Redox-dependent substrate-cofactor interactions in the Michaelis-complex of a flavin-dependent oxidoreductase

How an enzyme activates its substrate for turnover is fundamental for catalysis but incompletely understood on a structural level. With redox enzymes one typically analyses structures of enzyme-substrate complexes in the unreactive oxidation state of the cofactor, assuming that the interaction between enzyme and substrate is independent of the cofactors oxidation state. Here, we investigate the Michaelis complex of the flavoenzyme xenobiotic reductase A with the reactive reduced cofactor bound to its substrates by X-ray crystallography and resonance Raman spectroscopy and compare it to the non-reactive oxidized Michaelis complex mimics. We find that substrates bind in different orientations to the oxidized and reduced flavin, in both cases flattening its structure. But only authentic Michaelis complexes display an unexpected rich vibrational band pattern uncovering a strong donor-acceptor complex between reduced flavin and substrate. This interaction likely activates the catalytic ground state of the reduced flavin, accelerating the reaction within a compressed cofactor-substrate complex.

DDC Class

620: Ingenieurwissenschaften

Funding Organisations

Deutsche Forschungsgemeinschaft (DFG)

More Funding Information

The Deutsche Forschungsgemeinschaft is acknowledged for funding of the project (EXC 314 ‘Unifying Concepts in Catalysis—UniCat’)

Publication version

publishedVersion

Lizenz

https://creativecommons.org/licenses/by/4.0/

Name

ncomms16084.pdf

Size

909.57 KB

Format

Adobe PDF

Options

Redox-dependent substrate-cofactor interactions in the Michaelis-complex of a flavin-dependent oxidoreductase