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Pif1 is a force-regulated helicase.

Jing-Hua Li1, Wen-Xia Lin1, Bo Zhang2

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|April 22, 2016
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Summary
This summary is machine-generated.

The DNA helicase Pif1 unwinds DNA efficiently only at high protein concentrations. Research shows that applied force significantly enhances Pif1's DNA unwinding rate and length, suggesting force regulation of its activity.

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Area of Science:

  • Molecular Biology
  • Biophysics

Background:

  • Pif1 is a 5' to 3' DNA helicase with high DNA binding affinity but low unwinding processivity.
  • Efficient DNA unwinding by Pif1 typically requires high protein concentrations for dimerization.

Purpose of the Study:

  • To investigate the DNA unwinding activity of Saccharomyces cerevisiae Pif1 (Pif1) using single-molecule techniques.
  • To determine the effect of mechanical force on Pif1's DNA unwinding processivity and efficiency.

Main Methods:

  • Single-molecule fluorescence resonance energy transfer (smFRET).
  • Magnetic tweezers (MT) to apply controlled forces to forked double-stranded DNA (dsDNA).

Main Results:

  • Pif1 exhibits repetitive unwinding and re-zipping cycles at zero force but has limited processivity per cycle.
  • Applied force significantly enhances Pif1's unwinding rate (over 3-fold at 9 pN) and total unwinding length (over 20-fold at 9 pN).
  • Low processivity at zero force explains inefficient unwinding in bulk assays at low protein concentrations.

Conclusions:

  • Pif1's DNA unwinding activity is significantly regulated by mechanical force.
  • Force-dependent regulation may be crucial for Pif1's various cellular functions.