Kominami E, Hoffschulte H, Leuschel L, Maier K, Holzer H
Biochim Biophys Acta. 1981 Sep 15;661(1):136-41. doi: 10.1016/0005-2744(81)90092-9.
The substrate specificity of proteinase B (EC 3.4.22.9) from Baker's yeast was studied. Experiments with unblocked synthetic peptides indicated that the enzyme has no aminopeptidase activity. The proteinase cleaves trypsin substrates like Bz-Arg-OEt, Bz-Arg-pNA and Bz-Ile-Glu-Gly-Arg-pNA and chymotrypsin substrates like Ac-Tyr-OEt and Bz-Tyr-pNA. The Km value for Ac-Tyr-OEt is similar to that of chymotrypsin A, but the catalytic activity per mol proteinase B amounts to only 1/20 that of chymotrypsin A. Km and kcat for Bz-Arg-OEt are 1/50 and 1/7 as high as the corresponding values determined for trypsin. Proteinase B cleaved the oxidized insulin B chain with an initial rapid cleavage step at Leu(15)-Tyr(16) and Phe(24)-Phe(25). Slower hydrolysis was observed at Gln(4)-His(5), Leu(11)-Val(12) Tyr(16)-Leu(17), Leu(17)-Val(18), Arg(22)-Gly(23) and Phe(25)-Tyr(26). These results suggest that the specificity of proteinase B is comparable to the specificity of porcine chymotrypsin C as well as of trypsin. When the hexapeptide Leu-Trp-Met-Arg-Phe-Ala was used as a substrate for proteinase B, the enzyme preferentially attacked at Arg-Phe and more slowly at Trp-Met.
对面包酵母中的蛋白酶B（EC 3.4.22.9）的底物特异性进行了研究。对未封闭的合成肽进行的实验表明，该酶没有氨肽酶活性。该蛋白酶可切割胰蛋白酶底物，如Bz-Arg-OEt、Bz-Arg-pNA和Bz-Ile-Glu-Gly-Arg-pNA，以及胰凝乳蛋白酶底物，如Ac-Tyr-OEt和Bz-Tyr-pNA。Ac-Tyr-OEt的Km值与胰凝乳蛋白酶A的相似，但每摩尔蛋白酶B的催化活性仅为胰凝乳蛋白酶A的1/20。Bz-Arg-OEt的Km和kcat分别是胰蛋白酶相应值的1/50和1/7。蛋白酶B切割氧化胰岛素B链时，最初在Leu(15)-Tyr(16)和Phe(24)-Phe(25)处快速切割。在Gln(4)-His(5)、Leu(11)-Val(12)、Tyr(16)-Leu(17)、Leu(17)-Val(18)、Arg(22)-Gly(23)和Phe(25)-Tyr(26)处观察到较慢的水解。这些结果表明，蛋白酶B的特异性与猪胰凝乳蛋白酶C以及胰蛋白酶的特异性相当。当六肽Leu-Trp-Met-Arg-Phe-Ala用作蛋白酶B的底物时，该酶优先在Arg-Phe处攻击，在Trp-Met处攻击较慢。
