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Research article2017Peer reviewedOpen access

High-resolution structure of a lytic polysaccharide monooxygenase from Hypocrea jecorina reveals a predicted linker as an integral part of the catalytic domain

Hansson, Henrik; Karkehabadi, Saeid; Mikkelsen, Nils; Douglas, Nicholai R.; Kim, Steve; Lam, Anna; Kaper, Thijs; Kelemen, Brad; Meier, Katlyn K.; Jones, Stephen M.; Solomon, Edward I.; Sandgren, Mats

Abstract

For decades, the enzymes of the fungus Hypocrea jecorina have served as a model system for the breakdown of cellulose. Three-dimensional structures for almost all H. jecorina cellulose-degrading enzymes are available, except for HjLPMO9A, belonging to the AA9 family of lytic polysaccharide monooxygenases (LPMOs). These enzymes enhance the hydrolytic activity of cellulases and are essential for cost-efficient conversion of lignocellulosic biomass. Here, using structural and spectroscopic analyses, we found that native HjLPMO9A contains a catalytic domain and a family-1 carbohydrate-binding module (CBM1) connected via a linker sequence. A C terminally truncated variant of HjLPMO9A containing 21 residues of the predicted linker was expressed at levels sufficient for analysis. Here, using structural, spectroscopic, and biochemical analyses, we found that this truncated variant exhibited reduced binding to and activity on cellulose compared with the full-length enzyme. Importantly, a 0.95- resolution X-ray structure of truncated HjLPMO9A revealed that the linker forms an integral part of the catalytic domain structure, covering a hydrophobic patch on the catalytic AA9 module. We noted that the oxidized catalytic center contains a Cu(II) coordinated by two His ligands, one of which has a His-brace in which the His-1 terminal amine group also coordinates to a copper. The final equatorial position of the Cu(II) is occupied by a water-derived ligand. The spectroscopic characteristics of the truncated variant were not measurably different from those of full-length HjLPMO9A, indicating that the presence of the CBM1 module increases the affinity of HjLPMO9A for cellulose binding, but does not affect the active site.

Keywords

electron paramagnetic resonance (EPR); enzyme mechanism; enzyme structure; Hypocrea jecorina; X-ray crystallography; auxiliary activity family 9 (AA9); lytic polysaccharide monooxygenase (LPMO)

Published in

Journal of Biological Chemistry
2017, Volume: 292, number: 46, pages: 19099-19109 Publisher: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

      SLU Authors

    • Hansson, Henrik

      • The Department of Chemistry and Biotechnology, Swedish University of Agricultural Sciences
      • Karkehabadi, Saeid

        • The Department of Chemistry and Biotechnology, Swedish University of Agricultural Sciences
        • Mikkelsen, Nils

          • The Department of Chemistry and Biotechnology, Swedish University of Agricultural Sciences
          • Sandgren, Mats

            • The Department of Chemistry and Biotechnology, Swedish University of Agricultural Sciences

          UKÄ Subject classification

          Biochemistry and Molecular Biology

          Publication identifier

          DOI: https://doi.org/10.1074/jbc.M117.799767

          Permanent link to this page (URI)

          https://res.slu.se/id/publ/92626