# Development of an inhibitory monoclonal nanobody targeting Streptococcus pyogenes siderophore binding protein FtsB

## TL;DR

Researchers developed a nanobody that inhibits a key iron transporter in Streptococcus pyogenes, offering a new tool to study infection and potential antibacterial therapy.

## Contribution

A novel inhibitory nanobody targeting the FtsB protein in S. pyogenes was developed and characterized.

## Key findings

- Nb1 binds to FtsB with sub-nM affinity and a slow dissociation rate.
- The nanobody competitively inhibits hydroxamate siderophore binding and uptake in S. pyogenes.
- Nb1 provides a new tool to study the FtsABCD transporter's role in infection.

## Abstract

Due to the limited availability of metals inside the human body, pathogenic bacteria must produce multiple highly specialized metal transporters to cause infection. These transporters constitute attractive targets for developing novel antibacterial strategies. Streptococcus pyogenes possesses three iron transporters, of which the FtsABCD system is specialized in the uptake of ferric hydroxamates. The role of this transporter in infection remains unclear. In this study, we developed a monoclonal alpaca VHH, or nanobody, Nb1, targeting FtsB. Nb1 binds to FtsB with sub-nM affinity, in an enthalpy-driven manner, and with a characteristically slow dissociation rate. Solvent accessibility analysis by hydrogen/deuterium exchange coupled with mass spectrometry, mutational analyses, and X-ray crystallography revealed that the epitope of Nb1 is in the binding pocket of FtsB. The nanobody competitively inhibited the binding of multiple hydroxamate siderophores and partially inhibited the uptake of siderophores in S. pyogenes cells. The inhibitory activity of Nb1 on siderophore transport represents a new tool to study the role of the FtsABCD transporter and can be used as a potential inhibitor of S. pyogenes growth under iron-limited conditions.