A new role for lipoproteins LpqZ and FecB in orchestrating mycobacterial cell envelope biogenesis

Abstract

Accounting for more deaths than any other bacterial species, Mycobacte rium tuberculosis (Mtb) represents a critical threat to public health worldwide. A key factor contributing to Mtb’s virulence is its unique cell envelope, which acts as a protective barrier. Among the components of this envelope, lipoproteins represent a critical but understudied group of proteins. In this study, we focused on 79 con served putative lipoproteins, shared between Mtb and the closely related M. marinum. Leveraging the CRISPR/Cas9 gene editing system for Mycobacteria, we generated frameshift mutations, targeting one conserved lipoprotein-coding gene at a time. We identified two mutants, lpqZ and fecB, that exhibited increased susceptibility to all tested antibiotics, suggesting critical roles in cell envelope biogenesis. Interestingly, despite having homology to periplasmic substrate-binding proteins (SBPs), neither protein is associated with any inner membrane transporter complex. Instead, co-immunoprecipita tion experiments revealed that LpqZ interacts with AftA and FecB interacts with AftB. Both these interaction partners are essential enzymes involved in arabinogalactan and lipoarabinomannan synthesis. Accordingly, we observed alterations for both glycoconju gates in lpqZ and fecB mutants. Together, these findings show that orphaned SBP-like proteins have been repurposed in mycobacteria to aid key enzymes involved in cell envelope biosynthesis.