Fig. 4.
Gam binding stimulates E. coli DNA LigA activity. (A) The 550-bp substrate DNA (C) with noncomplementary sticky ends (EcoRI/SalI) was incubated with increasing molar ratios of tagless Gam before the addition of E. coli LigA (Left) or T4DNA ligase (Right). DNA:Gam molar ratios in Left were 1:10, 1:20, and 1:40; only the first two ratios were used in Right. Reaction products were analyzed by agarose gel electrophoresis and visualized with EtBr. (B) Representative kymograph of FLAG-Gam (green) colocalizing with (HA)2-Lig A (magenta). White arrow denotes LigA dissociation. The half-life of LigA with Gam localization is ∼3 min (n = 25).