TY - JOUR
T1 - Characterization of kinetics of DNA strand-exchange and ATP hydrolysis activities of recombinant PfRad51, a Plasmodium falciparum recombinase
AU - Bhattacharyya, Mrinal Kanti
AU - Bhattacharyya Nee Deb, Sunanda
AU - Jayabalasingham, Bamini
AU - Kumar, Nirbhay
N1 - Funding Information:
These studies were supported by grants from the NIH AI 46760 and Johns Hopkins Malaria Research Institute. The authors thank Dr. F.R. Bryant for valuable discussions and critical comments on the manuscript. We also thank Dr. Dario Calumo and Dr. Akhilesh Pandey for mass spectrometric analysis of recombinant PfRad51 protein.
PY - 2005/1
Y1 - 2005/1
N2 - Although homologous recombination-mediated DNA rearrangements are quite widespread in Plasmodium falciparum, the molecular mechanisms involved are essentially unknown. Recent identification of PfRad51 in P. falciparum has suggested that it may play central role during homologous recombination and DNA rearrangements. Full-length recombinant PfRad51 was over expressed in Escherichia coli and purified to near homogeneity. Using optimized enzymatic activity conditions recombinant PfRad51 protein was shown to catalyze DNA strand-exchange reaction, a central step during homologous recombination. Unlike bacterial RecA protein, PfRad51 promoted strand-exchange reaction does not require ATP hydrolysis. The PfRad51 protein also catalyzed ssDNA-dependent ATP hydrolysis and the k cat values were similar to those reported for human Rad51. The demonstration of strand-exchange activity of PfRad51 protein, first such report in any protozoan parasite, suggests importance of similar recombination mechanism during DNA rearrangements associated with antigenic variation in P. falciparum.
AB - Although homologous recombination-mediated DNA rearrangements are quite widespread in Plasmodium falciparum, the molecular mechanisms involved are essentially unknown. Recent identification of PfRad51 in P. falciparum has suggested that it may play central role during homologous recombination and DNA rearrangements. Full-length recombinant PfRad51 was over expressed in Escherichia coli and purified to near homogeneity. Using optimized enzymatic activity conditions recombinant PfRad51 protein was shown to catalyze DNA strand-exchange reaction, a central step during homologous recombination. Unlike bacterial RecA protein, PfRad51 promoted strand-exchange reaction does not require ATP hydrolysis. The PfRad51 protein also catalyzed ssDNA-dependent ATP hydrolysis and the k cat values were similar to those reported for human Rad51. The demonstration of strand-exchange activity of PfRad51 protein, first such report in any protozoan parasite, suggests importance of similar recombination mechanism during DNA rearrangements associated with antigenic variation in P. falciparum.
KW - ATPase assay
KW - Homologous recombination
KW - Malaria
KW - RecA
KW - Three strand-exchange assay
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U2 - 10.1016/j.molbiopara.2004.09.007
DO - 10.1016/j.molbiopara.2004.09.007
M3 - Article
C2 - 15610817
AN - SCOPUS:11144321643
VL - 139
SP - 33
EP - 39
JO - Molecular and Biochemical Parasitology
JF - Molecular and Biochemical Parasitology
SN - 0166-6851
IS - 1
ER -