RNA annealing activity of Ltp26/Ltp28 100-kD complex. (A) Specificity and cofactor requirements. Panel 1: lane 1, RNA-III at 2 nM was annealed with 50-fold excess of radioactively labeled RNA-IV by heating and slow cooling prior to addition of T1 nuclease in the absence of added protein. Lane 2, labeled RNA-IV + Ltp26/Ltp28 complex in the absence of complementary RNA followed by T1 nuclease digestion. Lane 3, RNA-III + labeled RNA-IV (2 nM each) in absence of protein, followed by T1 nuclease digestion. Lane 4, labeled RNA-IV + noncomplementary RNA-VI of approximately the same length + Ltp26/Ltp28 complex, followed by T1 nuclease digestion. Lane 5, RNA-III + labeled RNA-IV (2 nM) + Ltp26/Ltp28 complex, no T1 nuclease. Lane 6, labeled RNA-IV, no T1 nuclease. Panel 2: Ltp26/Ltp28 complex was added at increasing concentrations to RNA-III + labeled RNA-IV in absence of added NTPs, followed by T1 nuclease digestion. Panel 3: Same as Panel 2 but with 1mM GTP. Panel 4: Same as Panel 2 but with 1 mM ATP. Panel 5: Same as Panel 2 but without Mg2+. (B) Kinetics of annealing catalyzed by the recombinant p26 and p28 proteins and by the native 100-kD complex at 50 nM . SI, signal intensity of the T1-protected fragment. A control for the kinetics of annealing in the absence of protein is also shown.
