Abstract
The flavin-dependent sugar oxidoreductase pyranose dehydrogenase (PDH) from the plant litter-degrading fungus Agaricus meleagris oxidizes d-glucose (GLC) efficiently at positions C2 and C3. The closely related pyranose 2-oxidase (P2O) from Trametes multicolor oxidizes GLC only at position C2. Consequently, the electron output per molecule GLC is twofold for PDH compared to P2O making it a promising catalyst for bioelectrochemistry or for introducing novel carbonyl functionalities into sugars. The aim of this study was to rationalize the mechanism of GLC dioxidation employing molecular dynamics simulations of GLC–PDH interactions. Shape complementarity through nonpolar van der Waals interactions was identified as the main driving force for GLC binding. Together with a very diverse hydrogen-bonding pattern, this has the potential to explain the experimentally observed promiscuity of PDH towards different sugars. Based on geometrical analysis, we propose a similar reaction mechanism as in P2O involving a general base proton abstraction, stabilization of the transition state, an alkoxide intermediate, through interaction with a protonated catalytic histidine followed by a hydride transfer to the flavin N5 atom. Our data suggest that the presence of the two potential catalytic bases His-512 and His-556 increases the versatility of the enzyme, by employing the most suitably oriented base depending on the substrate and its orientation in the active site. Our findings corroborate and rationalize the experimentally observed dioxidation of GLC by PDH and its promiscuity towards different sugars.Electronic supplementary materialThe online version of this article (doi:10.1007/s10822-013-9645-7) contains supplementary material, which is available to authorized users.
Highlights
The sugar oxidoreductase pyranose dehydrogenase (PDH, EC 1.1.99.29) is a monomeric flavoprotein of approximately 65 kDa containing *7 % carbohydrates
We propose a similar reaction mechanism as in pyranose 2-oxidase (P2O) involving a general base proton abstraction, stabilization of the transition state, an alkoxide intermediate, through interaction with a protonated catalytic histidine followed by a hydride transfer to the flavin N5 atom
Phosphate ion PO4910, most likely a crystallization-buffer artifact surrounded by amino acid residues Arg-87 to Asp-90 and Pro-406 to b, c After superpositioning, D-glucose coordinates were grafted from the P2O structure with PDB accession codes (b) 3PL8 or (c) 2IGO into PDH
Summary
The sugar oxidoreductase pyranose dehydrogenase (PDH, EC 1.1.99.29) is a monomeric flavoprotein of approximately 65 kDa containing *7 % carbohydrates. Depending on the sugar-substrate, it can perform (di)oxidations at C1, C2, C3, or C4 positions of the hexapyranose ring [5, 7]. This property, depends strongly on the source of the enzyme as well as on the sugar substrate. The electron output per molecule of GLC is twofold for Agaricus PDH compared to P2O. This makes PDH a promising catalyst for applications in bioelectrochemistry or for introducing novel carbonyl functionalities into sugars [5]
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