An Overview of Plasmodium Protein Kinases

doi:10.1016/S0169-4758(99)01527-6

Parasitology Today

Volume 15, Issue 11, 1 November 1999, Pages 449-454

Parasitology Today

https://doi.org/10.1016/S0169-4758(99)01527-6 Get rights and content

Abstract

Protein kinases are key regulators of many biochemical processes in eukaryotic cells. Malaria parasites, in spite of all their peculiarities, are not likely to represent an exception in this respect. Over the past few years, several genes encoding Plasmodium protein kinases have been cloned and characterized; these molecular studies extend previous data on kinase activities in parasite extracts. Here, Barbara Kappes, Christian Doerig and Ralph Graeser present available data on this topic, with an emphasis on cloned protein kinase genes, and discuss the potential outcome of such research in the context of drug development.

Section snippets

CMGC group

The CMGC group includes the family of cyclin- dependent protein kinases (CDKs), the mitogen-activated protein kinase (MAPK) family, the glycogen-synthase kinase 3 (GSK-3) family, the CDK-like kinase (CLK) family and other close relatives. Most of these enzymes are proline-directed kinases, phosphorylating substrates at sites close to proline residues¹.

CaMK group

The Ca²⁺–calmodulin kinase (CaMK) group includes the family of protein kinases regulated by Ca²⁺–calmodulin, the calcium-dependent protein kinases (CDPKs), the SNF1 (sucrose non-fermenting)/AMP-activated kinase (AMPK) family and other close relatives. In general, the CaMK group of protein kinases are basic amino acid-directed enzymes, phosphorylating substrates at Ser/Thr sites close to Arg and Lys residues¹.

AGC group

The AGC group includes the cyclic nucleotide-dependent families of PKA and PKG, the PKC family, the β-adrenergic receptor kinase family, the ribosomal S6 kinase family and other close relatives. Like the kinases of the CaMK group, AGC protein kinases tend to be basic amino acid-directed¹.

Casein kinase 1 (CK1) family

Like CK2, CK1 has acidic recognition sequences, and is often involved in hierarchical protein phosphorylation reactions, acting on a wide range of substrates²⁷.

A malaria CK1 homologue has been isolated and characterized (Ref. 49). PfCK1 is very similar to the human α-isoform in that it lacks a C-terminal extension sequence, which in some CK1 isoforms mediates subcellular localization or is the target of inhibitory autophosphorylation50, 51. The kinase activity of recombinant PfCK1 can be

Conventional protein tyrosine kinase group

To date, no convincing evidence has emerged for the existence of tyrosine kinases in P. falciparum. This apparent lack of tyrosine kinases, and the relatedness of the apicomplexans to the plant kingdom suggests that if a receptor protein kinase system is present in the parasite it might be of the type found in plants. In contrast to the animal kingdom, with receptor kinases autophosphorylating at tyrosine residues, the plant receptor kinases belong to the Ser/Thr kinase family⁵⁷. However, none

How far have we come?

Validation of a given kinase as a drug target requires the proof that it is essential for parasite growth and/or differentiation. Gene knockout might present an attractive means of demonstrating the function of a protein kinase in sexual differentiation; the mutant parasites would be unaffected in their ability to proliferate, but could no longer form sexual stages. A similar experiment for a kinase with an essential function during the asexual multiplication cycle is hampered by the fact that

Acknowledgements

We thank Debopam Chakrabarti, Chiang Syin, Gordon Langsley and Ginette Jaureguiberry for unpublished information, and Tim Hunt for critically reading the manuscript. Furthermore, we would like to acknowledge the Malaria Genome Project. Barbara Kappes was funded in part by funds from the Canton City of Basel, the UNDP/World Bank/WHO Special Programme for Research and Training in Tropical Diseases (TDR) projects 920559 and 960140, and the Swiss National Foundation Grant 31-39122.93. Christian

References (59)

A. Garcia
A family of PP2 phosphatases in Plasmodium falciparum and parasitic protozoa
Parasitol. Today
(1999)
B. Andrews et al.
The cyclin family of budding yeast: abundant use of a good idea
Trends Genet.
(1998)
R. Beyaert
Cleavage of PITSLRE kinases by ICE/CASP-1 and CPP32/CASP-3 during apoptosis induced by tumor necrosis factor
J. Biol. Chem.
(1997)
C.D. Doerig
Pfcrk-1, a developmentally regulated cdc2-related protein kinase of Plasmodium falciparum
Mol. Biochem. Parasitol.
(1995)
R. Graeser
Mechanisms of activation of the cdc2-related kinase PfPK5 from Plasmodium falciparum
Mol. Biochem. Parasitol.
(1996)
R. Graeser
Plasmodium falciparum protein kinase 5 and the malarial nuclear division cycles
Mol. Biochem. Parasitol.
(1996)
G.L. Johnson et al.
Sequential protein kinase reactions controlling cell growth and differentiation
Curr. Opin. Cell Biol.
(1994)
H.D. Madhani et al.
The riddle of MAP kinase signaling specificity
Trends Genet.
(1998)
C.M. Doerig
A MAP kinase homologue from the human malaria parasite, Plasmodium falciparum
Gene
(1996)
D.T. Lin
Stage-specific expression of a Plasmodium falciparum protein related to the eukaryotic mitogen-activated protein kinases
Mol. Biochem. Parasitol.
(1996)

N.J. Kilby

Site-specific recombinases: tools for genome engineering

Trends Genet.

(1993)

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Proteins, such as the raf kinase inhibitory protein (RKIP), serve as modulators of signalling pathways by either promoting or inhibiting the formation of productive signalling complexes through protein–protein interactions. In the present study, the plasmodial RKIP ortholog, PfPE-PB1, was cloned, recombinantly expressed and purified to homogeneity. The purified protein was used to investigate the effect of plasmodial RKIP on the autophosphorylation and substrate phosphorylation activity of Plasmodium falciparum calcium-dependent protein kinase 1, PfCDPK1. Phosphorylation of RKIP by PfCDPK1 in in vitro kinase assays suggests that RKIP may be an in vivo substrate of this kinase, although the specific activity of PfCDPK1 is approximately seven-fold lower when RKIP, instead of casein, an exogenous substrate of this enzyme, is used as a substrate. In addition to the observed phosphorylation of RKIP itself, its presence in the assays greatly enhanced the autophosphorylation capacity of PfCDPK1 by approximately 5.5-fold. This substantial increase in autophosphorylation activity was associated with a diminished substrate phosphorylation activity of PfCDPK1 when casein was used. At the same time, RKIP phosphorylation slightly increased when casein was included into the assays. Thus, RKIP is recognized as a substrate under in vitro conditions and appears to act as a regulator of PfCDPK1 kinase activity, which possibly is one of its actual functions in the parasite.
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ReviewAn Overview of Plasmodium Protein Kinases

Abstract

Section snippets

CMGC group

CaMK group

AGC group

Casein kinase 1 (CK1) family

Conventional protein tyrosine kinase group

How far have we come?

Acknowledgements

Parasitol. Today

Trends Genet.

J. Biol. Chem.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Curr. Opin. Cell Biol.

Trends Genet.

Gene

Mol. Biochem. Parasitol.

J. Biol. Chem.

FEBS Lett.

J. Biol. Chem.

Mol. Biochem. Parasitol.

Biochim. Biophys. Acta

J. Biol. Chem.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

J. Biol. Chem.

Exp. Parasitol.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Trends Biochem. Sci.

Biochim. Biophys. Acta

Mol. Biochem. Parasitol.

Trends Biochem. Sci.

Trends Genet.

Review
An Overview of Plasmodium Protein Kinases