The Plasmodium falciparum cGMP-dependent protein kinase (PfPKG) uniquely contains three cGMP binding sites, but also has a ‘degenerate’ fourth site. The role of each cGMP-binding site in PfPKG activation remains unknown. We have analysed the effect of mutation of each cGMP-binding site (individually and in combination) on PfPKG activation in vitro. The most striking result was that mutation of cGMP site 3 resulted in a 10–49-fold increase in the Ka(cGMP) value and a 45–55% decrease in maximal activity compared with wild-type. Mutations involving only cGMP-binding sites 1 and 2 had less effect on both the Ka(cGMP) values and the maximal activities. These results suggest that, although all three cGMP-binding sites are involved in PfPKG activation, cGMP-binding site 3 has the greatest influence on activation. A mutation in the fourth, degenerate cGMP-binding site decreased PfPKG maximal activity by 40%, but did not change the Ka(cGMP) value for the PfPKG mutant, suggesting that this site does not bind cGMP, but is required for full activation of PfPKG. The distinct activation properties of PfPKG from mammalian isoforms may be exploitable in the design of a parasite-specific inhibitor and development of a novel anti-malarial drug.
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September 2003
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Research Article|
September 01 2003
The role of two novel regulatory sites in the activation of the cGMP-dependent protein kinase from Plasmodium falciparum
Wensheng DENG;
Wensheng DENG
Department of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine, Keppel Street, London WC1E 7HT, U.K.
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Asha PARBHU-PATEL;
Asha PARBHU-PATEL
Department of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine, Keppel Street, London WC1E 7HT, U.K.
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David J. MEYER;
David J. MEYER
Department of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine, Keppel Street, London WC1E 7HT, U.K.
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David A. BAKER
David A. BAKER
1
Department of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine, Keppel Street, London WC1E 7HT, U.K.
1To whom correspondence should be addressed (e-mail david.baker@lshtm.ac.uk).
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Publisher: Portland Press Ltd
Received:
March 27 2003
Revision Received:
June 13 2003
Accepted:
June 20 2003
Accepted Manuscript online:
June 20 2003
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2003
2003
Biochem J (2003) 374 (2): 559–565.
Article history
Received:
March 27 2003
Revision Received:
June 13 2003
Accepted:
June 20 2003
Accepted Manuscript online:
June 20 2003
Citation
Wensheng DENG, Asha PARBHU-PATEL, David J. MEYER, David A. BAKER; The role of two novel regulatory sites in the activation of the cGMP-dependent protein kinase from Plasmodium falciparum. Biochem J 1 September 2003; 374 (2): 559–565. doi: https://doi.org/10.1042/bj20030474
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