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A unique 45-amino-acid region in the toprim domain of Plasmodium falciparum gyrase B is essential for its activity.

ABSTRACT: DNA gyrase is the only topoisomerase that can introduce negative supercoils into the DNA at the cost of ATP hydrolysis. Some but not all the steps of the topoisomerization reaction are understood clearly for both eukaryotic topoII and DNA gyrase. This study is an attempt to understand whether the B subunit of DNA gyrase binds to DNA directly, which may be central to the stimulation of its ATPase activity essential for gyrase function. We have dissected the Plasmodium falciparum gyrase B (PfGyrB) subunit to identify a 45-amino-acid region in the toprim domain that is responsible for its intrinsic DNA binding activity, DNA-stimulated ATPase activity, and DNA cleavage. We find that DNA has to enter through the ATP-operated clamp of PfGyrB to gain access to the DNA binding region. Furthermore, the rate of ATP hydrolysis of PfGyrB increases significantly with increasing DNA length, suggesting a possible communication between the ATPase domain and the DNA binding region that can account for its optimal ATPase activity. These results not only highlight the mechanism of GyrB action in the deadly human parasite P. falciparum but also provide meaningful insights into the current mechanistic model of DNA transport by gyrase during the topoisomerization reaction.

SUBMITTER: Dar A

PROVIDER: S-EPMC2772398 | biostudies-literature | 2009 Nov

REPOSITORIES: biostudies-literature

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A unique 45-amino-acid region in the toprim domain of Plasmodium falciparum gyrase B is essential for its activity.

Dar Ashraf A Prusty Dhaneswar D Mondal Neelima N Dhar Suman K SK

Eukaryotic cell 20090821 11

DNA gyrase is the only topoisomerase that can introduce negative supercoils into the DNA at the cost of ATP hydrolysis. Some but not all the steps of the topoisomerization reaction are understood clearly for both eukaryotic topoII and DNA gyrase. This study is an attempt to understand whether the B subunit of DNA gyrase binds to DNA directly, which may be central to the stimulation of its ATPase activity essential for gyrase function. We have dissected the Plasmodium falciparum gyrase B (PfGyrB) ...[more]

PMID: 19700639

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Project description:The target of rapamycin complex 1 (TORC1) pathway is a highly conserved signaling pathway across eukaryotes that integrates nutrient and stress signals to regulate the cellular growth rate and the transition into and maintenance of dormancy. The majority of the pathway's components, including the central TOR kinase, have been lost in the apicomplexan lineage, and it is unknown how these organisms detect and respond to nutrient starvation in its absence. Plasmodium falciparum encodes a putative ortholog of the RNA polymerase (Pol) III repressor Maf1, which has been demonstrated to modulate Pol III transcription in a TOR-dependent manner in a number of organisms. Here, we investigate the role of P. falciparum Maf1 (PfMaf1) in regulating RNA Pol III expression under conditions of nutrient starvation and other stresses. Using a transposon insertion mutant with an altered Maf1 expression profile, we demonstrated that proper Maf1 expression is necessary for survival of the dormancy-like state induced by prolonged amino acid starvation and is needed for full recovery from other stresses that slow or stall the parasite cell cycle. This Maf1 mutant is defective in the downregulation of pre-tRNA synthesis under nutrient-limiting conditions, indicating that the function of Maf1 as a stress-responsive regulator of structural RNA transcription is conserved in P. falciparum Recent work has demonstrated that parasites carrying artemisinin-resistant K13 alleles display an enhanced ability to recover from drug-induced growth retardation. We show that one such artemisinin-resistant line displays greater regulation of pre-tRNA expression and higher survival upon prolonged amino acid starvation, suggesting that overlapping, PfMaf1-associated pathways may regulate growth recovery from both artemisinin treatment and amino acid starvation.IMPORTANCE Eukaryote organisms sense changes in their environment and integrate this information through signaling pathways to activate response programs to ensure survival. The TOR pathway is a well-studied signaling pathway found throughout eukaryotes that is known to integrate a variety of signals to regulate organismal growth in response to starvation and other stresses. The human malaria parasite Plasmodium falciparum appears to have lost the TOR pathway over the course of evolution, and it is unclear how the parasite modulates its growth in response to starvation and drug treatment. Here, we show that Maf1, a protein regulated by TOR in other eukaryotes, plays an important role in maintaining the parasite's viability in the face of starvation and other forms of stress. This suggests that PfMaf1 is a component of a yet-to-be-described nutrient and stress response pathway.

Dataset Information

A unique 45-amino-acid region in the toprim domain of Plasmodium falciparum gyrase B is essential for its activity.

Publications

A unique 45-amino-acid region in the toprim domain of Plasmodium falciparum gyrase B is essential for its activity.

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