Publication

Article Metrics

Citations


Online attention

The role of conserved residues in Fdc decarboxylase in prenylated flavin mononucleotide oxidative maturation, cofactor isomerisation and catalysis

DOI: 10.1074/jbc.RA117.000881 DOI Help

Authors: Samuel S. Bailey (The University of Manchester) , Karl A. P. Payne (Manchester Institute of Biotechnology) , Karl Fisher (The University of Manchester) , Stephen A. Marshall (The University of Manchester) , Matthew J. Cliff (The University of Manchester) , Reynard Spiess (The University of Manchester) , David A. Parker (Shell International Exploration and Production) , Stephen E. J. Rigby (The University of Manchester) , David Leys (The University of Manchester)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of Biological Chemistry

State: Published (Approved)
Published: December 2017
Diamond Proposal Number(s): 8997

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-ArgGlu/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, prFMNketimine and prFMNiminium have been observed. It also has been suggested that only the prFMNiminium form is relevant to catalysis, which requires transient cycloaddition between substrate and cofactor. Using Aspergillus niger Fdc1 as a model system, we reveal isomerization of prFMNiminium to prFMNketimine is a light-dependent process that is largely independent of the Glu277-Arg173-Glu282 network and accompanied by irreversible loss of activity. On the other hand, efficient catalysis was highly dependent on an intact Glu-Ar-Glu network, as only Glu to Asp substitutions retain activity. Surprisingly, oxidative maturation to form the prFMNiminium species is severely affected only for the R173A variant. In summary, the unusual irreversible isomerization of prFMN is light dependent and likely 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.

Journal Keywords: prFMN; UbiD; crystal structure; EPR; decarboxylase; enzyme catalysis; enzyme mechanism; oxidation-reduction (redox); flavin; radical

Subject Areas: Biology and Bio-materials, Chemistry


Instruments: I02-Macromolecular Crystallography , I03-Macromolecular Crystallography , I04-1-Macromolecular Crystallography (fixed wavelength) , I04-Macromolecular Crystallography