Switch-like control of helicase processivity by single-stranded DNA binding protein.

Area of Science:

• Molecular biology
• Biochemistry
• Structural biology

Background:

• Helicases unwind nucleic acid duplexes using NTP hydrolysis.
• Cellular helicase function is modulated by associated proteins, but mechanisms are unclear.
• Superfamily 2B helicases, like XPD, are crucial in DNA repair and replication.

Purpose of the Study:

• To elucidate the regulatory mechanism of XPD helicase activity by RPA2.
• To investigate how accessory proteins influence helicase processivity.
• To understand the molecular basis of helicase regulation by protein-protein interactions.

Main Methods:

• Utilized optical trap measurements to monitor single XPD helicase unwinding activity.
• Investigated the effect of replication protein A 2 (RPA2) on XPD translocation and unwinding.
• Introduced a point mutation at a regulatory DNA binding site on XPD to assess its impact.

Main Results:

• XPD helicase exists in two states with differing processivities.
• Transient interactions with RPA2 stabilize the highly processive state of XPD.
• A mutation at an XPD DNA binding site mimicked RPA2's stabilizing effect, activating a 'processivity switch'.
• These findings reveal a novel mechanism for helicase regulation by accessory proteins.

Conclusions:

• RPA2 acts as a regulator, stabilizing a more processive XPD state via transient interactions.
• XPD possesses a latent 'processivity switch' activated by RPA2 or specific mutations.
• This study provides critical insights into how accessory proteins modulate helicase function at a molecular level.