FC0060
Replication protein A (RPA)  -  DNA

Biological function
The human replication protein A (RPA) participates in both nucleotide excision repair and combinatorial repair using multiple ssDNA binding domains.

Domain organization/sequence features
The 70-kDa subunit of human RPA (hRPA70) contains weak and high-affinity DNA binding domains (DBDs), connected by a 78- amino-acid intrinsically unstructured linker domain (IULD).

Structural evidence
Backbone flexibility of several IULD homologues using nuclear magnetic resonance (NMR) spectroscopy. The backbone flexibility of five IULDs, representing three kingdoms, was measured and analyzed. Two IULDs from animals, one IULD from fungi, and two IULDs from plants showed similar levels of backbone flexibility that were consistent with the absence of a compact globular structure.

Biochemical evidence
RPA makes its first contacts with ssDNA using the high-affinity DBDs. The highly flexible IULD anchors the weak affinity DBD to DNA, thereby increasing its local concentration near the substrate. Conversely, the loss of the transient IULD–weak affinity DBD interaction results in a threefold reduction in binding to ssDNA. The functional relevance of the IULD is highlighted by point mutations isolated in the IULD from the yeast RPA70 homologue that inhibit the response to UV-induced DNA damage.

Structure/Mechanism
Transcriptional activation of the tumor suppressor p53 depends on its interactions with hRPA70. Binding sites for ssDNA and p53 overlap on the weak affinity DBD of hRPA70. The affinity of hRPA70 for ssDNA is higher than that for p53, and is primarily controlled by the disordered IULD. The ID region increases the local concentration of the weak-affinity DBD near its cognate site, which results in the occlusion of p53 from the basic cleft of hRPA70.

Mechanism category
Tethering

Significance
Thus, depending on its orientation, flexibility and length, the fuzzy hRPA70 IULD regulates the balance between binding p53 and damaged DNA.