Neutron and Atomic Resolution X-ray Structures of a Lytic Polysaccharide Monooxygenase Reveal Copper-Mediated Dioxygen Binding and Evidence for N-Terminal Deprotonation

Abstract

A 1.1 Å resolution, room-temperature X-ray structure and a 2.1 Å resolution neutron structure of a chitin-degrading lytic polysaccharide monooxygenase domain from the bacterium Jonesia denitrificans (JdLPMO10A) show a putative dioxygen species equatorially bound to the active site copper. Both structures show an elongated density for the dioxygen, most consistent with a Cu(II)-bound peroxide. The coordination environment is consistent with Cu(II). In the neutron and X-ray structures, difference maps reveal the N-terminal amino group, involved in copper coordination, is present as a mixed ND and ND, suggesting a role for the copper ion in shifting the pK of the amino terminus.

Targeting the reactive intermediate in polysaccharide monooxygenases

“…For instance, it participates in a hydrogen bonding network with the substrate [20] which increases the basicity and provides a channel for deprotonation of the N terminus. It has also been suggested to be deprotonated from studies on model complexes [30], and lately, indications have also come from neutron diffraction and X-ray crystallography [54]. Therefore, we also investigated the BDEs for reactions involving deprotonated complexes (i.e., involving –NH).…”

Formally Copper(III)–Alkylperoxo Complexes as Models of Possible Intermediates in Monooxygenase Enzymes

“…Such deprotonation of the amino group has been suggested on the basis of X-ray crystallography. 22 The enhanced basicity of 6 would facilitate protonation to yield a [CuOOH] 2+ species (7) analogous to the species we have prepared and similarly stabilized by an anionic supporting ligand (for 7, a trans amide donor). Whether 7 is suffciently reactive to be a viable oxidizing intermediate is an open question.…”

Bracing copper for the catalytic oxidation of C–H bonds

“…Accordingly, deprotonation of the NH2 in LPMOs to give an imido group gains credence, and such a possibility was raised in the 2011 paper which announced the copper histidine brace active site in LPMOs. 1 The notion gained further momentum from neutron diffraction experiments performed on AA10 LPMOs, which appeared to show a lack of density around the amino terminus nitrogen atom 37 and a notable average Cu NH2 C of ~117°; confirmation by spectroscopy is nonetheless needed before a definitive assignment can be made. If true, however, the enhanced electron donation by the deprotonated moiety would raise the d(x 2 y 2 ) orbital high enough in energy such that a formal Cu(III) species becomes realistic.…”