Coliphage 434 tof Protein: NH2-terminal amino acid sequence and kinetic and equilibrium measurements of DNA binding.

Coliphage 434 tof protein was purified to a substantially pure state from lambda imm434 cI dv carrier cells. The minimum molecular weight is 7,500 +/- 500 as estimated by polyacrylamide gel electrophoresis. The amino acid sequence of the nine NH2-terminal residues was determined, by manual Edman degradation of the intact protein, to be Met-Gln-Thr-Leu-Ser-Glu-Arg-Leu-(Lys)-. The purified protein at low concentrations binds specifically to lambda imm434 dv DNA and at high concentrations also binds to lambda imm21 dv and lambda dv DNA. The curve of the specific binding is of Michaelis type, while that of the nonspecific binding is sigmoidal. The specific binding does not show marked temperature dependency at 4 degrees - 37 degrees C. We have analyzed the equilibrium and kinetic data of specific binding. The equilibrium dissociation constant is 1.9 X 10(-11) M at O degree C. The association rate constant and the dissociation rate constant are 1.1-2.9 X 10(8) M-1s-1 and 2.7 X 10(-3)s-1, respectively, at 0 degrees C. The half life of the tof protein-operator DNA complex is 260 s. These results suggest that the tof protein-operator interaction is much weaker than the interaction between the cI repressor and the operator reported by other workers.