help protect the growing RNA chains from degradation by nucleases. 3) RNA chain termination 4) RNA chain termination Occurs when RNA polymerase encounters a termination signal The transcription complex dissociates releasing the nascent RNA molecule There are two types of transcription terminators in E. coli Rho-dependent terminators – req. protein factor (ρ) and rut (rho utilization) seq. on transc. RNA Rho-independent terminators – req. inverted repeats on complementary strand, form hairpin, RNA poly. Falls off a 6A’s seq. 1. Cleavage of endonuclease usually occurs 30 nucleotides downstream from a conserved polyadenylation signal (consensus AAUAAA), and upstream from a GU- rich sequence 2. Addition of poly(A) tail by poly(A) polymerase. The actual transcription termination events often occur at multiple sites that are located 1000 to 2000 nucleotides downstream from the site that will become the 3’ end of the mature transcript transcription proceeds beyond the site that will become the 3’ terminus Rho-Dependent Termination Rho-Independent Termination The mechanism by which rho-dependent termination of transcription occurs is similar to that of rho-independent termination (i.e. both involve the formation of hairpin structures upstream from the site of termination as hairpins impede the movement of RNA polymerase, causing it to pause). Rho-dependent terminators also contain additional sequences: 1) These sequences are ~50 to 90 bps upstream from the inverted repeats. 2) Rho protein binding site called rut (for rho utilization) near the 3’ end of the transcript. The presence of the rho protein indicates the presence of a termination signal. When RNA polymerase encounters this signal, the transcription complex pauses and allows the rho protein to unwind the DNA/RNA base pairing and releasing the nascent RNA molecule. Rho protein binds to the rut sequence in the transcript and moves 5’ to 3’following RNA polymerase. When polymerase encounters the hairpin, it pauses, allowing rho to catch up, pass through the hairpin, and use its helicase activity to unwind the DNA/RNA base- pairing at the terminus and release the RNA transcript. Rho-independent terminators contain a GC-rich region containing inverted repeats followed by six or more AT base pairs, with the A’s present in the template strand. 1) Transcription of inverted repeats (on complementary strand) Sequences of nucleotides in each DNA strand that are inverted and complementary.
2)Formation of a hairpin structure on the nascent RNA that impedes transcriptionWhen transcribed, these inverted repeat regions produce single- stranded RNA sequences that can base- pair and form hairpin structures.3)RNA Pol pauses and falls off at the ‘AAAAAA’ sequenceRNA hairpin structures form immediately after the synthesis of the participating regions of the RNA chain and retard the movement of RNA polymerase molecules along the DNA, causing pauses in chain extension.AU base-pairing is weak and therefore requires less energy to separateIn prokaryotes, a mutation that deletes the region from -10 to -35 will seriously down-regulate transcription of that gene.
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- Fall '13