# Mechanistic studies of peptide modifying radical S-adenosylmethionine enzymes PqqE and MftC

> **NIH NIH F32** · UNIVERSITY OF CALIFORNIA AT DAVIS · 2021 · $17,141

## Abstract

Project Summary/Abstract (Mark Nesbit)
 The superfamily of radical S-adenosylmethionine (SAM) enzymes (RSEs) are responsible for catalyzing
a wide variety of unusual and difficult chemical reactions which are critical for the survival of living organisms
from bacteria to humans. RSEs are identified by binding of a redox active [4Fe-4S] cluster through three
cysteine residues (typically from a CX3CX2C sequence). The initial step in RSE catalyzed reactions where
SAM binds to the [4Fe-4S] cluster and is reduced to generate Ado• (5’-deoxyadenosyl radical) appears to be
common to all RSEs. However, despite having many structural similarities and sharing a common initiation
step RSEs are able to catalyze a wide variety of chemical transformations and are involved in peptide
modification, metalloprotein cluster assembly and cofactor biosynthesis. Two RSEs involved in peptide
modification are PqqE and MftC. PqqE catalyzes a C-C bond forming step in the biosynthesis of the redox
cofactor PQQ. MftC catalyzes the oxidative decarboxylation of the C-terminus tyrosine residue in the MftA
peptide as a part of the biosynthesis of the proposed redox cofactor mycofactocin. Studying the mechanisms
by which these genetically related enzymes are able to catalyze vastly different chemical reactions will provide
valuable insight into the varied functionality of RSEs in nature.
 The planned investigation will interrogate the mechanism of the C-C bond forming step in PQQ
biosynthesis catalyzed by the radical SAM enzyme PqqE and the oxidative decarboxylation of a C-terminus
tyrosine residue catalyzed by MftC. Continuous wave EPR, and pulsed methods such as ENDOR will be able to
intimately probe the nature of potential radical organic and organometallic intermediates. The use of non-natural
amino acids designed to stabilize potential radical intermediates and incorporated into analogs of the peptide
substrates will assist in these experiments. Additional spectroscopic methods such as rapid freeze-quench 57Fe
Mössbauer spectroscopy will provide data which complements the EPR studies allowing for observation of all
Fe nuclei in a sample under turnover conditions. The data produced by these experiments will help to further the
mechanistic understanding of the diverse array of biological reactions catalyzed by radical SAM enzymes and
provide additional characterization of physical and electronic structures of the PqqE and MftC under catalytically
relevant conditions. Additionally, these methods may be used to study other RSEs and will provide a strong
spectroscopic foundation of knowledge about the mechanisms by which this diverse family of enzymes
accomplishes the unique biological roles they have adapted to fill.

## Key facts

- **NIH application ID:** 10373580
- **Project number:** 3F32GM126628-03S1
- **Recipient organization:** UNIVERSITY OF CALIFORNIA AT DAVIS
- **Principal Investigator:** Mark A Nesbit
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $17,141
- **Award type:** 3
- **Project period:** 2018-05-01 → 2021-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10373580

## Citation

> US National Institutes of Health, RePORTER application 10373580, Mechanistic studies of peptide modifying radical S-adenosylmethionine enzymes PqqE and MftC (3F32GM126628-03S1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10373580. Licensed CC0.

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