Villegas V, Vendrell J, Avilés X
Departament de Bioquímica i Biologia Molecular, Institut de Biologia Fonamental, Universitat Autònoma de Barcelona, Bellaterra, Spain.
Protein Sci. 1995 Sep;4(9):1792-800. doi: 10.1002/pro.5560040914.
The activation process of porcine pancreatic procarboxypeptidase B (pro-CPB) has been studied in detail by a number of complementary methodologies, and a description of the molecular events that lead to the generation of active carboxypeptidase B (CPB) has been deduced. The generated CPB participates in the degradation of its own activation segment by excising C-terminal residues from fragments produced by tryptic proteolysis. The trimming action of CPB is, however, not essential for the release of a fully functional enzyme, in contrast to what was previously reported for porcine procarboxypeptidase A (pro-CPA). In the model presented here, the activation process is solely dependent on the first tryptic cleavage, at the limit between the activation segment and the enzyme region, and the former piece loses all of its inhibitory capacity once severed from the proenzyme. The use of heterologous inhibitors of CPB activity during the study of the tryptic activation process of pro-CPB has been required for the capture of short-lived, otherwise nondetectable, intermediates. This has allowed a complete description of the process and shown that the first proteolytic action of trypsin can also take place on a second target bond. Structural considerations that take into account the three-dimensional structures of the A and B forms of the proenzymes lead us to propose that the differences in conformation at the region that connects the globular activation domain to the enzyme are the main responsible elements for the differences observed in the activation processes of both proenzymes.
已经通过多种互补方法对猪胰羧肽酶原B（pro-CPB）的激活过程进行了详细研究，并推导了导致活性羧肽酶B（CPB）产生的分子事件的描述。生成的CPB通过从胰蛋白酶解产生的片段中切除C末端残基来参与其自身激活片段的降解。然而，与先前报道的猪羧肽酶原A（pro-CPA）不同，CPB的修剪作用对于释放完全功能性的酶不是必需的。在此提出的模型中，激活过程仅取决于在激活片段和酶区域之间的界限处的第一次胰蛋白酶切割，并且一旦从前体酶中切断，前一片段就失去了所有的抑制能力。在研究pro-CPB的胰蛋白酶激活过程中，需要使用CPB活性的异源抑制剂来捕获短暂存在的、否则无法检测到的中间体。这使得能够完整描述该过程，并表明胰蛋白酶的第一次蛋白水解作用也可以发生在第二个靶标键上。考虑到酶原A和B形式的三维结构的结构因素使我们提出，连接球状激活域与酶的区域的构象差异是两种酶原激活过程中观察到的差异的主要原因。
