# Structural insights into the substrate binding mechanism of the class I dehydratase MadB

## TL;DR

This study reveals how the enzyme MadB binds and processes a specific antibiotic precursor, offering insights into its structure and function.

## Contribution

The paper provides the first structural and functional analysis of the MadB dehydratase with its substrate.

## Key findings

- Cryo-EM structures show MadB undergoes a conformational change when binding its substrate.
- Small-angle X-ray scattering identifies the C-terminal binding site of the maddinglicin leader peptide.
- A specific amino acid stretch in the leader sequence is critical for MadB substrate recognition.

## Abstract

In the battle against antimicrobial resistance, lantibiotics have emerged as promising new sources for antimicrobial drugs. Their exceptional stability is due to characteristic modifications termed (methyl-)lanthionine rings. Genome mining efforts have identified hundreds of lantibiotics by detecting gene operons, so-called biosynthetic gene clusters (BGC), which encode cysteine-rich peptides (30-50 amino acids in size) and enzymes responsible for dehydration and cyclization, catalyzing the post-translational ring formation. One such identified, class I lantibiotic is maddinglicin from Clostridium maddingley. Here, we present single particle cryo-EM structures of the dehydratase MadB in both, its apo-state and in complex with a leader peptide of maddinglicin, revealing a distinct conformational change upon substrate binding. Small-angle X-ray scattering studies elucidate the substrate binding site for the C-terminal part of maddinglicin. Furthermore, a substrate specificity analysis was performed highlighting a critical stretch of amino acids within the maddinglicin leader sequence that is crucial for binding. Here, we provide molecular insights into the conformational changes, principles of substrate recognition and ligand:protein stoichiometry of a class I lantibiotic dehydratase.

Class I lantibiotic dehydratases play a key role in the biosynthesis of ribosomally synthesized peptide antibiotics. We present structural insights into MadB in two states, revealing the conformational changes that underlie its catalytic mechanism during maddinglicin maturation.