Kinetic mechanism, ammonia channeling and domain cross-talk in Plasmodium falciparum GMP synthetase : an amido- transferase

Abstract

Glutamine amidotransferases (Gln-ATs) or transamidases, are the enzymes catalyzing amination of a wide variety of metabolites, like amino acids, purines, pyrimidines, amino sugars, coenzymes and antibiotics, that sometimes serve as entry points for nitrogen into the respective pathways Most of the amidotransferases reported till date can either directly utilize ammonia from an external source or generate it by the hydrolysis of glutamine, needed for the amination of the acceptor molecules. Generally, these enzymes are modular with spatially separated glutamianse and the acceptor domains, specific for catalyzing the complex amidotransferase reaction in two parts, the glutamine hydrolysis and the acceptor amination, respectively. A set of glutamine amidotransferases after glutamine hydrolysis, transfer the generated ammonia directly to the acceptor substrates, with glutamate synthase serving as an example. However, in other cases, the acceptor substrate is first pre-activated by utilization of a molecule of ATP that leads to the formation of catalytically active intermediate possessing an electron deficient centre. The intermediate finally reacts with ammonia (with lone pair of electrons) in a nucleophillic reaction to proceed to the formation of aminated product (Buchanan, 1973; Massiere and Badet-Denisot, 1998; Zalkin, 1985; Zalkin, 1993; Zalkin and Smith, 1998). Though the final nitrogen-incorporated products differ across amidotransferases, glutamate is the common product whenever glutamine is hydrolyzed. The amidotransferase reaction differs from that catalyzed by aminotransferases or transaminases (E.C. 2.6.1.) as the latter catalyse the transfer of α-amino group from an amino acid to an α-keto acid (generally α-keto glutaric acid), in a pyridoxal phosphate (PLP) dependent reaction (Hirotsu et al., 2005). A general amidotransferase reaction scheme (Scheme 1.1) is shown below: