Learning from oligosaccharide soaks of crystals of an AA13 lytic polysaccharide monooxygenase: crystal packing, ligand binding and active-site disorder

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• 作者：Kristian E. H. Frandsen ; Jens-Christian Navarro Poulsen ; Morten Tovborg ; Katja S. Johansen and Leila Lo Leggio
• 关键词：lytic polysaccharide monooxygenases ; LPMOs ; starch degradation ; active-site stability ; biomass utilization
• 刊名：Acta Crystallographica Section D
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：73
• 期：1
• 页码：64-76
• 全文大小：1988K
• ISSN：1399-0047

文摘

Lytic polysaccharide monooxygenases (LPMOs) are a class of copper-dependent enzymes discovered within the last ten years. They oxidatively cleave polysaccharides (chitin, lignocellulose, hemicellulose and starch-derived), presumably making recalcitrant substrates accessible to glycoside hydrolases. Recently, the first crystal structure of an LPMO–substrate complex was reported, giving insights into the interaction of LPMOs with β-linked substrates (Frandsen et al., 2016). The LPMOs acting on α-linked glycosidic bonds (family AA13) display binding surfaces that are quite different from those of LPMOs that act on β-linked glycosidic bonds (families AA9–AA11), as revealed from the first determined structure (Lo Leggio et al., 2015), and thus presumably the AA13s interact with their substrate in a distinct fashion. Here, several new structures of the same AA13 enzyme, Aspergillus oryzae AA13, are presented. Crystals obtained in the presence of high zinc-ion concentrations were used, as they can be obtained more reproducibly than those used to refine the deposited copper-containing structure. One structure with an ordered zinc-bound active site was solved at 1.65 Å resolution, and three structures from crystals soaked with maltooligosaccharides in solutions devoid of zinc ions were solved at resolutions of up to 1.10 Å. Despite similar unit-cell parameters, small rearrangements in the crystal packing occur when the crystals are depleted of zinc ions, resulting in a more occluded substrate-binding surface. In two of the three structures maltooligosaccharide ligands are bound, but not at the active site. Two of the structures presented show a His-ligand conformation that is incompatible with metal-ion binding. In one of these structures this conformation is the principal one (80% occupancy), giving a rare atomic resolution view of a substantially misfolded enzyme that is presumably rendered inactive.