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Hao Wang Dissertation.pdf (3.67 MB)

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Functional Studies of N-glycosylation in Human Corin

Wang, Hao
http://orcid.org/0000-0001-6881-9977
http://rave.ohiolink.edu/etdc/view?acc_num=csu1500479066332159

Abstract Details

2017, Doctor of Philosophy in Clinical-Bioanalytical Chemistry, Cleveland State University, College of Sciences and Health Professions.
Corin is a transmembrane serine protease essential for salt-water balance and normal blood pressure. Human corin has 19 N-glycosylation sites. In this study, we investigated the role of N-glycosylation in corin biosynthesis. First, we investigated the role of N-glycosylation in corin cell surface expression, zymogen activation and ectodomain shedding. We mutated each of the 19 N-glycosylation sites and characterized the corin mutants in functional studies. Our results show that N-glycosylation at Asn-80 and -231 protected corin from ectodomain shedding. N-glycosylation at Asn-697 and -1022 promoted corin cell surface expression and zymogen activation. Next, we analyzed the functional importance of N-glycosylation in the corin protease domain. We generated corin mutants that lacked the transmembrane domain or extracellular domains in the pro-peptide region and chimeric corin mutants containing the protease domain from a different serine protease. We also extended our study to include prothrombin, factor VII and enterokinase. Our results show that the function of N-glycosylation in the protease domain in promoting corin cell surface expression is independent of the pro-peptide domains and that N-glycosylation in the protease domain also promoted the secretion of prothrombin and factor VII and the cell surface expression of enterokinase. Finally, we performed proteomic and cellular studies to understand how N-glycans in the protease domain promote corin cell membrane targeting. Our results show that N-glycans in the protease domain facilitated corin trafficking in the endoplasmic reticulum (ER). Moreover, we found that corin proteins lacking the N-glycosylation site in the protease domain were bound to calnexin and BiP in the ER. These data indicate that N-glycans in the protease domain of corin are important in calnexin-assisted protein folding and ER exiting. We also show that such a mechanism may apply to the function of N-glycans in other trypsin-like serine proteases such as prothrombin, factor VII and enterokinase. Together, our results show that N-glycans at different sites of corin protein play distinct roles in regulating corin folding, intracellular trafficking, cell surface expression, zymogen activation and ectodomain shedding. Our results also suggest that N-glycosylation may have similar roles in the protein synthesis and posttranslational modifications of other trypsin-like serine proteases.
Qingyu Wu, Ph.D. (Advisor)
Xue-Long Sun, Ph.D. (Committee Chair)
Anthony Berdis, Ph.D. (Committee Member)
Michael Kalafatis, Ph.D. (Committee Member)
Bin Zhang, Ph.D. (Committee Member)
Biochemistry; Chemistry; Molecular Biology

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Citations

  • Wang, H. (2017). Functional Studies of N-glycosylation in Human Corin [Doctoral dissertation, Cleveland State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=csu1500479066332159

    APA Style (7th edition)

  • Wang, Hao. Functional Studies of N-glycosylation in Human Corin. 2017. Cleveland State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=csu1500479066332159.

    MLA Style (8th edition)

  • Wang, Hao. "Functional Studies of N-glycosylation in Human Corin." Doctoral dissertation, Cleveland State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=csu1500479066332159

    Chicago Manual of Style (17th edition)

csu1500479066332159
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© 2017, all rights reserved.
This open access ETD is published by Cleveland State University and OhioLINK.