Sequential cleavage of proinsulin by human pancreatic kallikrein and a human pancreatic kininase.
Sequential cleavage of proinsulin by human pancreatic kallikrein and a human pancreatic kininase.
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人胰激肽释放酶和人胰激肽酶对胰岛素原的连续裂解。
DOI：
10.1073/pnas.76.8.3612
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发表时间：
1979
影响因子：
11.1
通讯作者：
K. Austen
中科院分区：
文献类型：
--
作者：
O. O. Yoi;D. Seldin;J. Spragg;G. Pinkus;K. Austen
通过免疫组织化学技术定位于胰腺的β细胞的胰腺内肽酶已经通过遵循其作为腺激肽释放酶的功能和抗原特征而纯化至同质（EC 3.4.21.8）。该酶在碱性圆盘凝胶电泳上产生一条染色带，其位置与从重复凝胶中洗脱的激肽产生活性相对应，凝胶过滤的分子量为54，000，没有酪蛋白溶解活性，在兔中引起单特异性抗血清，并产生一条与胰腺提取物、粗尿和纯化的尿激肽释放酶。胰腺激肽释放酶从牛和猪胰岛素原产生三种增加阳极流动性的裂解产物，并且胰激肽酶或牛羧肽酶B的存在增加了这些产物的量。虽然转化产物并不对应于difuryl-和monocurylinsulin，产品的中间流动性也得到了当胰岛素原处理低浓度的胰蛋白酶在存在的激肽。迁移最快的产物确实对应于参比标准品中的desalanylinsulin。单独的激肽酶对胰岛素原没有作用，抑肽酶单独或与激肽酶联合使用可防止激肽释放酶裂解。尽管胰岛素原裂解产物的化学结构尚未确定，但人胰激肽释放酶被认为是胰岛素原的假定激活剂，因为其位于β细胞中，与偶氮酪蛋白相比，其对胰岛素原和激肽原的优先作用，以及其产生胰岛素中间产物的能力，所述胰岛素中间产物被人胰激肽酶或牛羧肽酶B进一步修饰。
A pancreatic endopeptidase localized to the beta-cells of the pancreas by immunohistochemical techniques has been purified to homogeneity by following its functional and antigenic characteristics as a glandular kallikrein (EC 3.4.21.8). The enzyme gave a single stained band on alkaline disc gel electrophoresis which corresponded in location with the kinin-generating activity eluted from a replicate gel, was of 54,000 molecular weight by gel filtration, was devoid of caseinolytic activity, elicited a monospecific antiserum in a rabbit, and gave a line of complete identity with a single constituent in pancreatic extract, crude urine, and purified urokallikrein when analyzed with monospecific antibody to urokallikrein. The pancreatic glandular kallikrein generated three cleavage products of increasing anodal mobility from bovine and porcine proinsulin, and the presence of pancreatic kininase or bovine carboxypeptidase B increased the quantity of these products. Although the conversion products did not correspond to diarginyl- and monoarginylinsulin, the product of intermediate mobility was also obtained when proinsulin was treated with a low concentration of trypsin in the presence of kininase. The most rapidly migrating product did correspond to desalanylinsulin in the reference standard. Kininase alone had no action on proinsulin, and aprotinin prevented cleavage by kallikrein alone or in combination with kininase. Although the chemical structure of the proinsulin cleavage products has not been established, human pancreatic kallikrein is considered a putative activator of proinsulin because of its location in the beta-cell, its preferential action on proinsulin and kininogen as compared to azocasein, and its capacity to generate insulin intermediate products that are further modified by human pancreatic kininase or bovine carboxypeptidase B.