Prokaryotic transcription Flashcards
RNA polymerase properties
(3)
- builds up RNA chain by attaching 5’-P of an NTP to the 3’OH of the nucleotide preceding it
- does not need a pre-existing primer to initiate synthesis of a new chain
- Composed of a sigma subunit that recognizes promoter and the core RNA pol complex is made up of 5 subunits
Desribe the bacterial RNA polymerase holoenzyme
6 subunits, 5 core subunits and 6th sigma unit is needed for initiation and cycles in and out of complex
Mechanism of RNA synthesis
addition of an rNTP in a Mg2+ dependent reaction that produces 5-3 phosphodiester linkages
What is the other strand called that is not the template
coding strand or non-template strand
direction of RNA synthesis
5’-3’
what is the transcription bubble?
the RNAP maintains a bubble of melted DNA to reveal the template strand
DNA duplex is unwound for about 17 bp, forming a bubble, which enables RNA polymerase to access the template strand.
What are promoters
regions of DNA where the RNAP holoenzyme can bind the dsDNA and begin transcription at a transcriptional start site on the template strand
Promoters recognized by the same sigma factor share a particular ______ _______
consensus sequence
Which promoters do RNAP bind to
sigma70
o-70
How can you predict the strength of a promoter
Promoters with a close match to the sigma 70 consensus and have an UP element are especially strong promoters
three distinct phases of RNA synthesis
initiation: RNA polymerase binds to promoters
Elongation: process of adding nucleotides to the growing RNA strand
Termination: release of the product RNA when the polymerase reaches the end of a gene
formation of transcription bubble happens before all
T/F RNA polymerase requires a primer like DNA polymerase
F
what is the -10 region
site of strand melting that reveals the template strand
binds sigma 2
describe the conversion of the closed complex to the open complex by sigma factors
(2)
- the DNA is melted around the -10 element to reveal the template strand
- The sigma factor repositions away to open the channel through which NTPs must enter the polymerase active site
Describe the transcription initiation
After open complex formation reveals the template strand the RNAP holoenzyme starts transcribing from the +1 position
How can extension of the RNA chain be halted
can be halted by a steric clash of the growing RNA chain with the s subunit, this sometimes causes transcription to abort after extension of the RNA chain to 8-9 nt, otherwise s is released and the RNAP core escapes the promoter
What is promoter clearance
movement of the transcription complex away from the promoter, which marks the beginning of the elongation stage of transcription
What is abortive initiation
release of an 8 to 10 base pair RNA transcript from the bacterial RNA polymerase initiation complex before it clears the promoter and enters the elongation stage.
what is the sigma facotr
directs the enzyme to the promoter
What is a consensus sequence
DNA or AA sequence consisting of residues that commonly occur
what is the transcription elongation complex
composed only of the RNAP core without the sigma factor. Initiates the processive function of elongation until termination signal is encountered
What is the transcription termination
signals for RNA extension to stop
2 types
1) rho-independent terminators
2)Rho-dependent terminators
Rho-independent terminators
composed of 2 elements
a) an inverted repeat that can form a hairpin structure in the RNA transcript
b) a run of A’s in the DNA that promote pausing and RNAP release (U’s in transcript)
mRNA forms a hairpin followed by three U residues, stalling and separating the polymerase from the mRNA
Rho-dependent terminators
1) A rut site where the Rho helicase loads
2) a string of A’s in the template strand shortly after the rut site. This slows down the RNAP core to allow Rho to catch up to the RNAP core complex.
Describe the regulation of gene expression in bacteria
Bacteria need some proteins most of the time, like the proteins involved in DNA replication so they are expressed at most times. However, many other genes are only expressed when their products are needed by the cell
When is it most efficient to regulate gene expression
Transcription initiation step or shortly after because gene expression is very expensive
Describe the regulation of the gal operon (mechanism, repressors, operators)
repressed when galactose is absent. When present, GalR binds galactose and falls off the operator removing the repression
2 repressors: GalR and GalS
2 operators: Oe(external), Oi(internal)
GalR is bound to Oe and Oi that represses Gal
What is catabolite repression? what is it mediated by
Bacterial cells prefer to use glucose as an energy and carbon source and repress expression of operons for utilizing other sugar sources until the glucose is used up
this process is mediated by cAMP and cAMP receptor protein (CRP)
Describe the cAMP-CRP mechanism
Binds to operator sites near the promoters of sugar utilization operons and stimulate transcription of these neurons when glucose is absent
Acts like an accelerator
what is a corepressor
a protein or small molecule that inhibits transcription by binding a repressor or repressors, without binding the DNA directly
What is the trp operon in E. coli? Describe the regulation
produces enzymes needed to form the amino acid tryptophan
In the absence of tryptophan the Trp repressor cannot bind the operator and transcription is initiated, tryptophan serves as an effector molecule for the repressor to bind the operator
What is transcriptional attenuation
a process for the regulation of expression of a bacterial operon in which transcription constitutively initiated but is halted in the leader region.
Termination occurs if products are not needed in the cell
What is the leader sequence
a short sequence near the 5′ end of an RNA that has a specialized targeting or regulatory function
what is a leader peptide
a short sequence near the amino terminus of a protein that has a specialized targeting or regulatory function
What is the SOS response
a coordinated induction of a variety of genes in response to high levels of DNA damage
LexA interaction with RecA-ssDNA causes LexA to inactivate itself. Lack of LexA allows SOS to be expressed
What is a riboswitch
Structure within an mRNA that binds to a specific ligand and undergo a conformational change to regulate gene expression affecting the translation or processing of the mRNA.
Describe the expression of the B. subtilis trp operon
responsive to cellular tryptophan
uses a tryptophan sensing RNA-binding repressor
What is the TRAP
Binds to mRNA if it is itself bound to tryptophan
Binding of TRAP to leader RNA sequesters the RNA at an essential point (antiterminator point)
What is the glmS ribozyme
enzyme needed for the biosynthesis of a cell wall component in B. subtilis
the glmS riboswitch is a ribozyme that binds to glucosamine 6-phosphate
upon binding, the glmS cleaves itself and promotes its own degradation
