Disentangling polydispersity in the PCNA-p15PAF complex, a disordered, transient and multivalent macromolecular...

Area of Science:

• Structural Biology
• Molecular Biophysics
• Biochemistry

Background:

• The intrinsically disordered p15PAF protein is crucial for DNA replication and repair.
• p15PAF interacts with the Proliferating Cell Nuclear Antigen (PCNA) sliding clamp.
• The transient, low-affinity complex between p15PAF and PCNA is challenging to study structurally due to polydispersity.

Purpose of the Study:

• To determine the structure, conformational dynamics, and population of the p15PAF-PCNA complex in solution.
• To elucidate the binding mode of p15PAF to the PCNA ring.
• To understand the role of p15PAF in regulating PCNA function during DNA repair.

Main Methods:

• Small-angle X-ray scattering (SAXS) was combined with molecular modeling.
• Integrative approaches and molecular dynamics (MD) simulations were used to build explicit ensemble descriptions.
• SAXS profiles were interpreted as population-weighted thermodynamic mixtures.

Main Results:

• The study determined the structure, conformational fluctuations, and relative populations of five coexisting species in the p15PAF-PCNA complex.
• The N-terminus of p15PAF was observed to penetrate the PCNA ring and emerge on the back face.
• This binding mode supports the role of p15PAF as a regulator of PCNA processivity in DNA repair.

Conclusions:

• Ensemble-based approaches utilizing SAXS data are powerful for decoding structural, dynamic, and thermodynamic information of flexible macromolecular assemblies.
• The findings provide structural insights into the mechanism of p15PAF in DNA repair.
• This strategy can be applied to study other transient and multivalent protein complexes involved in critical biological processes.