Relationship between N-linked oligosaccharides in SEA module and proteolytic cleavage within C-terminal domain of rodent Muc3
- VernacularTitle:Muc3羧基端内SEA组件的蛋白酶切与N糖基化关系的研究
- Author:
Yicheng LI
;
Yonghong HE
;
Zhihong PENG
;
Rongquan WANG
- Publication Type:Journal Article
- Keywords:
Mucin 3;
proteolytic cleavage;
SEA module;
glycosylation
- From:Journal of Third Military Medical University
2003;0(21):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective The C-terminal domain of rodent Muc3 is proteolytically cleaved.This study is to explore the relationship between N-linked oligosaccharides in SEA module and the proteolytic cleavage within C-terminal domain of rodent Muc3.Methods Truncated rodent Muc3 C-terminal domains with complete SEA module(p20SEA) were produced by site-directed mutagenesis to insert a stop code in the required place.Proteins were detected by pulse/chase and immunoprecipitation method,or SDS/PAGE and Western blot.Inhibition of glycosylation of the expressed protein was performed by using tunicamycin.Results Muc3 C-terminal domain was posttranslationally cleaved to produce a V5-tagged 30 000 extracellular glycopeptide and a Myc-tagged 49 000 membrane-associated glycopeptide.Treatment with tunicamycin to transfected COS-1 cells led to the abundant production of 60 000 uncleaved and whole-length Muc3 C-terminal domain,the 30 000 N-terminal fragment shifted to 22 000 and 49 000 C-terminal fragment shifted to 41 000 after deglycosylation.The truncated Muc3 C-terminal domain containing complete SEA module but without the following residues led to production of 36 000 uncleaved and whole-length protein,and 30 000 cleaved product shifted to 22 000 after deglycosylation.Conclusion Proteolytic cleavage in both complete rodent C-terminal domain and complete SEA module without the following residues were partially inhibited by tunicamycin.
