CARBAMOYL-PHOSPHATE SYNTHETASE - A CROOKED PATH FROM SUBSTRATES TO PRODUCTS

The formation of carbamoyl phosphate is catalyzed by a single enzyme u sing glutamine, bicarbonate and two molecules of ATP via a reaction me chanism that requires a minimum of four consecutive reactions and thre e unstable intermediates. The recently determined X-ray crystal struct ure of carbamoyl phosphate synthetase has revealed the location of thr ee separate active sites connected by two molecular tunnels that run t hrough the interior of the protein. It has been demonstrated that the amidotransferase domain within the small subunit of the enzyme from Es cherichia coli hydrolyzes glutamine to ammonia via a thioester interme diate with Cys269. The ammonia migrates through the interior of the pr otein, where it reacts with carboxy phosphate to produce the carbamate intermediate. The carboxy phosphate intermediate is formed by the pho sphorylation of bicarbonate by ATP at a site contained within the amin o-terminal half of the large subunit. The carbamate intermediate is tr ansported through the interior of the protein to a second site within the carboxy-terminal half of the large subunit, where it is phosphoryl ated by another ATP to yield the final product, carbamoyl phosphate. T he entire journey from substrate to product covers a distance of nearl y 100 Angstrom.