Studies towards the in vivo inhibition of oligosaccharyl transferase
Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2003.
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| Format: | Thesis |
| Language: | eng |
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Massachusetts Institute of Technology
2006
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| Online Access: | http://hdl.handle.net/1721.1/30018 |
| _version_ | 1811092537194053632 |
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| author | Ufret-Vincenty, María de L. (María de Lourdes), 1974- |
| author2 | Barbara Imperiali. |
| author_facet | Barbara Imperiali. Ufret-Vincenty, María de L. (María de Lourdes), 1974- |
| author_sort | Ufret-Vincenty, María de L. (María de Lourdes), 1974- |
| collection | MIT |
| description | Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2003. |
| first_indexed | 2024-09-23T15:19:47Z |
| format | Thesis |
| id | mit-1721.1/30018 |
| institution | Massachusetts Institute of Technology |
| language | eng |
| last_indexed | 2024-09-23T15:19:47Z |
| publishDate | 2006 |
| publisher | Massachusetts Institute of Technology |
| record_format | dspace |
| spelling | mit-1721.1/300182019-04-10T10:15:15Z Studies towards the in vivo inhibition of oligosaccharyl transferase Ufret-Vincenty, María de L. (María de Lourdes), 1974- Barbara Imperiali. Massachusetts Institute of Technology. Dept. of Chemistry. Massachusetts Institute of Technology. Dept. of Chemistry. Chemistry. Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2003. Vita. Includes bibliographical references. Protein glycosylation is an important process because of the great diversity of glycoproteins that can be produced by the introduction of different oligosaccharide sequences. Our group has made significant progress in the study of asparagine-linked glycosylation. This process is catalyzed by oligosaccharyl transferase (OT), which is a membrane-associated enzyme found in the lumen of the endoplasmic reticulum (ER). A variety of inhibitors that bind tightly to OT in vitro, with K[sub]i s as low as 10 nM, have been synthesized. The development of peptides capable of inhibiting OT in vivo would be desirable, since there is no bio-available inhibitor that targets N-linked glycosylation directly. Both active and passive strategies for delivering inhibitors into cells were studied. Internalization sequences were attached to the inhibitors to be used as delivery vectors. An ER retrieval sequence was used to target inhibitors to the ER and fluorescent labels were used to trace the inhibitors in the interior of the cell. It was determined that inhibitors with the internalization sequences, both with and without the ER retrieval sequence, were internalized by cells in culture. Also, inhibitors with a BODIPY fluorophore were internalized by cells in culture. Those that contained the C-terminus ER retrieval sequence were concentrated in the ER, while the inhibitors without the ER retrieval sequence were found distributed throughout the cell. Nevertheless, inhibitors with an amide C-terminus were internalized with more ease that those with a free acid at the C-terminus, which were the ones that contained the ER retrieval sequence. Compounds were assayed for in vivo inhibition of OT with a system in which the activity of secreted alkaline phosphatase is related to (cont.) the glycosylation state of this glycoprotein, since unglycosylated protein is not secreted from the cell. Some of the compounds containing internalization sequences were determined to be in vivo inhibitors of OT, as well as some of the compounds labeled with the BODIPY fluorophore. by María de L. Ufret-Vincenty. Ph.D. 2006-03-24T18:12:19Z 2006-03-24T18:12:19Z 2003 2003 Thesis http://hdl.handle.net/1721.1/30018 55030032 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/1721.1/7582 163 leaves 6461122 bytes 6460929 bytes application/pdf application/pdf application/pdf Massachusetts Institute of Technology |
| spellingShingle | Chemistry. Ufret-Vincenty, María de L. (María de Lourdes), 1974- Studies towards the in vivo inhibition of oligosaccharyl transferase |
| title | Studies towards the in vivo inhibition of oligosaccharyl transferase |
| title_full | Studies towards the in vivo inhibition of oligosaccharyl transferase |
| title_fullStr | Studies towards the in vivo inhibition of oligosaccharyl transferase |
| title_full_unstemmed | Studies towards the in vivo inhibition of oligosaccharyl transferase |
| title_short | Studies towards the in vivo inhibition of oligosaccharyl transferase |
| title_sort | studies towards the in vivo inhibition of oligosaccharyl transferase |
| topic | Chemistry. |
| url | http://hdl.handle.net/1721.1/30018 |
| work_keys_str_mv | AT ufretvincentymariadelmariadelourdes1974 studiestowardstheinvivoinhibitionofoligosaccharyltransferase |