Download MendeleyThe role of conserved residues in Fdc decarboxylase in prenylated flavin mononucleotide oxidative maturation, cofactor isomerization, and catalysis.
Bailey, S.S., Payne, K.A.P., Fisher, K., Marshall, S.A., Cliff, M.J., Spiess, R., Parker, D.A., Rigby, S.E.J., Leys, D.(2018) J Biol Chem 293: 2272-2287
- PubMed: 29259125
- DOI: https://doi.org/10.1074/jbc.RA117.000881
- Primary Citation of Related Structures:
6EV3, 6EV4, 6EV5, 6EV6, 6EV7, 6EV8, 6EV9, 6EVA, 6EVB, 6EVC, 6EVD, 6EVE, 6EVF - PubMed Abstract:
The UbiD family of reversible decarboxylases act on aromatic, heteroaromatic, and unsaturated aliphatic acids and utilize a prenylated flavin mononucleotide (prFMN) as cofactor, bound adjacent to a conserved Glu-Arg-Glu/Asp ionic network in the enzyme's active site. It is proposed that UbiD activation requires oxidative maturation of the cofactor, for which two distinct isomers, prFMN ketimine and prFMN iminium , have been observed. It also has been suggested that only the prFMN iminium form is relevant to catalysis, which requires transient cycloaddition between substrate and cofactor. Using Aspergillus niger Fdc1 as a model system, we reveal that isomerization of prFMN iminium to prFMN ketimine is a light-dependent process that is largely independent of the Glu 277 -Arg 173 -Glu 282 network and accompanied by irreversible loss of activity. On the other hand, efficient catalysis was highly dependent on an intact Glu-Arg-Glu network, as only Glu ¡ú Asp substitutions retain activity. Surprisingly, oxidative maturation to form the prFMN iminium species is severely affected only for the R173A variant. In summary, the unusual irreversible isomerization of prFMN is light-dependent and probably proceeds via high-energy intermediates but is independent of the Glu-Arg-Glu network. Our results from mutagenesis, crystallographic, spectroscopic, and kinetic experiments indicate a clear role for the Glu-Arg-Glu network in both catalysis and oxidative maturation.
Organizational Affiliation:
From the Manchester Institute of Biotechnology, University of Manchester, 131 Princess Street, M1 7DN, United Kingdom and.
