Fig. 4: PabRPA assembles as a tetrameric supercomplex that dissociates upon binding to ssDNA.
From: DNA-binding mechanism and evolution of replication protein A
a Characterization of PabRPA oligomeric states at low concentrations by using mass photometry. Expected molecular weights for monomeric (M), dimeric (D), and tetrameric (T) PabRPA supercomplexes are indicated with dotted lines. b SEC-SLS characterization of PabRPA ΔWH and PabRPA Tri-C in the presence or absence of poly-d25T ssDNA. The theoretical (MWth) and calculated molecular weights (MWcalc) for each complex are given in kDa. PabRPA complexes were injected at 20 µM. c 3.4 Å cryo-EM structure of the tetrameric PabRPA super-structure. d Focused view on critical contacts within the PabRPA tetrameric assembly. e SEC profiles of PabRPA, primase, or PabRPA-primase complexes, injected at 20 µM. The experiment was repeated three times independently. f Specific binding of PabRPA, ΔWH, or ΔAROD mutants at 1 µM to immobilized primase measured by BLI. Source data are provided as a source data file. g Schematic representation of a DNA replication fork in Archaea. Four PabRPA molecules clustered within the tetramer could efficiently coat and protect the stretches of ssDNA created by the advancing replisome. PabRPA tetramers may also play a role in recruiting and delivering the primase to DNA, thereby contributing to the efficient synthesis of Okazaki fragment, which requires repeated priming by the DNA primase.
