RIG: Toward the Development of an All Enzymatic Method to Generate Peptidomimetics

University Of Southern Mississippi, Hattiesburg MS

Investigators

Abstract

The long-term goal of this project is to develop a unique method to generate libraries of novel compounds that are likely to contain specific molecules that have important biological activities. These libraries will be composed or peptides with enhanced capabilities for in vivo applications. Peptides are ideally suited for in vivo studies (i.e. biomolecular imaging and gene delivery) due to their high target binding specificity, potency and low toxicity. As a result, peptides provide an effective means to investigate specific aspects of cellular pathways without disrupting critical cellular functions. Despite their inherent specificity and potency, peptides suffer from low stability and reduced membrane permeability -- factors that limit their biological stability and utility for in vivo studies. To overcome these limitations, a recombinant expression system (the "PURE" system) will be used to synthesize peptides composed of nonstandard or 'unnatural' amino acid building blocks. The premise for using unnatural amino acids is based upon the chemical composition of a potent class of natural products known as nonribosomal peptides (NRPs). NRPs contain a variety of unnatural residues that equip these biomolecules with enhanced functional capabilities. However, the synthetic pathways involved in NRP production are complex, and have not proven amenable to screening libraries for novel functionalities. Unlike conventional expression systems, the PURE system affords precise control over each of the reaction components required for protein/peptide synthesis. This allows potential substitution of the standard L-amino acids by a variety of unnatural amino acids to generate NRP-like peptides. Efficient in vitro assays (i.e. adenylation, nuclease protection, etc.) have been identified that can be used to determine quantitative criteria (binding affinity constants) that establish a threshold for translation of a substrate. With these criteria in hand, diverse peptide libraries composed of L- and unnatural amino acids will be synthesized. The research project merges well established and emerging cell expression techniques; the intellectual and scientific merits of this translation system could greatly enhance the development and economic expression of peptide diagnostics and reagents that will impact the field of chemical biology in general. Broader Impacts The project will also offer educational training and experience in basic research to the students of The University of Southern Mississippi (USM). As a member of an underrepresented minority group, the principle investigator is keenly aware of the need to increase the level of exposure and participation of minority groups in science. At USM there is a significant pool of students from underrepresented minority groups that the PI can involve in research and mentor. This summer an underrepresented undergraduate student participating in the Research Experience for Undergraduate (REU) program at USM will participate in the research project. The principle investigator also currently mentors students from the Alliance for Graduate Education in Mississippi (AGEM). A primary goal of AGEM is to increase the number of underrepresented minorities entering academe in STEM related fields. To raise the level of scientific literacy of the public through outreach activities, the PI will partner with the eMerging Careers Institute (ECI) to disseminate scientific information through an online publication designed to impact youth and young adults. ECI provides high school and junior college youth of Alameda County with pathways to explore career options, and compete in the job market. ECI is developing a similar program in Mississippi and the PI will work with ECI on the development of its scientific focus. The PI believes that these activities will be an effective means to increase the participation of underrepresented and underserved groups to help benefit science and society as a whole.