R01AI162961

Advancing Ribosome-Targeting Antibacterial Peptides with a Unique Mechanism of Action - Project Summary

Apidaecin (API) and Drosocin (DRO) are proline-rich antimicrobial peptides (PRAMPs) produced by honeybees and fruit flies, respectively, which share a unique mechanism of action. Our previous studies of API showed that upon entering gram-negative bacterial cells through the SBMA transporter, API binds in the exit tunnel of ribosomes that have just released the newly made protein and arrests the ribosomes at stop codons by trapping the associated tRNA and release factor. As such, API represents the first-ever described specific inhibitor of translation termination.

Our subsequent whole-genome studies revealed that arresting terminating ribosomes triggers several downstream events that accentuate the inhibitory action of this PRAMP, including ribosome queuing and readthrough of stop codons. Our preliminary data indicate that DRO, despite its distinct amino acid sequence, inhibits the termination step of translation as well, by a mechanism likely resembling that of API. Their idiosyncratic mode of binding to the target, the unique mechanism of action, and the triggering of downstream effects harmful for the bacterial cell make these antibacterial peptides an attractive model for developing novel antibiotics.

Furthermore, the biological nature of these PRAMPs opens unique opportunities for their screening and optimization by generating hundreds of thousands of peptide variants directly in bacterial cells. In the current proposal, we will use the combined effort of three laboratories with expertise in biochemistry and genomics of ribosomal antibiotics, in peptide chemistry, and in structural analysis of ribosome-antibiotic complexes to advance the fundamental understanding of the mechanism of action of API- and DRO-like translation termination inhibitors and identify derivatives with superior on-target activity and expanded spectrum of antibacterial action.

In order to achieve these goals, we will test arrays of API and DRO variants in bacterial cells by the tunable expression of peptide gene libraries, determine high-resolution X-ray crystal structures of ribosome-peptide complexes, and employ rational structure-based design to generate via chemical synthesis peptide variants with superior properties.

Specifically, in Aim 1, we will identify API-derived peptides with improved activity upon ribosomes from gram-negative and gram-positive pathogens. In Aim 2, the spectrum of action of API-like peptides will be expanded by bypassing the necessity for uptake by the SBMA transporter. Finally, in Aim 3, we will analyze the ribosome binding and mechanism of action of DRO-like peptides and use comparative analysis to identify the key features that define the class of antimicrobial peptides that target translation termination. The three aims are tightly interconnected but completely independent from each other.

The reagents and tools that will be generated in the course of the proposed work are aimed to serve as leads for future clinical development. Importantly, the results obtained in the proposed studies will significantly advance the fundamental understanding of the properties and mechanisms of action of PRAMPs and will stimulate the progress of the field of ribosome-targeting antibacterial peptides, which currently is still in its infancy.

University Of Illinois

TO ASSIST PUBLIC AND PRIVATE NONPROFIT INSTITUTIONS AND INDIVIDUALS TO ESTABLISH, EXPAND AND IMPROVE BIOMEDICAL RESEARCH AND RESEARCH TRAINING IN INFECTIOUS DISEASES AND RELATED AREAS, TO CONDUCT DEVELOPMENTAL RESEARCH, TO PRODUCE AND TEST RESEARCH MATERIALS. TO ASSIST PUBLIC, PRIVATE AND COMMERCIAL INSTITUTIONS TO CONDUCT DEVELOPMENTAL RESEARCH, TO PRODUCE AND TEST RESEARCH MATERIALS, TO PROVIDE RESEARCH SERVICES AS REQUIRED BY THE AGENCY FOR PROGRAMS IN INFECTIOUS DISEASES, AND CONTROLLING DISEASE CAUSED BY INFECTIOUS OR PARASITIC AGENTS, ALLERGIC AND IMMUNOLOGIC DISEASES AND RELATED AREAS. PROJECTS RANGE FROM STUDIES OF MICROBIAL PHYSIOLOGY AND ANTIGENIC STRUCTURE TO COLLABORATIVE TRIALS OF EXPERIMENTAL DRUGS AND VACCINES, MECHANISMS OF RESISTANCE TO ANTIBIOTICS AS WELL AS RESEARCH DEALING WITH EPIDEMIOLOGICAL OBSERVATIONS IN HOSPITALIZED PATIENTS OR COMMUNITY POPULATIONS AND PROGRESS IN ALLERGIC AND IMMUNOLOGIC DISEASES. BECAUSE OF THIS DUAL FOCUS, THE PROGRAM ENCOMPASSES BOTH BASIC RESEARCH AND CLINICAL RESEARCH. SMALL BUSINESS INNOVATION RESEARCH (SBIR) PROGRAM EXPANDS AND IMPROVES PRIVATE SECTOR PARTICIPATION IN BIOMEDICAL RESEARCH. THE SBIR PROGRAM INTENDS TO INCREASE AND FACILITATE PRIVATE SECTOR COMMERCIALIZATION OF INNOVATIONS DERIVED FROM FEDERAL RESEARCH AND DEVELOPMENT, TO INCREASE SMALL BUSINESS PARTICIPATION IN FEDERAL RESEARCH AND DEVELOPMENT, AND TO FOSTER AND ENCOURAGE PARTICIPATION OF SOCIALLY AND ECONOMICALLY DISADVANTAGED SMALL BUSINESS CONCERNS AND WOMEN-OWNED SMALL BUSINESS CONCERNS IN TECHNOLOGICAL INNOVATION. THE SMALL BUSINESS TECHNOLOGY TRANSFER (STTR) PROGRAM STIMULATES AND FOSTERS SCIENTIFIC AND TECHNOLOGICAL INNOVATION THROUGH COOPERATIVE RESEARCH AND DEVELOPMENT CARRIED OUT BETWEEN SMALL BUSINESS CONCERNS AND RESEARCH INSTITUTIONS, TO FOSTER TECHNOLOGY TRANSFER BETWEEN SMALL BUSINESS CONCERNS AND RESEARCH INSTITUTIONS, TO INCREASE PRIVATE SECTOR COMMERCIALIZATION OF INNOVATIONS DERIVED FROM FEDERAL RESEARCH AND DEVELOPMENT, AND TO FOSTER AND ENCOURAGE PARTICIPATION OF SOCIALLY AND ECONOMICALLY DISADVANTAGED SMALL BUSINESS CONCERNS AND WOMEN-OWNED SMALL BUSINESS CONCERNS IN TECHNOLOGICAL INNOVATION. RESEARCH CAREER DEVELOPMENT AWARDS SUPPORT THE DEVELOPMENT OF SCIENTISTS DURING THE FORMATIVE STAGES OF THEIR CAREERS. INDIVIDUAL NATIONAL RESEARCH SERVICE AWARDS (NRSAS) ARE MADE DIRECTLY TO APPROVE APPLICANTS FOR RESEARCH TRAINING IN SPECIFIED BIOMEDICAL SHORTAGE AREAS. IN ADDITION, INSTITUTIONAL NATIONAL RESEARCH SERVICE AWARDS ARE MADE TO ENABLE INSTITUTIONS TO SELECT AND MAKE AWARDS TO INDIVIDUALS TO RECEIVE TRAINING UNDER THE AEGIS OF THEIR INSTITUTIONAL PROGRAM.

93.855 - Allergy and Infectious Diseases Research

National Institute of Allergy and Infectious Diseases (NIAID) [HHS - NIH]

Chicago, Illinois 60612 United States

Single Zip Code

NIH Research Project Grant (Parent R01 Clinical Trial Not Allowed) (PA-20-185)

Amendment Since initial award the total obligations have increased 601% from $454,250 to $3,183,300.