Molecular visualization of the yeast Dmc1 protein ring and Dmc1-ssDNA nucleoprotein complex.
Saccharomyces cerevisiae Dmc1, a meiosis-specific homologue of RecA, catalyzes homologous pairing and strand exchange during meiotic DNA recombination. The purified budding yeast Dmc1 (ScDmc1) protein exhibits much weaker recombinase activity in vitro as compared to that of the Escherichia coli RecA protein. Using atomic force microscopy (AFM) with carbon nanotube tips, we found ScDmc1 forms rings with an external diameter of 18 nm and a central cavity of 4 nm. In the presence of single-stranded DNA (ssDNA), the majority of the ScDmc1 protein (90%) bound DNA as protein rings; only a small faction (10%) was able to form filamentous structure. In contrast, nearly all RecA proteins form fine helical nucleoprotein filaments with ssDNA under identical conditions. RecA-mediated recombinase activity is initiated through the nucleation of RecA onto ssDNA to form helical nucleoprotein filaments. Our results support the notion that ScDmc1 becomes catalytically active only when it forms a helical nucleoprotein filament with ssDNA.
Chang
Yuan-Chih
YC
Institute of Physics, Academia Sinica, Taipei 11529, Taiwan.
Lo
Yu-Hui
YH
Lee
Ming-Hui
MH
Leng
Chih-Hsiang
CH
Hu
Su-Ming
SM
Chang
Chia-Seng
CS
Wang
Ting-Fang
TF
D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D004271:DNA, Fungal / Q000737:chemistry* / Q000235:genetics; D004277:DNA, Single-Stranded / Q000737:chemistry* / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D046911:Macromolecular Substances; D018625:Microscopy, Atomic Force; D037742:Nanotubes, Carbon; D009698:Nucleoproteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics
D018797:Cell Cycle Proteins; C513033:DMC1 protein, S cerevisiae; D004271:DNA, Fungal; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D037742:Nanotubes, Carbon; D009698:Nucleoproteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins