Conserved copper(I)-binding auxiliary domains link virulence-associated LPMOs to copper homeostasis in the intestinal environment

Authors

Kommedal, E. G., Berggreen, H., Ronnekleiv, S. E., Vinther Sorensen, H., Runningen, A., Zhou, Y., Krengel, U., Rohr, A. K., Eijsink, V. G. H., Forsberg, Z.

Abstract

Multidomain lytic polysaccharide monooxygenases (LPMOs) are widely distributed in human pathogenic bacteria and increasingly recognized for promoting host interactions and virulence. Recent work has shown that immunizing mice with CbpD, a trimodular LPMO from Pseudomonas aeruginosa, provides protection against lethal P. aeruginosa infection. In several human intestinal pathogenic bacteria, LPMOs occur as tetra- and pentamodular proteins and their ability to interact with the host and facilitate virulence has primarily been ascribed to their ability to bind to N-acetylglucosamine containing glycans. Recently, AlphaFold predictions and substrate-binding studies of the Vibrio cholerae colonization factor GbpA suggested the presence of a copper-binding site in its non-catalytic third domain. In light of the recent demonstration that MUC2, the primary intestinal mucin, harbors two conserved and distinct copper binding sites for Cu(II) and Cu(I), we hypothesized that the auxiliary non-catalytic copper-binding domains of intestinal LPMOs evolved as a response to the intestinal environment. Here, we examine GbpA and homologous multidomain LPMOs from food-borne intestinal disease-causing Gram-positive bacteria from the genera Bacillus and Listeria. By combining mutagenesis with biochemical, spectroscopic, and computational approaches, we demonstrate that these multidomain LPMOs contain conserved, yet structurally distinct copper(I)-binding motifs on their non-catalytic third domain. A comprehensive review and reassessment of the literature spanning the past two decades reveals consistent links between the copper-binding ability of these LPMOs and host-pathogen interactions. Thus, our findings link these LPMOs to copper management in the intestine, offering an explanation for why their unique architecture is preserved across different bacterial lineages causing intestinal disease.

Preprint server: bioRxiv
The authors list and abstract were imported from bioRxiv on 07 Aug 2026.

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