Investigations of tetraspanin functions using large extracellular loops
Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2005.
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| Format: | Thesis |
| Language: | eng |
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Massachusetts Institute of Technology
2008
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| Online Access: | http://dspace.mit.edu/handle/1721.1/33751 http://hdl.handle.net/1721.1/33751 |
| _version_ | 1811098383313534976 |
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| author | Liu, Christopher C |
| author2 | Peter S. Kim and Richard O. Hynes. |
| author_facet | Peter S. Kim and Richard O. Hynes. Liu, Christopher C |
| author_sort | Liu, Christopher C |
| collection | MIT |
| description | Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2005. |
| first_indexed | 2024-09-23T17:14:12Z |
| format | Thesis |
| id | mit-1721.1/33751 |
| institution | Massachusetts Institute of Technology |
| language | eng |
| last_indexed | 2024-09-23T17:14:12Z |
| publishDate | 2008 |
| publisher | Massachusetts Institute of Technology |
| record_format | dspace |
| spelling | mit-1721.1/337512019-04-12T11:54:53Z Investigations of tetraspanin functions using large extracellular loops Liu, Christopher C Peter S. Kim and Richard O. Hynes. Massachusetts Institute of Technology. Dept. of Biology. Massachusetts Institute of Technology. Dept. of Biology. Biology. Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2005. Includes bibliographical references. This thesis describes our characterization of a specific tetraspanin domain: the large extracellular loop (LEL). Tetraspanins are involved in cellular migration, adhesion, and metastasis, sperm-egg fusion, and viral infectivity. The large extracellular loop domain is the major extracellular domain of tetraspanins and the binding of a monoclonal antibody against the tetraspanin CD9 serves to inhibit fertilization, consistent with the CD9-null mouse model. The first area of focus in this thesis is the characterization of the murine CD9-LEL domain. We present a methodology to express and purify the mCD9-LEL to homogeneity. Biophysical characterization of the mCD9-LEL protein reveals that it is an autonomously folding, [alpha]-helical dimer. Mutagenesis over much of the mCD9-LEL protein reveals that it is composed of two subdomains: a dimerization subdomain and a variable subdomain proposed to mediate heterotypic interactions. These results suggest both a means for exploring endogenous tetraspanins functions and a mechanism by which tetraspanins may oligomerize. Surprisingly, we were not able to detect oligomerization of the intact CD9 molecule, in discordance with our biophysical data on the mCD9-LEL. (cont.) In the latter part of this thesis, we expand our methodology to purify and characterize three different tetraspanins-LELs, the hCD9-LEL, the hCD63-LEL, and the hCD8 1-LEL. These tetraspanins-LELs all exhibit similar characteristics to the mCD9-LEL, consistent with a published crystal structure of the hCD81-LEL. Lastly, we demonstrate the ability of our tetraspanin-LEL proteins to bind integrins, to inhibit sperm-egg fusion, and to inhibit hepatitis C viral infectivity. Taken as a whole, these studies present novel, biophysically validated tetraspanins-LELs that lend insight into endogenous tetraspanins functions. by Christopher C. Liu. Ph.D. 2008-04-24T08:53:11Z 2008-04-24T08:53:11Z 2005 2005 Thesis http://dspace.mit.edu/handle/1721.1/33751 http://hdl.handle.net/1721.1/33751 65195713 eng M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission. http://dspace.mit.edu/handle/1721.1/33751 http://dspace.mit.edu/handle/1721.1/7582 224 leaves application/pdf Massachusetts Institute of Technology |
| spellingShingle | Biology. Liu, Christopher C Investigations of tetraspanin functions using large extracellular loops |
| title | Investigations of tetraspanin functions using large extracellular loops |
| title_full | Investigations of tetraspanin functions using large extracellular loops |
| title_fullStr | Investigations of tetraspanin functions using large extracellular loops |
| title_full_unstemmed | Investigations of tetraspanin functions using large extracellular loops |
| title_short | Investigations of tetraspanin functions using large extracellular loops |
| title_sort | investigations of tetraspanin functions using large extracellular loops |
| topic | Biology. |
| url | http://dspace.mit.edu/handle/1721.1/33751 http://hdl.handle.net/1721.1/33751 |
| work_keys_str_mv | AT liuchristopherc investigationsoftetraspaninfunctionsusinglargeextracellularloops |