Combinatorial library approach to decipher the stability effects of loop mutagenesis in model protein scaffold via high throughput methods

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dc.contributor.advisor Magliery, Thomas J.
dc.creator Sen, Shiladitya
dc.date.accessioned 2012-04-24T17:22:07Z
dc.date.available 2012-04-24T17:22:07Z
dc.date.issued 2012-02
dc.identifier.uri http://hdl.handle.net/1811/51768
dc.description Mathematical and Physical Sciences: 3rd Place (The Ohio State University Edward F. Hayes Graduate Research Forum) en_US
dc.description.abstract The inability to accurately predict the relationship between protein’s primary sequence and its overall stability presents a major hurdle in determining the effects of mutations on protein folding. To study the effects of mutation, combinatorial libraries that involve randomizing many selected positions of a protein simultaneously, offer a broader, statistically relevant dataset. In this study, we have analyzed this complicated sequence-stability relationship using rigorous high-throughput (HT) methods. Our studies focus on the four-helix model bundle protein Rop through the means of combinatorial repacking the loop. The loop was randomized with and without an additional residue to make it more flexible. Using a novel screen we identified functional variants of Rop that were of similar or higher in stability than the parent scaffold. HT biophysical characterization and detailed Gibbs-Helmholtz studies were carried out to probe the entropy and enthalphy contribution on stability for selected variants. A potential salt bridge interaction was necessary to enhance the stability of the protein without affecting its function or fold. en_US
dc.language.iso en_US en_US
dc.relation.ispartofseries 2012. Edward F. Hayes Graduate Research Forum. 26th en_US
dc.subject Combinatorial en_US
dc.subject High-throughput en_US
dc.subject Stability en_US
dc.subject Loop mutagenesis en_US
dc.subject Gibbs-Helmholtz en_US
dc.subject protein folding en_US
dc.title Combinatorial library approach to decipher the stability effects of loop mutagenesis in model protein scaffold via high throughput methods en_US
dc.type Article en_US
dc.description.embargo A five-year embargo was granted for this item. en_US