Proteolytic enzymes. Papain-catalysed hydrolysis of specific aryl esters

Abstract

Aryl hippurates (I) and mesylglycinates (II) are hydrolysed in the presence of papain and have constant k₀ parameters of ca. 4 s^–1 and 12 s^–1 respectively at pH 6·00, 35° and 0·3M ionic strength. The low sensitivity of k₀/K_m to change in phenyl substituent [ρ= 0·47 and 0·78 for (II) and (I) respectively] is argued to arise from electrophilic assistance on the carbonyl oxygen of the ester during acylation but the involvement of imidazolium ion (of histidine-159) as the electrophile is discounted. Evidence is reported for the involvement of a cationic acid (pK_aca. 4) in acylation and deacylation and a neutral acid (pK_aca. 8) in acylation. 4-Nitrophenyl acetate (III) is a substrate of papain with a typical bell-shaped profile for k₀/K_m; the ester is shown to differ from specific substrates only in its electronic reactivity.