The biochemistry and fidelity of synthesis by the apicoplast genome replication DNA polymerase Pfprex from the malaria parasite Plasmodium falciparum

Scott R. Kennedy, Cheng Yao Chen, Michael W. Schmitt, Cole N. Bower, Lawrence A. Loeb

Research output: Contribution to journalArticlepeer-review

12 Citations (Scopus)

Abstract

Plasmodium falciparum, the major causative agent of human malaria, contains three separate genomes. The apicoplast (an intracellular organelle) contains an ∼ 35-kb circular DNA genome of unusually high A/T content (> 86%) that is replicated by the nuclear-encoded replication complex Pfprex. Herein, we have expressed and purified the DNA polymerase domain of Pfprex [KPom1 (Klenow-like polymerase of malaria 1)] and measured its fidelity using a LacZ-based forward mutation assay. In addition, we analyzed the kinetic parameters for the incorporation of both complementary and noncomplementary nucleotides using Kpom1 lacking 3′ → 5′ exonucleolytic activity. KPom1 exhibits a strongly biased mutational spectrum in which T → C is the most frequent single-base substitution and differs significantly from the closely related Escherichia coli DNA polymerase I. Using E. coli harboring a temperature-sensitive polymerase I allele, we established that KPom1 can complement the growth-defective phenotype at an elevated temperature. We propose that the error bias of KPom1 may be exploited in the complementation assay to identify nucleoside analogs that mimic this base-mispairing and preferentially inhibit apicoplast DNA replication.

Original languageEnglish
Pages (from-to)27-38
Number of pages12
JournalJournal of Molecular Biology
Volume410
Issue number1
DOIs
Publication statusPublished - 2011 Jul 1

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Molecular Biology

Fingerprint Dive into the research topics of 'The biochemistry and fidelity of synthesis by the apicoplast genome replication DNA polymerase Pfprex from the malaria parasite Plasmodium falciparum'. Together they form a unique fingerprint.

Cite this