Kinin-generating and esterolytic activity of purified human urinary kallikrein (urokallikrein).
Kinin-generating and esterolytic activity of purified human urinary kallikrein (urokallikrein).
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纯化的人尿激肽释放酶（urokallikrein）的激肽生成和酯解活性。
DOI：
10.1016/0006-2952(77)90163-0
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发表时间：
1977
影响因子：
5.8
通讯作者：
J. Spragg
中科院分区：
文献类型：
--
作者：
Onesmo ole;K. Austen;J. Spragg
尿激肽释放酶 (urokallikrein) 的定义是其从热灭活血浆或纯化的人激肽原中产生激肽的能力，是从新鲜浓缩男性人尿中分离出来的，并被证明是一种抗原性独特的尿对甲苯磺酰基-L-精氨酸甲酯盐酸盐 (TAMe) 酯酶。该分离程序实现了每毫克蛋白质激肽生成活性的 400 至 576 倍纯化，并产生了白蛋白作为等电聚焦步骤中唯一显着污染物的产品。纯化的尿激肽释放酶由其激肽生成活性定义，其等电点范围为 pH 3.9 至 4.2，具有电荷异质性，Sephadex 凝胶过滤的表观分子量为 25,000-40,000，碱性圆盘凝胶上的阳极迁移率。尿激肽释放酶从圆盘凝胶中洗脱出来，并通过激肽生成活性在兔子体内引发单特异性抗血清进行鉴定。纯化的尿激肽释放酶是一种 TAMe 酯酶，这从激肽生成活性、与显示识别尿激肽释放酶的驴抗胰血清的抗原反应性以及等电聚焦后评估的酯解能力的一致性中可以明显看出。通过增加 TAMe 或苯甲酰基-L-精氨酸甲酯盐酸盐 (BAMe) 的剂量，可以抑制纯化的尿激肽释放酶的酯解活性，并且用 Dixon 图分析这些数据表明存在底物抑制作用。
Urinary kallikrein (urokallikrein), as defined by its capacity to generate kinin from heat-inactivated plasma or from purified human kininogen, was isolated from fresh concentrated male human urine and shown to be an antigenically unique urinaryp-tosyl-L-arginine methyl ester HCl (TAMe) esterase. The isolation procedure achieved a 400- to 576-fold purification of the kinin-generating activity/mg of protein and yielded a product with albumin as the only significant contaminant at the isoelectric focusing step. The purified urokallikrein, defined by its kinin-generating activity, exhibited an isoelectric point with a range from pH 3.9 to 4.2 with charge heterogeneity, an apparent molecular weight of 25,000–40,000 on Sephadex gel filtration, and an anodal mobility on alkaline disc gels. Urokallikrein eluted from disc gels and identified by kinin-generating activity elicited monospecific antiserum in the rabbit. That purified urokallikrein is a TAMe esterase was evident from the concordance of kinin-generating activity, antigenic reactivity with a donkey antipancreas serum shown to recognize urokallikrein and esterolytic capacity as assessed after isoelectric focusing. There was suppression of the esterolytic activity of purified urokallikrein by increasing doses of TAMe or benzoyl-L-arginine methyl ester HCl (BAMe), and analysis of these data with Dixon plots indicated substrate inhibition.