Kinetic and biochemical characterization of Plasmodium falciparum GMP synthetase

Bhat, Javaid Yousuf ; Shastri, Brahmanaspati Ganapathi ; Balaram, Hemalatha (2008) Kinetic and biochemical characterization of Plasmodium falciparum GMP synthetase Biochemical Journal, 409 (1). pp. 263-273. ISSN 0264-6021

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Official URL: http://www.biochemj.org/bj/409/bj4090263.htm

Related URL: http://dx.doi.org/10.1042/BJ20070996

Abstract

Plasmodium falciparum, the causative agent of the fatal form of malaria, synthesizes GMP primarily from IMP and, hence, needs active GMPS (GMP synthetase) for its survival. GMPS, a G-type amidotransferase, catalyses the amination of XMP to GMP with the reaction occurring in two domains, the GAT (glutamine amidotransferase) and ATPPase (ATP pyrophosphatase). The GAT domain hydrolyses glutamine to glutamate and ammonia, while the ATPPase domain catalyses the formation of the intermediate AMP-XMP from ATP and XMP. Co-ordination of activity across the two domains, achieved through channelling of ammonia from GAT to the effector domain, is the hallmark of amidotransferases. Our studies aimed at understanding the kinetic mechanism of PfGMPS (Plasmodium falciparum GMPS) indicated steady-state ordered binding of ATP followed by XMP to the ATPPase domain with glutamine binding in a random manner to the GAT domain. We attribute the irreversible, Ping Pong step seen in initial velocity kinetics to the release of glutamate before the attack of the adenyl-XMP intermediate by ammonia. Specific aspects of the overall kinetic mechanism of PfGMPS are different from that reported for the human and Escherichia coli enzymes. Unlike human GMPS, absence of tight co-ordination of activity across the two domains was evident in the parasite enzyme. Variations seen in the inhibition by nucleosides and nucleotide analogues between human GMPS and PfGMPS highlighted differences in ligand specificity that could serve as a basis for the design of specific inhibitors. The present study represents the first report on recombinant His-tagged GMPS from parasitic protozoa.

Item Type:Article
Source:Copyright of this article belongs to Portland Press Limited.
Keywords:Amidotransferase; Atp Pyrophosphatase (ATPPase) Domain; Glutaminase Domain; Plasmodium Falciparum Gmp Synthetase; Interdomain Cross-talk; Irreversible Inactivation; Two-site Ping Pong Mechanism
ID Code:1259
Deposited On:04 Oct 2010 07:59
Last Modified:08 Jan 2011 03:16

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