Crystallographic snapshots of oxalyl-CoA decarboxylase give insights into catalysis by nonoxidative ThDP-dependent decarboxylases.
Despite more than five decades of extensive studies of thiamin diphosphate (ThDP) enzymes, there remain many uncertainties as to how these enzymes achieve their rate enhancements. Here, we present a clear picture of catalysis for the simple nonoxidative decarboxylase, oxalyl-coenzyme A (CoA) decarboxylase, based on crystallographic snapshots along the catalytic cycle and kinetic data on active site mutants. First, we provide crystallographic evidence that, upon binding of oxalyl-CoA, the C-terminal 13 residues fold over the substrate, aligning the substrate alpha-carbon for attack by the ThDP-C2 atom. The second structure presented shows a covalent reaction intermediate after decarboxylation, interpreted as being nonplanar. Finally, the structure of a product complex is presented. In accordance with mutagenesis data, no side chains of the enzyme are implied to directly participate in proton transfer except the glutamic acid (Glu-56), which promotes formation of the 1',4'-iminopyrimidine tautomer of ThDP needed for activation.
Berthold
Catrine L
CL
Department of Medical Biochemistry and Biophysics, Molecular Structural Biology, Karolinska Institutet, S-17177 Stockholm, Sweden.
Toyota
Cory G
CG
Moussatche
Patricia
P
Wood
Martin D
MD
Leeper
Finian
F
Richards
Nigel G J
NG
Lindqvist
Ylva
Y
D000214:Acyl Coenzyme A / Q000737:chemistry; D001665:Binding Sites; D002262:Carboxy-Lyases / Q000737:chemistry* / Q000235:genetics; D002384:Catalysis; D003065:Coenzyme A / Q000737:chemistry; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D009154:Mutation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry; D013379:Substrate Specificity; D013835:Thiamine Pyrophosphate / Q000737:chemistry*
D000214:Acyl Coenzyme A; D011994:Recombinant Proteins; C523110:oxalyl-coenzyme A; C106238:formyl-coenzyme A; D002262:Carboxy-Lyases; C059418:oxalyl CoA decarboxylase; D013835:Thiamine Pyrophosphate; D003065:Coenzyme A