; C039553:N-benzyloxycarbonylphenylalanylalanine