Studies of QSOX, a medically important flavoenzyme: mechanistic insights, searching for inhibitors, and a potential new diagnostic assay
| Author(s) | Israel, Benjamin A. | |
| Date Accessioned | 2022-12-09T17:02:11Z | |
| Date Available | 2022-12-09T17:02:11Z | |
| Publication Date | 2014 | |
| SWORD Update | 2022-08-11T19:08:26Z | |
| Abstract | The Quiescin sulfhydryl oxidase (QSOX) enzymes are facile catalysts of disulfide bond formation in reduced, unfolded proteins. They are a family of flavoenzymes with emerging, medically-relevant biological roles. Recent studies have demonstrated the utility of QSOX as a diagnostic biomarker for pancreatic cancer and certain types of heart failure, and several reports have begun to characterize the roles of QSOX in promoting the invasiveness of a number of different cancers. In the first portion of this work we explore an in-depth understanding the catalytic mechanism of the simplest of the QSOX enzymes, from Trypanosoma brucei. After developing a novel substrate analog, we obtained results that suggest that QSOX does not have a significant prototypical substrate binding site. Methods were also developed to measure, for the first time, the redox potentials of all three redox centers of QSOX. This study revealed an unexpected redox potential mismatch in the mechanism. Mutagenesis studies provided data suggesting a novel mechanism that QSOX and the unrelated DsbA/B system use to bypass apparent thermodynamic barriers to catalysis. Next, work toward the discovery and development of QSOX inhibitors is discussed. Two new, sensitive assays for the enzymes are developed and utilized for high-throughput screening of small molecule libraries. The negative results of this screening effort, combined with our earlier mechanistic studies led to our pursuance of antibody-based inhibitors. Early achievements toward this collaborative project are presented. Finally we adapt and implement the new, sensitive, fluorescence-based assay for QSOX activity in biological samples. This study found, surprisingly, that QSOX activity is relatively abundant in normal adult blood, and provides implications for the utilization of our assay for diagnostic purposes. | |
| Advisor | Thorpe, Colin | |
| Degree | Ph.D. | |
| Department | University of Delaware, Department of Chemistry and Biochemistry | |
| DOI | https://doi.org/10.58088/ss4y-5c70 | |
| Unique Identifier | 1350917389 | |
| URL | https://udspace.udel.edu/handle/19716/31630 | |
| Language | en | |
| Publisher | University of Delaware | |
| URI | https://login.udel.idm.oclc.org/login?url=https://www.proquest.com/dissertations-theses/studies-qsox-medically-important-flavoenzyme/docview/2708570349/se-2?accountid=10457 | |
| Keywords | Sulfhydryl oxidase | |
| Keywords | Redox potential | |
| Keywords | QSOX | |
| Keywords | Trypanosoma brucei | |
| Keywords | Sulfide | |
| Keywords | Fluorescence-based assay | |
| Title | Studies of QSOX, a medically important flavoenzyme: mechanistic insights, searching for inhibitors, and a potential new diagnostic assay | |
| Type | Thesis |