Baudys M, Meloun B, Gan-Erdene T, Fusek M, Mares M, Kostka V, Pohl J, Blake C C
Institute of Organic Chemistry and Biochemistry, Czechoslovak Academy of Sciences, Prague, CSFR.
Biomed Biochim Acta. 1991;50(4-6):569-77.
Bovine spleen cathepsin B contains 7 disulfide bridges. Using different chemical and enzymatic cleavage methods we isolated fragments representing the individual disulfides: Cys14-Cys43, Cys26-Cys71, Cys62-Cys128, Cys63-Cys67, Cys100-Cys132, Cys108-Cys119, and Cys148-Cys252. A similar line of approach was applied to determine the S-S bridges of bovine spleen cathepsin H: Cys23-Cys66, Cys57-Cys99, Cys157-Cys207, and Cys212-Cys5A, where Cys5A is located in the propart portion of the procathepsin H chain. On the basis of the knowledge of the S-S bridges of cathepsin B a novel sequence alignment of papain and cathepsin B has been proposed. This enabled us to construct a reasonable 3D-model of cathepsin B and propose the region (a 18 residue insertion between Glu89 and Gly90 of papain) responsible for the carboxypeptidase activity of cathepsin B functioning as a "closure". A similar approach was applied to explain the aminopeptidase activity of cathepsin H. A general model of steric regulation of accessibility of the preformed "endopeptidase-like" binding cleft by distant parts of the polypeptide chain of the proteinases discussed is proposed as a factor determining the mode of binding and thus cleavage of polypeptide substrates.
牛脾组织蛋白酶B含有7个二硫键。我们使用不同的化学和酶切方法分离出了代表各个二硫键的片段：半胱氨酸14-半胱氨酸43、半胱氨酸26-半胱氨酸71、半胱氨酸62-半胱氨酸128、半胱氨酸63-半胱氨酸67、半胱氨酸100-半胱氨酸132、半胱氨酸108-半胱氨酸119以及半胱氨酸148-半胱氨酸252。我们采用类似的方法来确定牛脾组织蛋白酶H的二硫键：半胱氨酸23-半胱氨酸66、半胱氨酸57-半胱氨酸99、半胱氨酸157-半胱氨酸207以及半胱氨酸212-半胱氨酸5A，其中半胱氨酸5A位于组织蛋白酶H原链的前肽部分。基于对组织蛋白酶B二硫键的了解，我们提出了木瓜蛋白酶和组织蛋白酶B的一种新的序列比对。这使我们能够构建一个合理的组织蛋白酶B三维模型，并提出负责组织蛋白酶B羧肽酶活性的区域（木瓜蛋白酶谷氨酸89和甘氨酸90之间18个残基的插入片段）起到“封闭物”的作用。我们采用类似的方法来解释组织蛋白酶H的氨肽酶活性。本文提出了一个通用模型，即所讨论的蛋白酶多肽链的远端部分对预先形成的“类内肽酶”结合裂隙可及性的空间调节，作为决定多肽底物结合模式进而切割模式的一个因素。
