Characterization of Glycocomponents and Protein p38 in Mouse Sperm Using Lectins

• Main Authors: CHEN, XUAN-REN, 陳炫任
• Other Authors: LIU, MIN
• Format: Others
• Language: zh-TW
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/52105298443289614671

碩士 === 中國文化大學 === 生物科技研究所 === 104 === Testicular sperm are not motile and have to undergo extratesticular maturation in epididymis. Nevertheless, freshly ejaculated sperm cannot fertilize oocytes. As sperm reside in female reproductive tract, a final step of maturation, termed capacitation, takes place and sperm become fully fertilizable. As sperm approach oocytes, the extracellular matrix, zona pellucida glycoproteins, surrounding the oocytes induce sperm to undergo acrosome reaction. This exocytotic event allows sperm to release enzymes that facilitate sperm to move forward prior to gamete fusion. Recent studies, using carbohydrates-binding lectins, have demonstrated that sperm glycocomponents participate in sperm maturation, capacitation, acrosome reaction, and fertilization. The purpose of this study is to investigate the presence and the distribution of sperm specific galactosylated, N-acetylgalactosamylated and N-acetylglucosamylated glycocomponents during epididymal maturation, prior to and following capacitation and acrosome reaction. Fluorescent staining study using three lectins, PNA, SBA, WGA, showed that the presence of galactosylated, N-acetylgalactosamylated and N-acetylglucosamylated glycocomponents and these glycosylated materials distributed in sperm head region. Following extratesticular maturation and development, sperm glycocomponents underwent relocalization and they were differentially expressed. Using PNA affinity chromatography, we isolated fifteen sperm galactosylated glycoproteins for LC/MS MS protein identification and our spectrometric results showed that a sperm specific protein, p38, was present. In epididymal sperm, two major protein bands with molecular weight close to 38kDa and 40kDa were identified by antibody specific to p38. Following capacitation and acrosome reaction, p38 was only identified as a single protein band and there was no significant change in p38 protein expression, compare to that of in epididymal sperm. Finally, p38 was localized to the sperm head region and underwent redistribution following acrosome reaction.