Regulation of the activity of a tetra-modular lytic polysaccharide monooxygenase from Enterobacter cloacae by inter-domain copper coordination
Abstract
Enterobacter cloacae is a Gram-negative nosocomial human pathogen that inhabits diverse ecological niches. Its genome encodes a conserved set of putative chitin-active enzymes, including a lytic polysaccharide monooxygenase (LPMO), termed EcLPMO, which we functionally characterized in this study. EcLPMO is a tetra-modular protein consisting of an auxiliary activity family 10 (AA10) catalytic domain, two central domains of unknown function (DUF-A and DUF-B), and a C-terminal carbohydrate-binding module (CBM73). Functional assays using full-length EcLPMO and its truncated variants demonstrated that the AA10 domain oxidatively cleaves chitin at the C1 position. The CBM73 module enhances chitin binding and promotes synergy with endogenous chitinases. EcLPMO exhibited synergy with the unimodular chitinase EcChiA, resulting in up to 14-fold and 60-fold increase in GlcNAc release from alpha- and beta-chitin, respectively. Deletion of both DUFs reduced EcLPMO activity. While DUF-A alone and the association of DUF-A and DUF-B showed limited chitin binding, DUF-B alone exhibited no binding, suggesting a distinct role. Using AlphaFold3, we observed that the DUF-B domain contains two highly conserved histidines that coordinate the AA10-bound copper, forming a previously unreported 'inter-domain tetra-histidine copper coordination' center. These findings highlight the structural and functional complexity of EcLPMO and suggest that its accessory domains, particularly DUF-B, may contribute to enzyme stability and substrate interaction. We speculate that DUF-B may protect the LPMO active site from oxidative damage, a feature that could prove crucial in its ecological and pathogenic contexts.
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BibTeX
@article{bardhan2026regulation,
title = {{Regulation of the activity of a tetra-modular lytic polysaccharide monooxygenase from Enterobacter cloacae by inter-domain copper coordination}},
author = {Koyel Bardhan and Saumashish Mukherjee and Theruvothu Madathil Vandhana and Alessia Munzone and Andres Posbeyikian and Sacha Grisel and Lal Duhsaki and Jean-Guy Berrin and Bastien Bissaro and Jogi Madhuprakash},
journal = {Journal of Biological Chemistry},
year = {2026},
month = sep,
doi = {10.1016/j.jbc.2025.110964},
url = {https://hal.inrae.fr/hal-05440314v1},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Regulation of the activity of a tetra-modular lytic polysaccharide monooxygenase from Enterobacter cloacae by inter-domain copper coordination
AU - Koyel Bardhan
AU - Saumashish Mukherjee
AU - Theruvothu Madathil Vandhana
AU - Alessia Munzone
AU - Andres Posbeyikian
AU - Sacha Grisel
AU - Lal Duhsaki
AU - Jean-Guy Berrin
AU - Bastien Bissaro
AU - Jogi Madhuprakash
PY - 2026
DA - 2026/09/08
JO - Journal of Biological Chemistry
DO - 10.1016/j.jbc.2025.110964
UR - https://hal.inrae.fr/hal-05440314v1
AB - Enterobacter cloacae is a Gram-negative nosocomial human pathogen that inhabits diverse ecological niches. Its genome encodes a conserved set of putative chitin-active enzymes, including a lytic polysaccharide monooxygenase (LPMO), termed EcLPMO, which we functionally characterized in this study. EcLPMO is a tetra-modular protein consisting of an auxiliary activity family 10 (AA10) catalytic domain, two central domains of unknown function (DUF-A and DUF-B), and a C-terminal carbohydrate-binding module (CBM73). Functional assays using full-length EcLPMO and its truncated variants demonstrated that the AA10 domain oxidatively cleaves chitin at the C1 position. The CBM73 module enhances chitin binding and promotes synergy with endogenous chitinases. EcLPMO exhibited synergy with the unimodular chitinase EcChiA, resulting in up to 14-fold and 60-fold increase in GlcNAc release from alpha- and beta-chitin, respectively. Deletion of both DUFs reduced EcLPMO activity. While DUF-A alone and the association of DUF-A and DUF-B showed limited chitin binding, DUF-B alone exhibited no binding, suggesting a distinct role. Using AlphaFold3, we observed that the DUF-B domain contains two highly conserved histidines that coordinate the AA10-bound copper, forming a previously unreported 'inter-domain tetra-histidine copper coordination' center. These findings highlight the structural and functional complexity of EcLPMO and suggest that its accessory domains, particularly DUF-B, may contribute to enzyme stability and substrate interaction. We speculate that DUF-B may protect the LPMO active site from oxidative damage, a feature that could prove crucial in its ecological and pathogenic contexts.
SN - 0021-9258
SN - 1083-351X
ER -