Sillerico T, Valdivia M, de Ioannes A, Barros C
Laboratory of Embryology, Laboratory of Immunology, Faculty of Biological Sciences, Pontifical Catholic University of Chile, Santiago, Chile.
Biocell. 1996 Aug;20(2):133-42.
The proacrosin/acrosin system has been immunolocalized, by the silver enhanced immunogold technique on the acrosomal region of capacitated and perivitelline rabbit spermatozoa. The purpose of the present work was to investigate the kinetics of the activation of acrosin by determining the proportion of proacrosin and acrosin present in the acrosome of the rabbit spermatozoa during capacitation and the induction of the acrosome reaction by the calcium ionophore A23187. Rabbit spermatozoa selected by the percoll gradient technique were incubated for 0, 0.5 and 6 hours and then the acrosome reaction was induced at 0 and 6 hours with 1.9 microM of the calcium ionophore A23187. It was found that 95% of acrosin activity in rabbit spermatozoa at zero time corresponds to proacrosin and after capacitation and acrosome reaction, a diminution of the activity of proacrosin/acrosin system was found. However, proacrosin represents the large majority of the system activity. Western blot prepared with sperm extract obtained at the start of incubation showed the characteristic doublet band about 53-55 kDa that may correspond to proacrosin and alpha-acrosin. After six hours of incubation and with induction with the calcium ionophore A23187 the same doublet was seen in addition to a third band of 49 kDa that could correspond to a transition form between alpha-acrosin to beta-acrosin. In conclusion, rabbit proacrosin/acrosin system remains in the large proportion as proacrosin during capacitation and acrosome reaction.
采用银增强免疫金技术，已在获能的兔精子顶体区域和卵周隙精子顶体区域对前顶体蛋白酶/顶体蛋白酶系统进行了免疫定位。本研究的目的是通过测定兔精子在获能过程中顶体内前顶体蛋白酶和顶体蛋白酶的比例以及钙离子载体A23187诱导顶体反应的情况，来研究顶体蛋白酶激活的动力学。通过Percoll梯度技术筛选的兔精子分别孵育0、0.5和6小时，然后在0和6小时用1.9微摩尔的钙离子载体A23187诱导顶体反应。结果发现，兔精子在零时刻95%的顶体蛋白酶活性对应于前顶体蛋白酶，在获能和顶体反应后，前顶体蛋白酶/顶体蛋白酶系统的活性降低。然而，前顶体蛋白酶占该系统活性的绝大部分。用孵育开始时获得的精子提取物制备的蛋白质免疫印迹显示出特征性的约53 - 55 kDa的双峰条带，可能对应于前顶体蛋白酶和α-顶体蛋白酶。孵育6小时并用钙离子载体A23187诱导后，除了一条49 kDa的第三条带外，还可见到相同的双峰条带，该条带可能对应于α-顶体蛋白酶向β-顶体蛋白酶的转变形式。总之，在获能和顶体反应过程中，兔前顶体蛋白酶/顶体蛋白酶系统大部分仍以前顶体蛋白酶的形式存在。
