Skip to Main Content
 

Global Search Box

 
 
 
 

Files

File List


osu1281719548.pdf (1.86 MB)

ETD Abstract Container

Abstract Header

Protein Engineering Surface Residues of the Four-helix Bundle Protein Rop: Evaluating the Relationship of Sequence-Stability through High-Throughput Approaches

Nguyen, Chau Q.
http://rave.ohiolink.edu/etdc/view?acc_num=osu1281719548

Abstract Details

2010, Master of Science, Ohio State University, Chemistry.
Recent developments in the field of rational engineering have increased the awareness that surface residues and surface electrostatics of proteins are more important for protein stability than previously thought. In this study, we have analyzed the complicated sequence-stability-function relationship of the four helix-bundle protein Rop through combinatorial engineering of surface residues. Taking advantage of the function-structure relationship, we are able to screen a large library for functional variants with assumed native-like structure, based on a novel cell-based screen with GFP reporter. High-throughput thermal scanning (HTTS) and colony-based sequencing provide data to analyze the thermodynamic consequence of mutations. The data from 72 active variants reveal a number of interesting results. According to the expression screen, the four-helix bundle Rop displays some degree of tolerance to hydrophobic residues on its solvent-exposed region. Due to the simplicity of Rop’s structure (entirely helix), charge neutralization and especially charge reversal substitutions do not seem to affect the protein stability. Moreover, it is encouraging to find a number of highly stabilized variants through rational and combinatorial engineering of surface residues. By using a four-helix bundle as a model protein, our results indicate great potential for stabilizing proteins with surface mutations; and the sites of stabilization can be selected so they do not interfere with protein function. Establishing a solid conclusion for the sequence-stability relationship of the surface mutants requires further ongoing work regarding high resolution structure analysis.
Thomas Magliery, PhD (Advisor)
Ross Dalbey, PhD (Committee Member)
103 p.
Chemistry

Recommended Citations

Citations

  • Nguyen, C. Q. (2010). Protein Engineering Surface Residues of the Four-helix Bundle Protein Rop: Evaluating the Relationship of Sequence-Stability through High-Throughput Approaches [Master's thesis, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1281719548

    APA Style (7th edition)

  • Nguyen, Chau. Protein Engineering Surface Residues of the Four-helix Bundle Protein Rop: Evaluating the Relationship of Sequence-Stability through High-Throughput Approaches. 2010. Ohio State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1281719548.

    MLA Style (8th edition)

  • Nguyen, Chau. "Protein Engineering Surface Residues of the Four-helix Bundle Protein Rop: Evaluating the Relationship of Sequence-Stability through High-Throughput Approaches." Master's thesis, Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1281719548

    Chicago Manual of Style (17th edition)

osu1281719548
491
© 2010, all rights reserved.
This open access ETD is published by The Ohio State University and OhioLINK.