Structural insights into the avian AICAR transformylase mechanism.
ATIC encompasses both AICAR transformylase and IMP cyclohydrolase activities that are responsible for the catalysis of the penultimate and final steps of the purine de novo synthesis pathway. The formyl transfer reaction catalyzed by the AICAR Tfase domain is substantially more demanding than that catalyzed by the other folate-dependent enzyme of the purine biosynthesis pathway, GAR transformylase. Identification of the AICAR Tfase active site and key catalytic residues is essential to elucidate how the non-nucleophilic AICAR amino group is activated for formyl transfer. Hence, the crystal structure of dimeric avian ATIC was determined as a complex with the AICAR Tfase substrate AICAR, as well as with an IMP cyclohydrolase inhibitor, XMP, to 1.93 A resolution. AICAR is bound at the dimer interface of the transformylase domains and forms an extensive hydrogen bonding network with a multitude of active site residues. The crystal structure suggests that the conformation of the 4-carboxamide of AICAR is poised to increase the nucleophilicity of the C5 amine, while proton abstraction occurs via His(268) concomitant with formyl transfer. Lys(267) is likely to be involved in the stabilization of the anionic formyl transfer transition state and in subsequent protonation of the THF leaving group.
Wolan
Dennis W
DW
Department of Molecular Biology and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Greasley
Samantha E
SE
Beardsley
G Peter
GP
Wilson
Ian A
IA
D000818:Animals; D001665:Binding Sites; D001717:Birds; D003198:Computer Simulation; D018360:Crystallography, X-Ray; D019281:Dimerization; D004789:Enzyme Activation; D019877:Hydroxymethyl and Formyl Transferases / Q000737:chemistry*; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry*; D009709:Nucleotide Deaminases / Q000737:chemistry*; D051027:Phosphoribosylaminoimidazolecarboxamide Formyltransferase; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D009097:Multienzyme Complexes; C113120:inosine monophosphate synthase; D019877:Hydroxymethyl and Formyl Transferases; D051027:Phosphoribosylaminoimidazolecarboxamide Formyltransferase; D009709:Nucleotide Deaminases; C021436:IMP cyclohydrolase