Exam 4: lecture 12-transcription, mRNA, and splicing Flashcards
function of promoter
recognition site for RNA to bind
tell rna to start (me, me, me)
What is the direction that RNA is read?
5’ by 3’
What is another name for template strand?
sense/ coding strand
what is another name for the non-template strand/
antisense
Does mRNA can be made wo a primer?
yes, de novo (from newness)
-added @ the 3’ end of growing chain
-they can be read one direction
How to know where transcription?
based on distance
-the beginning of the polymerase
the sequence of prokaryotes
-only has 1 RNA polymerase
TTGACAT ————————————–TATAAT
-35 region (15-17bp) -10 (the most common sequence
-> TATAAT box is 10 bp upstream of the polymerase (where transcription starts)
->15-17 bp(Y: any pyrimidine)
Why do certain sequences appear in all organisms?
they are conserved- not loss in evolution
where does transcription starts
<——upstream———–+1———–downstream———->
Eukaryotic promoters: RNA polymerase II
TATA box: -25
Py2CAPy5 (initiator) near +1
Eukaryotic promoters: RNA polymerase I
the majority of the rRNAs
-core promoter: -45—+1—–+20 (core promoter)
-upstream control element: -180+-107
Eukaryotic promoters: RNA polymerase III
-mostly tRNA
- tRNA gene
+1—box A—-box B - 5S-rRNA gene
+1————–box A—–box B
What is an overview of transcription?
5’ —–===(promoter)===————===(terminator)====3”
3’—– ===(promoter)===————===(terminator)====5”
1) RNA polymerase binds & unwind DNA
—–( )———— (RNA polymerase binds to promoter)
2) Initiation of RNA synthesis (+NTPs)
5’—–(coding-antisense)
3’—–(template-sense) /<- RNA
3) elongation of RNA + further elongation
(elongates until termination signal)
4)Termination of RNA synthesis
5’—————————-3’
RNA transcript
what happens in Rho-independent termination regarding a hairpin loop?
+not dependent on enzymes
To terminate transcription, the RNA transcript goes towards each other that form a stem+ loop where the bases are complementary
-the loop leading to termination will be near the end of the transcription site- only when transcription is complete.
small loop: pauses
big loop: cause everything to dissociate
what happens in rho dependent termination
Rut, an enzyme that is being used as Rho utilization site
-found in mRNA near the end of transcription
Procaryotic mRNA vs Eukaryotic mRNA
Prokaryotic mRNA:
+5’ & 3’ untranslated regions
+usually polycistronic (a sequence that coordinately transcribed)
=> Operon: a cluster of genes that are transcribed together to give a single messenger RNA (mRNA) molecule, which therefore encodes multiple proteins
O: a: b: c
+is mature after being transcribed (ready to be translated immediately after)
+can be both transcribed & translated at the same time
Eukaryotic mRNA;
+5’ & 3’ untranslated regions
+ is not polycistronic
+is not mature after being transcribed (RNA has to move out of nucleus to be translated)
+can not be both transcribed and translated at the same time
=transcription: in the nucleus
=translation: cytoplasm
maturation of eukaryotic mRNA
1) addition of cap- help bind to ribosome
2) addition of poly(A) tail- stabilizing RNA (short lived)
3) intron excision:
-introns: interrupting sequences that needed to be removed because exons needed to be connected
What is the “standard” type of intron?
recognition sequence:
+5’: GU - 100% conserved
+3’: AG
(branch point(A)——-20-50bp———–(AG)3’ splice site
how are introns cut out
the mRNA goes thru the splicing enzyme complex
the strand goes thru small nuclear RNA
-3’ ontop/ 5’ inside the RNA
an OH connects the extrons tgt and is removed w the lariat structure
how many bases are in a codon?
1? no, bc 20 aa, 4 diff singlet codons would not cover it
2? no , bc 20aa, there are only 16 possible doublet combinations
3? YES, there would be 64 possibilities of triplets that covers all the possible aa
=> Occam’s razor: the simplest explanation is the best-unless proven otherwise
what is the central dogma
DNA—-transcription—–> RNA—–translation—–> protein
things to know about codons
- codes are degenerate “redundant”
- every gene start w AUG (met)
3: stop codons (UAA,UAG,UGA) - some codes only changes the 3rd position (Leu, Ser, Thr, Val, Ala)- allows leeway in 3rd position
- universal
+codon usage: some organism uses certain sets more regularly
the molecular basis of sickle cell disease+ an example of what mutation?
normal hemoglobin DNA : CTT
mRNA : GAA
normal hemoglobin : Glu (hydrophilic)
mutant hemoglobin DNA: CAT
mRNA :GUA
sickle cell hemoglobin: : Val (hydrophobic)
=subtitution mutation: replace 1 letter by another
causes:
hard radiation
chemicals
UV
viruses
Mutation: Base insertion
+ another letter
=> reading frame will be out of position
mutation: Base substitution
replace 1 letter with another
=> change the aa
mutation: base deletion
a letter is taken out
mutation: nonsense
UAC (Tyr)—> UAG (stop)
putting a stop codon
mutation: missense
altering from one aa to another
mutation: frame shift
base deletion/ base insertion
mutation: null
replacing w something chemically similar
dna change–> mRNA change–> AA (not losing much function)
mutation: silent
have a mutation but do not have any effect
-a change in DNA, but no change in mRNA
In order to get translation, what is needed?
a. an mRNA transcript
b. a ribosome (rRNA)
c. charged tRNA (w aa attached)
Bases in the anticodon
Alanine —O(ester bond)—ACC [conserved]
UH2: altered bases (dihydrouracil)
M2G: dimethyl guandine
MG: methyl guadine
mI: methyl Inosine
Y: pseutouracil
structure of tRNA
all tRNA have similar but not identical structure =cloverleaf
*~75 nt
*acceptor arm: CCA-3’
*TYC arm
*variable length extra arm (3~20nt)
*anticodon arm
*D arm
*canonical position
*identity elements
What occurs when tRNA(ala)-> Alanyl-tRNA(ala)
tRNA(ala0
3’–HO–ACC
===========
+Alanine
=========
ester bond+ ACC
What is the process of aa activation by aminoacyl-tRNA synthetase?
-aa & pyrophosphate bind to aminoacyl-tRNA synthetase (PPP–> P: 2 r taken out)
-tRNA bind to whats left of of the one P
= aminoacyl tRNA (“activated aa”) is an AA+tRNA
what is the wobble postion in tRNA anticodon loop?
refers to the 3rd nucleotide in a codon
its much looser in this position so that its easier reduce the number of tRNA needed
-61 codons codes for aa (1 tRNA for each)
wobble hypothesis
-proposed by francis crick in 1966
-occurs at 3’ end of codon/5’ end of anticodon
-result of the arrangement of H-bonds of base pairs @ 3rd pos.
-degeneracy of the code is such that wobble always results in translation of the same aa
-complete set of codons can be read by fewer than 61 tRNAs
5’ anticodon. 3’codon
G U/C
C G
A U
U A/G
I A,U,C
I(post-transcription modified purine)
What is the structure of prokaryotic rRNA
small subunit (30s)
-16S RNA
-21 proteins (S1(small)-S21): scaffold proteins that help support+ hold up 16sRNA to give it structure
large subunit (50s)
-5S and 23S RNA
-36 proteins [L1(large)-L36)
=70S
Eukaryote rRNA
40S+ 60S=80S
Ribosome structure in specific (prokaryotic)
23S: contains the peptidyl transferase activity (where ribosomal activity is; attach 1 aa to the next=makes a peptid bond)
16S: directly involved in both initiation & termination of translation
5S: mostly structural
=overall , structure is highly conserved
what is the Shine-Dalgarno sequence
this is where the binding of bacterial mRNA to ribosome takes place in the 5’ UTR
-allows mRNA to bind to the ribosome proten
5’ ———- SD——-N5-10-AUG-//—3’n
Initiation of translation in prokaryotes
- IFs bind to 30s ribosomal subunit
- Initiator tRNA (w/ fMet formyl methenamine) and mRNA binds to the 30s subunit
- 50S subunit binds thru GTP hydrolysis = 70S initiation complex
Function of IF 2
chaperones in initator tRNA (ensure tRNA get to its desired location)
function of IF3
chaperones the 30S subunit to help bind to S-D site
function IF1
hold small+large subunits from each other
initiation of translation of eukaryotes
- small subunit binds to the 5’ Cap
- scan for AUG
- find the AUG based on the kozak sequence nearby
Kozak sequence: mRNA 5’-ACCAUGG_
ribosomal scanning: move down the length of RNA until bind at the end of the initiation site (KZ)
the process of polypeptide chain elongation in prokaryotes
- binding of aminoacyl tRNA
- peptide bond formation
3.translocation
A site: AA site, new tRNA come in as new aa needed to be added
P site: polypeptide site, newly made sit as new aa come in (nascent: growing, unfinished)
E site: exit site
EF-Tu: chaperone protein for initiation
ET-Ts: carrier protein, carry Gtp transfer GTP to EF-TU (transfer energy)
EF-G: move the ribosome down 1 triplet
termination of translation
release factor (stop codon) binds to A site
-> causes the whole things to dissociate
