Cho K
Enzyme Systems Products, Dublin, California 94568.
Anal Biochem. 1987 Jul;164(1):248-53. doi: 10.1016/0003-2697(87)90393-9.
Fluorescent peptidyl thioneamides are synthesized for the first time. The carbonyl oxygen of the scissile amide bond of the substrates was replaced by a sulfur atom. The proteolytic activities of trypsin and papain were measured against 5-(benzyloxycarbonyllysylthioamido)-isophthalic acid dimethyl ester (Z-Lys-psi[CS]-AIE) and 5-(benzyloxycarbonylphenylalanylarginylthioamido)-isophthalic++ + acid dimethyl ester (Z-Phe-Arg-psi[CS]-AIE) and were compared to the corresponding oxyamides. Kinetic constants were measured. With thioneamide substrates, no tryptic hydrolysis was observed. Papain, on the other hand, hydrolyzed both oxy and thioneamides. The Km values of the thioneamides were shown to be slightly lower for papain than for the oxyamides, but the efficiency of the overall catalytic activity was off set by the lower turnover number for the thio derivatives. With the present synthetic substrate technology, selective detection of cysteine proteases in the presence of serine proteases is difficult. The thioneamides reported here were hydrolyzed by papain alone in the presence of trypsin.
首次合成了荧光肽基硫代酰胺。底物可裂解酰胺键的羰基氧被硫原子取代。测定了胰蛋白酶和木瓜蛋白酶对5-(苄氧羰基赖氨硫代酰胺基)-间苯二甲酸二甲酯（Z-Lys-psi[CS]-AIE）和5-(苄氧羰基苯丙氨酰精氨硫代酰胺基)-间苯二甲酸二甲酯（Z-Phe-Arg-psi[CS]-AIE）的蛋白水解活性，并与相应的氧酰胺进行比较。测定了动力学常数。对于硫代酰胺底物，未观察到胰蛋白酶水解。另一方面，木瓜蛋白酶可水解氧酰胺和硫代酰胺。结果表明，木瓜蛋白酶对硫代酰胺的Km值略低于氧酰胺，但硫代衍生物较低的周转数抵消了其整体催化活性的效率。利用目前的合成底物技术，在丝氨酸蛋白酶存在的情况下选择性检测半胱氨酸蛋白酶很困难。本文报道的硫代酰胺在胰蛋白酶存在下仅被木瓜蛋白酶水解。
