The crystal structure of the formiminotransferase domain of formiminotransferase-cyclodeaminase: implications for substrate channeling in a bifunctional enzyme

Background:Background: The bifunctional enzyme formiminotransferase-cyclode aminase (FTCD) contains two active sites at different positions on the prot ein structure. The enzyme binds a gamma-linked polyglutamylated form of the tetrahydrofolate substrate and channels the product of the transferase rea ction from the transferase active site to the cyclodeaminase active site. S tructural studies of this bifunctional enzyme and its monofunctional domain s will provide insight into the mechanism of substrate channeling and the t wo catalytic reactions. Results: The crystal structure of the formiminotransferase (FT) domain of F TCD has been determined in the presence of a product analog, folinic acid. The overall structure shows that the FT domain comprises two subdomains tha t adopt a novel alpha/beta fold. Inspection of the folinic acid binding sit e reveals an electrostatic tunnel traversing the width of the molecule. The distribution of charged residues in the tunnel provides insight into the p ossible mode of substrate binding and channeling. The electron density reve als that the non-natural stereoisomer, (6R)-folinic acid, binds to the prot ein; this observation suggests a mechanism for product release. In addition , a single molecule of glycerol is bound to the enzyme and indicates a puta tive binding site for formiminoglutamate. Conclusions: The structure of the FT domain in the presence of folinic acid reveals a possible novel mechanism for substrate channeling. The position of the folinic acid and a bound glycerol molecule near to the sidechain of His82 suggests that this residue may act as the catalytic base required for the formiminotransferase mechanism.