Purification and Characterization of LPMOs

Recombinant ScAA10C (UniProt Q9RJY2) from S. coelicolor was produced and purified as previously described (7 (link)). ScAA10C was copper saturated according to Loose et al (42 (link)), carefully desalted using a PD MidiTrap column [G-25, GE Healthcare (57 (link))], and stored in sodium phosphate (25 mM, pH 6.0). The LPMO from S. marcescens (SmAA10A; UniProt O83009) was produced and purified as previously described (58 (link)), copper-saturated in the same way as ScAA10C, and stored in the same buffer. Manganese-dependent SOD (MnSOD) from Escherichia coli (Sigma-Aldrich, S5639) was solubilized in Tris⋅HCl (10 mM, pH 8.0) and desalted (PD MidiTrap G-25, GE Healthcare) in the same buffer before use. HRP (type II) (Sigma-Aldrich, P8250) was solubilized in Milli-Q water and filtered (Filtropur S, 0.2 µm polyethersulfone (PES), Sarstedt). All enzymes were stored at 4 °C.

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Kommedal E.G., Sæther F., Hahn T, & Eijsink V.G. (2022). Natural photoredox catalysts promote light-driven lytic polysaccharide monooxygenase reactions and enzymatic turnover of biomass. Proceedings of the National Academy of Sciences of the United States of America, 119(34), e2204510119.

Publication 2022

PD MidiTrap column PD MidiTrap G-25 Filtropur S

Buffer Copper Enzymes Escherichia coli Manganese Polyethersulfone Sodium phosphate Tris

Corresponding Organization :

Other organizations : Norwegian University of Life Sciences, Fraunhofer Institute for Interfacial Engineering and Biotechnology

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Variable analysis

independent variables

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dependent variables

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control variables

Production and purification methods for recombinant ScAA10C, SmAA10A, and MnSOD enzymes
Copper saturation of ScAA10C and SmAA10A enzymes
Desalting and buffer conditions for ScAA10C, SmAA10A, and MnSOD enzymes
Storage conditions (4°C) for all enzymes

positive controls

None explicitly mentioned

negative controls

None explicitly mentioned

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