Characterization and localization of beta-1,4-N-acetyl-galactosaminyl transferase 2 in testicular cells

• Main Authors: Chech-Ang Liu, 劉傑昂
• Other Authors: Sin-tak chu
• Format: Others
• Language: zh-TW
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/88422582994308644629

碩士 === 國立臺灣大學 === 生化科學研究所 === 99 === The Sda β-1,4-N-acetyl-galactosaminyl transferase Ⅱ(B4GALNT2) catalyze the addition of GalNAc to the Gal of the [Neu5Acα2-3]Galβ1-4GlcNAcβ1-3Gal terminal structure via the β-1,4 linkage to form Sda antigen. The Sda antigen was first reported as a human blood surface antigen. This antigen was found in several tissue types, including stomach, colon, kidney, and oocytes. Its presence was also demonstrated in various body fluids, such as saliva, milk, serum, and urine. Current studies revealed that Sda antigen reduced remarkably in cancer lesions of the gastrointestinal tract. It indicated the significance of this antigen in biosystem and also associated it is importance to B4GALNT2. So far the research of B4GALNT2 physiological function is still unclear. Dell’s group first found Sda antigen in reproductive system, however, the significance of sda is unclear. Using RT-PCR and western blotting analysis to examine the expression of B4GALNT2 in mouse testis, and the results ¬¬revealed that two ¬¬¬genes expressed ¬¬higher in mature mice (12 WK) than in immature mice (3 WK). It suggested that B4GALNT2 is involved in mouse reproduction development. According to Shur’s study and the primary sequence prediction, it suggests that B4GALNT2 may be as a membrane protein. To identify the location of this membrane proteins, first we have extracted the total proteins from testis, and separated via SDS-PAGE, then analyzed by immunostaining. At the same time we overexpressed EGFP-b4galnt2 in 293T cell and observed by flurorescence microscope to detect B4GALNT2. In addition, the immunohistochemical analysis was performed in sertoli cell line (TM4) and leydig cell line (TM3) via B4GALNT2 antibody, and then observed under microscope. We found that B4GALNT2 is localized on plasma membrane by a sequence near its N-terminal, and the major structure may face to outside of the plasma membrane.