Interactions of helices on interdomain interface couple the chemistry of intermediate formation with allostery in Plasmodium falciparum GMP synthetase

GMP synthetase, a member of the glutamine amidotransferase family of enzymes catalyses the substitution of the C2 oxo-group of the purine base in XMP with an amino-group generating GMP, the last step in the biosynthesis of GMP. This seemingly simple reaction involves a complex series of catalytic events that include hydrolysis of the side chain of Gln generating ammonia in the glutamine amidotransferase domain (GATase), activation of XMP through generation of adenyl-XMP intermediate in the ATP pyrophosphatase domain (ATPPase) and tunnelling of ammonia from the GATase to the ATPPase domain where it reacts with the intermediate generating GMP. The catalytic events across the two domains are highly coordinated, resulting in an enzymatic machinery that functions without wastage of either ATP or Gln. Here, we have taken recourse to the analysis of structures and sequences of GMP synthetases, site-directed mutagenesis and enzymatic assays on the Plasmodium falciparum enzyme to decipher the molecular basis of domain coordination. Our studies show that residues on three helices at the domain interface are crucial for interdomain crosstalk. Further, we show that residues on these helices along with those on a loop referred to as lid-loop, play essential roles in catalysing adenyl-XMP formation. By way of intricately connecting residues involved in domain crosstalk with catalysis in ATPPase domain, the two domains in P. falciparum GMP synthetase synchronize their catalytic events. These results add significantly to our understanding of the working of GMP synthetases, which are drug targets in many infectious pathogens.