Sperm-surface chymotrypsin-like protease activity required for fertilization in ascidians.
Sperm-surface chymotrypsin-like protease activity required for fertilization in ascidians.
复制标题
海鞘受精所需的精子表面胰凝乳蛋白酶样蛋白酶活性。
DOI：
10.1006/dbio.1994.1100
复制
发表时间：
1994
影响因子：
2.7
通讯作者：
Lambert,CC
中科院分区：
文献类型：
--
作者：
Koch,RA;Norton,ML;Vazquez,H;Lambert,CC
During fertilization, species-specific gamete binding must be followed by sperm penetration of egg vestments before gamete fusion can occur. Sperm proteases, called lysins, aid this process. Sperm fromAscidia ceratodes, Ascidia callosa, andAscidia paratropawere found to have a surface-mounted chymotrypsin-like protease when studied by enzymology, biotinylation, immunolabeling, and histochemistry. Chymotrypsin substrates and inhibitors blocked fertilization in a concentration-dependent manner in A. ceratodes and decreased the number of sperm heads which penetrated the egg's vitelline coat, but had no effect on sperm binding to follicle cells. Sperm bound to agarose beads coated with the chymotrypsin inhibitor α2-macroglobulin. Chymotrypsin-like enzyme activity, assayed fluorimetrically usingN-succinyl-leucinyl-leucinyl-valinyl-tyrosinyl-7-amido-4-methyl-coumarin as the substrate, was associated with head fractions prepared by differential centrifugation. Biotinylation of live sperm followed by detergent extraction showed that chymotrypsin-like activity could be removed from the detergent extract using avidinagarose beads. Indirect immunofluorescence of unreacted and reacted sperm heavily labeled membrane domains overlying the mitochondrion and at the base of the head with occasional labeling of the sperm tip. Histochemical studies, which usedN-succinyl-alanyl-alanyl-prolyl-phenylalanyl-β-napthylamide as the substrate, colocalized enzyme activity in head regions of unreacted and near the mitochondrion of reacted sperm. Thus, we conclude that in ascidian sperm a chymotrypsin-like protease is exposed on the external surface of the plasma membrane of the head, is required for fertilization, and plays a role in sperm penetration but not binding.
