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Design and Study of Novel Antimicrobial Peptides with Proline Substitution

He, Jing
http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1257779581

Abstract Details

2009, Doctor of Philosophy (PhD), Ohio University, Chemistry and Biochemistry (Arts and Sciences).
Microorganism-related diseases and their resistance to conventional antibiotics are proliferating at an alarming rate and becoming a severe clinical problem. Therefore, it is urgent to develop novel approaches in antimicrobial therapy. Most living organisms produce and utilize at least some small peptides as part of their defensive system in combating infections by virulent pathogens. Research focusing on the structure and function of these antimicrobial peptides from diverse sources has gained a great number of interests in the past three decades. Generally, many naturally existing antimicrobial peptides are positively charged and have the potential to adopt either amphipathic α-helix or β-sheet conformation. In this project, based on preliminary studies of β-sheet-forming peptides developed in our lab, analogs were designed to investigate the effects of introducing a single leucine-to-proline substitution on the structure and function of the peptides. The leucine-to-proline substitution was selected as a structural perturbation that could influence the antimicrobial and the cytolytic activity toward mammalian cells of these peptides. A series of experiments were performed in this project to investigate these potential changes, beginning with the determination of the antimicrobial activity and hemolytic activity of these peptides. Peptide conformation was determined by circular dichroism spectroscopy. Membrane permeability changes in both synthetic lipid bilayers and bacterial membranes were assessed by measuring peptide-induced calcein leakage from large unilamellar vesicles (LUV) and by peptide-induced entry of o-nitrophenyl-β-D-galactopyranoside into E.coli ML-35 cells. The ability of peptides to bind to lipid bilayers of defined composition was measured by tryptophan fluorescence enhancement, acrylamide quenching and 10-doxylnonadecane quenching. The activity of these peptides was further studied by measuring the planktonic bacterial cell killing (the live vs. dead bacterial viability), bacterial biofilm formation inhibition and inhibition of bacterial cells growth within established bacterial biofilms. The results show that the position of proline has a significant influence on the antimicrobial and hemolytic activity of these peptides. Some of these proline-containing analogs show high antimicrobial activity and good selectivity between bacterial vs. mammalian cells. In addition, the peptide binding studies suggest that at least some of the proline-containing peptides may kill bacteria by mechanisms other than simply inducing membrane leakage. In summary, amphipathic β-sheet-forming antimicrobial peptides with proline substitutions appear to be promising models for novel antimicrobial agents.
John Blazyk, PhD (Advisor)
Douglas Goetz, PhD (Committee Member)
Jennifer Hines, PhD (Committee Member)
Marcia Kieliszewski, PhD (Committee Member)
Hugh Richardson, PhD (Committee Member)
228 p.
Biochemistry
antimicrobial peptides; amphipathic beta-sheet forming; proline substitution; bacterial biofilm

Recommended Citations

Citations

  • He, J. (2009). Design and Study of Novel Antimicrobial Peptides with Proline Substitution [Doctoral dissertation, Ohio University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1257779581

    APA Style (7th edition)

  • He, Jing. Design and Study of Novel Antimicrobial Peptides with Proline Substitution. 2009. Ohio University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1257779581.

    MLA Style (8th edition)

  • He, Jing. "Design and Study of Novel Antimicrobial Peptides with Proline Substitution." Doctoral dissertation, Ohio University, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1257779581

    Chicago Manual of Style (17th edition)

ohiou1257779581
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© 2009, all rights reserved.
This open access ETD is published by Ohio University and OhioLINK.