SDS-PAGE and biological activity of purified SSB
. (a) SDS-PAGE analysis of SSBBA (fraction V) used in this study. (b) FRET helicase analysis of DnaBBA in the presence and absence of SSBBA: Emission spectra of the substrate (4.2 nM), after 15 min incubation with SSBBA (3 μg/ml) DnaBBA (0.5 μg/ml), and both SSBBA and DnaBBA at 37°C; (c) Kinetic analysis of helicase activity: The helicase substrate was rapidly mixed with indicated protein(s) and fluorescence emission at 662 ± 8 nm was recorded as a function of time for 500 s using a Slow Kinetic mode in PC1 spectrofluorometer using Vinci software (ISS Inc. Champaign, IL). Using FRET spectra of native and heat denatured substrate, we predetermined that 1% decrease in FRET is equivalent to ~3.1 pmol of nucleotide (or bp) unwinding of duplex DNA.