Emerging biotechnology to detect weak and/or transient protein-protein interactions
| dc.contributor.advisor | Zhang, Zhiwen Jonathan | |
| dc.creator | Thibodeaux, Gabrielle Nina | en |
| dc.date.accessioned | 2014-04-30T17:23:43Z | en |
| dc.date.accessioned | 2017-05-11T23:01:28Z | |
| dc.date.available | 2014-04-30T17:23:43Z | en |
| dc.date.available | 2017-05-11T23:01:28Z | |
| dc.date.issued | 2009-12 | en |
| dc.description | text | en |
| dc.description.abstract | Protein-protein interactions are of great importance to a number of essential biological processes including cell cycle regulation, cell-cell interactions, DNA replication, transcription and translation. Thus, an understanding of protein-protein interactions is critical for understanding many facets of cell function. Unfortunately, the tools and methods currently in use to identify and study protein-protein interactions focus largely on high affinity, stable interactions. However, the majority of the protein-protein interactions involved in regulatory processes have weak affinities and are transient in nature. Therefore, it is important to develop new biotechnology capable of detecting weak and/or transient protein-protein interactions in vivo. Here, we describe four new methods that allow for the identification and study of weak and/or transient protein-protein interactions in vivo. First, we developed a rapid method to convert Escherichia coli orthogonal tRNA/synthetase pairs into an orthogonal system for mammalian cells in order to site-specifically incorporate unnatural amino acids into any gene of interest using stop codon suppression. This method will allow the expression and purification of proteins that carry normally transient post-translational modifications. Second, we successfully employed site-specific unnatural amino acid incorporation to chemically cross-link a known homodimer, Sortase A, in vivo. Third, we developed a novel tetracycline repressor-based mammalian two-hybrid system and successfully detected homo- and hetero-dimers that are known to have weak binding constants. Finally, a synthetic antibody (termed a synbody) that binds weakly to the SH3 domain of the proto-oncogene Abelson tyrosine kinase was developed. The synbody can potentially be used as a first generation drug and/or biomarker. We hope that the methods developed in this dissertation will enable the scientific community to better understand weak/transient protein-protein interactions in vivo. | en |
| dc.description.department | Pharmacy | en |
| dc.format.medium | electronic | en |
| dc.identifier.uri | http://hdl.handle.net/2152/24366 | en |
| dc.language.iso | eng | en |
| dc.rights | Copyright is held by the author. Presentation of this material on the Libraries' web site by University Libraries, The University of Texas at Austin was made possible under a limited license grant from the author who has retained all copyrights in the works. | en |
| dc.subject | Protein-protein interactions | en |
| dc.subject | In vivo | en |
| dc.subject | Unnatural amino acid incorporation | en |
| dc.subject | Weak/transient interactions | en |
| dc.title | Emerging biotechnology to detect weak and/or transient protein-protein interactions | en |
| dc.type | Thesis | en |