Chapter 29 Flashcards
splicing removes >
introns
homology between RNAPs
have central metal atom in active sites. important c terminal domain critical for processing RNa
3 steps of RNA transcription
initiation, elongation, termination
post-transcriptional processing in RNA
base modification (methylation), addition of nucleotides (poly A tails), splicing
RNA polymerases bing to DNA sequences called ?
promoters
In the absence of ? (2), RNAP can bind to promoters but cannot continue elongation steps
Mg++, NTPs
binding of ? protects DNa from enzyme digestion
RNAP
stronger promoters are better matches to ?
consensus
optimal promoters have ? bp between consensus sequences
17
changes in growth conditions lead to appearance of additional ?
sigma factors
housekeeping sigma factor
sigma 70
RNAP error
1 in 10^4
RNAP elongation reaction
PPPXOH + YTP»_space; PPPXPYOH + PPi
RNAP polymerization reaction is identical to DNA polymerase and also requires ? assisted phosphoryl transfer
Mg++
RNAP elongates at ? nucleotides/second
50
? polymerase needs no primer
RNA
RNAP uses free energy of ? to drive conformational change
PPi release
Incorporation of a ? changes the active site and pushes newly formed duplex base pair into the ? region. Release of PPi returns the active site to a ? binding region
Incorporation of a NMP changes the active site and pushes newly formed duplex base pair into the helix binding region. Release of PPi returns the active site to a NTP binding region
RNA:DNA hybrid is slightly more/less stable than DNA:DNA
more
determines when transcription stops
the RNA molecule being made
Three methods of Transcription pausing or termination
1) form stem loop 2) Recruit accessory proteins (attenuators) 3) additional factors can antagonize attenuation
rho independent termination
formation of step loop structure causes pause. if next residues are uridines, thranscription stops and rna is released. free energy of stem loop is greater than RNA:DNA duplex
Rho
a hexameric RNA helicase. binds to C rich regions. removes rna from polymerase.
rifampicin
inhibitor of many bacterial RNAPs. binds to aromatic AAs at site of formation of RNA:DNA hybrid and prevents initial elongation steps. Mutations in aromatic rings leads to rifampicin resistance.
In procaryotes, ? is directly used for protein synthesis
mRNA
rRNA and tRNA undergo these types of processing
Cleavage, modification, addition of nucleotides (methylation, flipping)
In E Coli, a primary transcript is cleaved by RNase 3 and RNase P to produce rRNAs and a tRNA that undergo ?. tRNAs have the nucleotide sequence ? added to their 3’ ends if needed
base modifications. CCA
eucaryotes have ? RNAPs differentiated by their sensitivity to ?
- alpha-amanitin
RNAP that is strongly inhibited by alpha-amanitin
type 2
RNAP that is inhibited by alpha-aminitin at high concentrations
type 3
A type 4 RNAP found in plants is used to make ?
siRNAs and RNA dependent RNA polymerases involved in RNA interference
Type 1 RNAPs used to make ?
rinosomal RNA
Type 2 RNAPs used to make ?
mRNA precursors and snRNA
different RNAPs transcribe from different ?
promoters
RNAP ? and ? produces RNAs needed by all cell types and have relatively simple sequences in their promoters
1 and 3
? promoters are complex
RNAP
eucaryotic RNAP binding
general transcription factors bind to promoter elements and recruit RNAPs
? elements of promoters are recognition sites for protein factors
cis
RNAP 2 is recruited by GTFs ?
TFIIA-TFIIH
In TATA-box promoters, ?, containing the TATA-binding protein binds first and distorts the DNA helix
TFIID
TATA box binding protein binds TATA box that contains sequences related to TATAAAA and ?
bends the DNA into a horseshoe shape
TFIID provides ? to form basal transcription apparatus
docking sites for additional GTFs and RNAP2
Assembly of transcription initiation complex generates a ? complex that determines the orientation of transcription
asymmetric
TFIIH functions
helicase activity opens DNA helix to start transcription. phosphorylation of C-terminal domain of RNAPII releases RNAP from initiation factors that recruited it and binds RNA processing enzymes
basal transcription complex initiates transcription at a low/high frequency
low
many promoters are activated by ? that may be far from transcription start sites either up or down stream
enhancer sequences
Cis promoter elements are recognized by ?
protein factors
complex called ? coordinates positive activators with recruitment of RNAP 2
mediator
SrB8-II represses
RNAP 2
RNAP 1 produces ? that undergo base and ribose modification in the nucleolus by ?
rRNA. snoRNPs
RNAP III transcript processing to tRNA
5’ and 3’ cleavage. additon of 3’ CCA sequence. base and ribose modification. spliced by endonuclease and ligase
RNAP II transcript processing to mRNA
5’-5’ triphosphate cap at 5’ end by GTP marks as pre-mRNA and protects from exonucleases. PolyA polymerase adds a tail to 3’ end. methylation can occur at 2’-OH groups of nucleotides 1 and 2
Tissue specific RNA editing. Liver produces mRNA that translates to ? while intestines generate a deaminase for the same mRNA that modifies it to translate to ?
ApoB-100. ApoB-48
ApoB-100
transports lipids to other cells
ApoB-48
Carries fat in chylomicrons
? is most common way eucaryotes generate diversity
splicing
splicing involves ? reactions
transesterification
splicing transesterification reactions are similar to tyrosine mediated reactions by ?
topo 2
Draw splicing reaction
do it. 2’ Oh of branch site attacks 3’ OH of phosphodiester bond at 5’ splice site to generate lariat intermediate. Newly generated 3’ OH attacks 3’ OH of phosphodiester bond at 3’ splice site
spliceosome
snRNPs that contain RNA and a large number of proteins assemble to help guide and catalyze splicing reactions
Since transesterification reactions are freely reversible, what is the energy source that drives splicing to proceed
ATP powered helicases drive the spliceosome through the conformational changes that involve the release of U1 and U4
specificity of splicing due to ?
complementary pairing of pre-mRNA and snRNAs
phosphorylation of CTD causes
release of RNAP from initiation complex and sequentially recruits enzymes of processing
15% of all genetic diseases due to ?
mutations that affect splicing
Cis effects
mutations in the pre-mRNA
Trans effects
mutations in the spliceosome
some defects in hemoglobin production are the result of a mutation within an ? that creates a new splice site
intron
ribozymes
catalytic RNAs
group 1 introns use ? to attack 5’ splice site
guanosine
group 2 introns and the spliceosome activate an ? to initiate splicing
adenine