# Discovery and characterization of biocatalysts for ribosomally encoded alpha-N-methylated peptide natural products

> **NIH NIH R35** · UNIVERSITY OF MINNESOTA · 2020 · $359,849

## Abstract

PROJECT SUMMARY / ABSTRACT
 Since the discovery of penicillin in the 1920s, bioactive peptide natural products have been used as
antibiotic, antiviral, immunosuppressive, and anti-cancer agents. Many of these bioactive peptides harbor
backbone α-N-methylations and/or macrocyclizations, since these tailorings significantly improve peptide
pharmacokinetics. As seen in the blockbuster immunosuppressant cyclosporin A, α-N-methylated peptides have
increased structural rigidity, proteolytic resistance, and membrane permeability. Additionally, cyclic α-N-
methylated peptides are able to bind large, flat surfaces with high affinity, making them attractive targets for
disrupting protein-protein interactions. Despite these advantages, inefficient synthetic and in vitro processes
hinder the production, screening, and optimization of α-N-methylated peptides. In addition, natural sources of
amide backbone-methylated peptides were thought to be completely limited to inflexible nonribosomal peptide
biosynthetic pathways. The goal of this work is to tease out the mechanistic and structural constraints of α-N-
methylating biocatalysts within our newly discovered ribosomally encoded peptide natural product family called
the borosins. Our first objective identifies a potent bioactive model system encoded in a basidiomycete fungus
to tease out the rules and limitations of borosin α-N-methylation, N-to-C macrocyclization, and bioactivity. Our
second objective focuses on structurally distinct borosin family members, where we will perform detailed kinetic
measurements to tease out the mechanism for chemically challenging α-N-methylation. We also outline our
longer-term visions to develop methods for studying peptide natural products in basidiomycete fungi as well as
our efforts to tease out the potential biological roles of borosin metabolites. This research will create a diverse
toolbox of flexible and efficient catalysts for the biological production, screening, and optimization of genetically
templated bioactive α-N-methylated peptides.

## Key facts

- **NIH application ID:** 9980950
- **Project number:** 5R35GM133475-02
- **Recipient organization:** UNIVERSITY OF MINNESOTA
- **Principal Investigator:** Michael F Freeman
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $359,849
- **Award type:** 5
- **Project period:** 2019-08-01 → 2024-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9980950

## Citation

> US National Institutes of Health, RePORTER application 9980950, Discovery and characterization of biocatalysts for ribosomally encoded alpha-N-methylated peptide natural products (5R35GM133475-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9980950. Licensed CC0.

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