Briseid K, Hoem N O, Johannesen S, Marthinsen K
Department of Pharmacology, University of Oslo, Blindern, Norway.
Thromb Res. 1990 Mar 15;57(6):945-56. doi: 10.1016/0049-3848(90)90161-5.
Plasma kallikrein and FXIIa were assayed in acetone-treated human citrated plasma (CPLa) with the chromogenic peptide Bz-Ile-Glu-Gly-Arg-pNA (S-2222) as substrate. In end point assays with short incubation periods (1-10 min.) nearly all kallikrein present could be blocked by a low concentration of soybean trypsin inhibitor (STI). In 30 min. assays the main part of the kallikrein was recovered in a functional state not inhibited by STI, and at the same time the level of FXIIa (as amidase activity blocked by corn inhibitor, C.I.) was reduced to about 2/3 of the initial value. The formation of an association between FXIIa and kallikrein is suggested. In fractions from gel filtration of CPLa kallikrein was assayed as S-2302 amidase, high molecular weight kininogen (HK) was measured in rocket immunoassays, and HK and FXII were studied in PAGE immunoblot experiments. Kallikrein appeared as one peak together with HK (gel mol. wt. 300 KD), about 40% of HK was free (220 KD), and no FXII was observed in the kallikrein or HK peaks, but in two areas corresponding to 78-79 KD and 39-42 KD. When experiments, however, were carried out with plasma acetone-activated and gel filtered in the presence of benzamidine (5 mM), part of the amidase activity present in kallikrein peak fractions was blocked by C.I. This observation supports the above suggestion of an association between FXIIa and kallikrein.
以发色肽Bz-Ile-Glu-Gly-Arg-pNA（S-2222）为底物，在丙酮处理的人枸橼酸盐血浆（CPLa）中检测血浆激肽释放酶和FXIIa。在短孵育期（1 - 10分钟）的终点测定中，几乎所有存在的激肽释放酶都可被低浓度的大豆胰蛋白酶抑制剂（STI）阻断。在30分钟的测定中，激肽释放酶的主要部分以未被STI抑制的功能状态被回收，同时FXIIa的水平（以被玉米抑制剂C.I.阻断的酰胺酶活性计）降至初始值的约2/3。提示FXIIa与激肽释放酶之间形成了一种结合。在CPLa的凝胶过滤组分中，激肽释放酶作为S-2302酰胺酶进行检测，在火箭免疫测定中测量高分子量激肽原（HK），并在PAGE免疫印迹实验中研究HK和FXII。激肽释放酶与HK（凝胶分子量300 KD）一起呈现为一个峰，约40%的HK是游离的（220 KD），在激肽释放酶或HK峰中未观察到FXII，但在对应于78 - 79 KD和39 - 42 KD的两个区域中观察到。然而，当在苯甲脒（5 mM）存在下对血浆进行丙酮激活和凝胶过滤实验时，激肽释放酶峰组分中存在的部分酰胺酶活性被C.I.阻断。这一观察结果支持了上述关于FXIIa与激肽释放酶之间结合的提示。
