Figure 7. ssDNA binding properties of EcSSB versus the EcSSB Tyr22Ala Lys73Glu mutant.

(a) Occluded site sizes (nucleotides per tetramer) on poly(dT) plotted as a function of [NaCl] for the EcSSB Tyr22Ala Lys73Glu variant (orange circles) determined from analysis of stoichiometric titration curves, examples of which are shown in panel (b) (occluded site sizes for EcSSB from [24] are shown for comparison (green triangles). (b) Representative reverse equilibrium titrations of EcSSB Tyr22Ala Lys73Glu (0.2μM) with poly(dT) (monitoring intrinsic Trp fluorescence quenching) at different NaCl concentrations: 10mM – dark yellow, 40 mM – magenta, 100 mM – green, 300 mM – grey and 1 M – blue. The occluded site sizes were determined as described [57]. (c) and (d) Sedimentation coefficient distributions, c(s_20,W), for EcSSB (c) and EcSSB Tyr22Ala Lys73Glu (d), in complex with M13 ssDNA in 10 mM NaCl at different protein to DNA ratios, R₆₅, where R₆₅=[SSB_{tetr, tot}]×65/[M13ssDNA_{nts, tot}]. Data were analyzed as described [10].