کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2830262 1163372 2008 12 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Expression and biochemical characterization of the Plasmodium falciparum DNA repair enzyme, flap endonuclease-1 (PfFEN-1)
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شناسی مولکولی
پیش نمایش صفحه اول مقاله
Expression and biochemical characterization of the Plasmodium falciparum DNA repair enzyme, flap endonuclease-1 (PfFEN-1)
چکیده انگلیسی

Flap endonuclease-1 (FEN-1) is a structure-specific endonuclease that is critical for the resolution of single-stranded DNA flap intermediates that form during long patch DNA base excision repair (BER). This investigation reports that Plasmodium species encode FEN-1 homologs. Protein sequence analysis revealed the N and I domains of Plasmodium falciparum (PfFEN-1) and Plasmodium yoelii (PyFEN-1) to be homologous to FEN-1 from other species. However, each possessed an extended C domain which had limited homology to apicomplexan FEN-1s and no homology to eukaryotic FEN-1s. A conserved proliferating cell nuclear antigen (PCNA)-binding site was identified at an internal location rather than the extreme C-terminal location typically seen in FEN-1 from other organisms. The endonuclease and exonuclease activities of PfFEN-1 and PyFEN-1 were investigated using recombinant protein produced in Escherichia coli. Pf and PyFEN-1 possessed DNA structure-specific flap endonuclease and 5′ → 3′ exonuclease activities, similar to FEN-1s from other species. Endonuclease activity was stimulated by Mg2+ or Mn2+ and inhibited by monovalent ions (>20.0 mM). A PfFEN-1 C-terminal truncation mutant lacking the terminal 250 amino acids (PfFEN-1ΔC) had endonuclease activity that was ∼130-fold greater (kcat = 1.2 × 10−1) than full-length PfFEN-1 (kcat = 9.1 × 10−4) or ∼240-fold greater than PyFEN-1 (kcat = 4.9 × 10−4) in vitro. PfFEN-1 generated a nicked DNA substrate that was ligated by recombinant Pf DNA Ligase I (PfLigI) using an in vitro DNA repair assay. Plasmodium FEN-1s have enzymatic activities similar to other species but contain extended C-termini and a more internally located PCNA-binding site.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Molecular and Biochemical Parasitology - Volume 157, Issue 1, January 2008, Pages 1–12
نویسندگان
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