Initiation and re-initiation of DNA unwinding by the E. coli Rep helicase
نویسندگان
چکیده
Figure I. a. In the absence of ATP, EFRET distributions of DNA III (Fig. 2) at various [Rep] showed at least four conformations. EFRET was determined from each DNA by averaging the first second of observation. The peak at EFRET=0 is due to inactive acceptor dyes and will not be discussed further. With no Rep in solution, the distribution is peaked at EFRET=0.7, reflecting the DNA-only state (S). At sub-nanomolar [Rep], two new peaks appear at EFRET =0.95 and 0.5 (ES and ES* states, respectively) while the DNA-only peak decreases. The two conformations, ES and ES*, persist up to 20nM Rep, whereas above this concentration, a new broad peak appears near EFRET =0.25 reflecting binding of two or more proteins. b. The EFRET time trace upon addition of a Rep solution shows two abrupt transitions: At t=0, 13.4nM Rep was added. DNA starts from the S state and shows stepwise changes to the ES state, and then to the ES* state. From each trace, 0 15 0 1
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