Biology subjects

Voedts, H.

Publications and source records attributed to Voedts, H..

4 recordsLinked to original sources

bioRxiv2026

Polysaccharide release factors drive lipid-carrier recycling in bacteria

Gram-negative bacteria build their cell envelope by transporting various sugar-based precursors across the inner membrane on the shared lipid carrier undecaprenyl (also known as C55). Because the cellular pool of undecaprenyl is finite and used by multiple competing pathways, it must be efficiently regenerated to sustain envelope assembly. Yet the enzymes that release polysaccharide cargo from the lipid carrier have remained elusive, leaving a major gap in our understanding of bacterial envelope biogenesis. Here we identify three periplasmic enzymes in Escherichia coli - UprA, UprB, and UprC - that hydrolyze the bond linking polysaccharide precursors to undecaprenyl. UprA releases elongating peptidoglycan glycan strands from the inner membrane, catalyzing the long-sought terminal step of peptidoglycan synthesis. UprB cleaves the undecaprenyl-linked initiating unit common to the enterobacterial common antigen and O-antigen pathways, while UprC is the likely release factor for the colanic acid pathway. These enzymes define a widespread Upr-like protein family (formerly DUF218) as polysaccharide release factors, revealing a unifying mechanism by which bacteria coordinate envelope assembly. Because all three Upr-like proteins share a conserved catalytic core, this family represents a potential target for antibiotics that simultaneously disrupt multiple envelope pathways.

microbiology↗

bioRxiv2026

Stress-induced Membrane Insertion at the β-Barrel Assembly Machinery Complex Regulates BepA Metalloprotease Activity

Proteases must be tightly regulated to prevent uncontrolled degradation, yet the mechanisms ensuring such control remain poorly understood. Members of the widespread M48 metalloprotease family are kept inactive by an autoinhibitory plug that blocks catalytic water activation, but how this plug is released was unknown. Here, using genetic, biochemical and cryo-EM approaches, we discover the activation mechanism of BepA, a quality-control protease that preserves outer membrane integrity by surveilling the {beta}-barrel assembly machinery (BAM) in Gram-negative bacteria. Our cryo-EM analysis of BepA engaged with a stalled BAM-substrate assembly complex revealed that a flexible, unstructured 6-lid covering the active site in the latent protein functions as a molecular harpoon, inserting into the outer membrane when a substrate stalls at BAM and thereby docking BepA at the complex. This membrane anchoring promotes displacement of the autoinhibitory plug and unlocks protease activity precisely where and when it is needed. Thus, a dual enzyme activation mechanism is coupled to membrane association under stress, ensuring that BepA remains inactive until properly localized. Our findings reveal how membranes themselves can license protease activation, a principle that may extend beyond M48 metalloproteases.

microbiology↗

bioRxiv2026

Extreme envelope plasticity drives temperature-dependent morphogenesis in the LPS-free bacterium Sphingobium yanoikuyae

Bacterial growth patterns are generally considered to be constrained and species-specific. Here, we show that Sphingobium yanoikuyae exhibits exceptional plasticity in envelope architecture and morphogenesis supported by a glycosphingolipid (GSL)-based outer membrane and an atypical peptidoglycan structure. At 30{degrees}C, cells expand asymmetrically via a rare bipolar envelope synthesis mode, whereas growth at 37{degrees}C triggers a transition toward longitudinal elongation accompanied by increased outer membrane vesiculation, indicating a reversible reprogramming of morphogenesis under host-relevant temperature. This switch is supported by rapid outer membrane dynamics, including high membrane fluidity and fast redistribution of envelope components. In addition to these pronounced morphological changes, peptidoglycan contains unusually short glycan strands and displays specific temperature-dependent remodeling suggesting that growth plasticity is not only governed by spatial regulation but also by changes in cell wall cross-linking pattern. Genetic analyses further identify essential roles for proteins involved in outer membrane-peptidoglycan and outer membrane-inner membrane coupling, as well as GSL transport systems, and core cell wall synthesis machinery, while revealing extensive redundancy in envelope remodeling enzymes. Together, these results establish S. yanoikuyae as a model for extreme envelope adaptability, where a highly fluid outer membrane and structurally unconventional peptidoglycan enable reversible transitions between distinct growth programs, potentially shaping environmental fitness and host-associated interactions.

microbiology↗

bioRxiv2021

Role of endopeptidases in peptidoglycan synthesis mediated by alternative cross-linking enzymes in Escherichia coli

Bacteria resist to the turgor pressure of the cytoplasm through a net-like macromolecule, the peptidoglycan, made of glycan strands connected via peptides cross-linked by penicillin-binding proteins (PBPs). We recently reported the emergence of {beta}-lactam resistance resulting from a bypass of PBPs by the YcbB L,D-transpeptidase (LdtD), which form chemically distinct 3[->]3 cross-links compared to 4[->]3 formed by PBPs. Here we show that peptidoglycan expansion requires controlled hydrolysis of cross-links and identify amongst eight endopeptidase paralogues the minimum enzyme complements essential for bacterial growth with 4[->]3 (MepM) and 3[->]3 (MepM and MepK) cross-links. Purified Mep endopeptidases unexpectedly displayed a 4[->]3 and 3[->]3 dual specificity implying recognition of a common motif in the two cross-link types. Uncoupling of the polymerization of glycan chains from the 4[->]3 cross-linking reaction was found to facilitate the bypass of PBPs by YcbB. These results illustrate the plasticity of the peptidoglycan polymerization machinery in response to the selective pressure of {beta}-lactams.

microbiology↗