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Sequence‐specific termination by T7 RNA polymerase requires formation of paused conformation prior to the point of RNA release
Author(s) -
Song Hoseok,
Kang Changwon
Publication year - 2001
Publication title -
genes to cells
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.912
H-Index - 115
eISSN - 1365-2443
pISSN - 1356-9597
DOI - 10.1046/j.1365-2443.2001.00420.x
Subject(s) - termination factor , terminator (solar) , rna , biology , antitermination , duplex (building) , polymerase , transcription (linguistics) , rna polymerase , elongation , dna , biophysics , microbiology and biotechnology , biochemistry , gene , physics , materials science , ionosphere , linguistics , philosophy , astronomy , ultimate tensile strength , metallurgy
Background The sequence‐specific, hairpin‐independent termination signal for the bacteriophage RNA polymerases in Escherichia coli rrn B t1 terminator consists of two modules. The upstream module includes the conserved sequence and the downstream one is U‐rich .Results Elongation complexes of T7 RNA polymerase paused 2 bp before reaching the termination site at a 500 µ m concentration of NTP. At 5–50 µ m NTP, however, they paused and terminated there or resumed elongation beyond the termination site. Only at higher concentrations of NTP (500 µ m ), the pause complex proceeded slowly to and became incompetent at the termination site. At 4 bp or more before the termination site, the unprotected single‐stranded region of transcription bubble shrank at the trailing edge to 4–5 bp from ≈10 bp, resulting from duplex formation of the conserved sequence. The pause and bubble collapse were not observed with an inactive mutant of the termination signal .Conclusion Sequence‐specific termination requires the slow elongation mode of paused conformation, working only at high concentrations of NTP for a few bp prior to the RNA release site. The collapse of bubble that was observed several base pairs before the termination site and/or the resulting duplex might subsequently lead to the paused conformation of T7 elongation complexes .