Final Exam Flashcards
what part of an mrna sequence helps e.coli distinguish the start codon from a methionine codon
shine delgarno sequence (towards the 5’ end of AUG)
what modification helps e.coli distinguish between an initiator trna and a regular trna
fMet
what does IF3 do
inhibits premature formation of 70S ribosome
what does EF-Tu do
delivers aminoscyl-trna to the A site
binds GTP/GDp
what does the 50S subunit do
forms the peptide bond
what does IF2 do
delivers fMet-tRNA to the P site
binds GTP/GDP
what does eIF4f do
initiates cap dependent translation
what does EF-G do
binds GTP/GDP
promotes translocation of the ribosome
what proteins/complexes hydrolyze GTP during translation
EF-Tu, RF3, IF2, EF-G
what functional change must be made to Cas9 to be used for CRISPRi or CRISPRa
mutate so it wont cut (dCAS9)
how can you modify the cas9 system in order to inhibit gene expression without modifying the DNA sequence
fuse a protein domain that will prevent binding of RNA pol III
dCAS9 will target specific region, inhibitor domain will prevent transcription, reducing gene expression
T or F: the 30S ribosome associates with the mrna before the 50S
true
T or F: N-formylmethionyl-trna^fmet binds in the p site
true
T or F: EF-Tu-GTP binds to the 50S ribosome
false
T or F: IF-1, IF-2, IF-3 dissociate before the 50S subunit binding
true
describe the wobble hypothesis
the variance in base pairing is greater at the 5’ end of the anticodon, thus accounting for more than one codon
non WC base pairing is sometimes allowed in the last position of the codon
G and U can pair as well
are trna molecules ds or ss
single stranded RNA
which terminal of the trna is the site of AA attachment
3’ terminal
T or F: the anticodon arm of trna contains a three nucleotide sequence that is identical to a specific mrna codon
false
can a single trna bind to multiple mrna codons?
yes
where does the energy to create a peptide bond in the peptidyl transferase reaction come from
energy is stored in ester bond, positioned by EF-Tu and peptidyl transferase
is a promoter region found within the mrna transcript
no
is a kozak sequence found within the mrna transcript
yes
is a stop codon found within the mrna transcript
yes
is a shine delgarno sequence found within a mrna transcript
yes
what is the sequence of mRNA for a polypeptide that encodes 4 Met residues
AUG AUG AUG AUG
what is the coding strand of DNA for the peptide 5’ AUG AUG AUG 3’
5’ ATG ATG ATG ATG 3’
what is the template strand of DNA for the polypeptide 5’ AUG AUG AUG AUG 3’
3’ TAC TAC TAC TAC 5’
what is the Met tRNA anticodon
5’ CAU 3’
what part of the mRNA sequence help E. coli distinguish the start codon from other met codons
shine-delgarno sequence (towards 5’ end of AUG)
what modification helps E. coli distinguish between an initiator tRNA and a regular tRNA
fMET
describe the wobble hypothesis
variance in base pairing is greater at 5’ end of anticodon, accounting for more than one codon
non W-C base pairing can be allowed in last position of the codon
imagine a mutation that makes gln aminoacyl synthetase recognize and bind to the tRNA^asn
would this mutation change the mRNA sequence
no
guanine nucleotide exchange factors are a set of proteins that activate GTPases by exchanging a GDP for a GTP
imagine a new eukaryotic protein that phosphorylated to a site on eIF2 that prevented eIF2 from binding its guanine nucleotide exchange factor
what would happen to the number of tRNAi^met
the number of charged initiator met would increase since they are not being used to start new protein synthesis; number of uncharged tRNAi^met would decrease
guanine nucleotide exchange factors are a set of proteins that activate GTPases by exchanging a GDP for a GTP
imagine a new eukaryotic protein that phosphorylated to a site on eIF2 that prevented eIF2 from binding its guanine nucleotide exchange factor
what would happen to the levels of protein translation
protein translation would decrease since they cant initiate new proteins
mRNA: 5’ ACC GCC AUG CAU CAC CAC CAU 3’
what is the coding strand of DNA
5’ ACC GCC ATG CAT CAC CAC CAT 3’
mRNA: 5’ ACC GCC AUG CAU CAC CAC CAU 3’
what is the AA sequence translated by the ribosome
met-his-his-his-his
mRNA: 5’ ACC GCC AUG CAU CAC CAC CAU 3’
what is the AA sequence once the peptide sequence has been processed into a functional protein
His His His His
mRNA: 5’ ACC GCC AUG CAU CAC CAC CAU 3’
what are the anticodons used to translate this mRNA
5' ---> 3' CAU (met) GUG (his) GUG (his) GUG (his) GUG (his)
insertions or deletions of one or two DNA nucleotides can cause what kind of mutations
frameshift mutations
where is the anticodon on the tRNA
bottom of the “t”
which end of the tRNA does the specific amino acid bind to
the 3’ end
T or F: the aminoacyl synthetase uses base pairing in the anticodon region to determine tRNA identity
false
what is aminoacyl synthetase
enzyme that attaches the right amino acid onto the tRNA
how does aminoacyl synthetase determine tRNA identity
the overall configuration of the tRNA
what is the role of IF1
prevents premature binding of the incoming tRNA to the A site
what is the role of IF2
loads in the fMET (initiator) tRNA into the 30s subunit
what is the role of IF3
prevents premature binding of the 50s subunit
what is the role of fMET
initiator tRNA that binds to the start codon
what is the role of the sine delgarno sequence
binds to the 16s rRNA to position start codon in the P-site
in EUKARYOTIC transcription initiation
what are the two roles of eIF4F in translation
bind mRNA and load it into small subunit
induce mRNA circularization
in EUKARYOTIC transcription initiation
which protein binds and then loads the initiator tRNA into the small subunit
eIF2
T or F: the initiator tRNA is loaded into the small subunit before the mRNA
true
if the genome annotation only used the AUG start codon to identify ORFs, but the leaderless start codon does not need to be AUG, what is suggested about the potential proteins in bacterium
there could be other proteins that do not use the AUG start codon that would not be recognized by the genome annotation algorithm
what are the two non-canonical mechanisms that give rise to the wobble hypothesis
U can basepair with G and A
I can basebair with C,U,A
in a BACTERIAL ribosome
what are the sizes of the subunits
large subunit: 50s
small subinit: 30s
in a BACTERIAL ribosome
which way does the mRNA run from L to R
5’ –> 3’
in a BACTERIAL ribosome
what order do the ribosomal tRNA sites go from L—>R
EPA
considering BACTERIAL translation initiation
T or F: EF-Tu-GTP bind to the 50s ribosome
false
considering BACTERIAL translation initiation
T or F: the 30s ribosome associated with the mRNA before the 50s ribosome
true
considering BACTERIAL translation initiation
T or F: IF-1, IF-2, IF-3 dissociate before the 50s subunit binding
true
considering BACTERIAL translation initiation
T or F: N-formylmethionyl-tRNS^fmet binds in the p site
true
consider the aspartic acid (asp) aminoacyl synthetase
what is the first step required to charge the tRNA
adenylating the AA (linking the free AA with an ATP)
consider the aspartic acid (asp) aminoacyl synthetase
what is the notation of the tRNA upon charging with the right AA
asp-tRNA^asp
consider the aspartic acid (asp) aminoacyl synthetase
imagine a mutation in the asp tRNA such that it is only recognized by the glu synthetase. what is the notation of the tRNA in this mutated charged context
glu-tRNA^asp
consider the synthetitc protein GCaMP (Ca binding domain) sensor
given that this protein is synthetic, why can it work inside cells
protein function is modular so the pieces can still do their roles when fused together in the cell
consider the synthetitc protein GCaMP (Ca binding domain) sensor
how could you make a new synthetic protein that has a fluorescent readout that senses Ca only near the ribosome
fuse the GCaMP proein onto a ribosomal protein that targets only to ribosomes
when considering ribosomal sizes, what does S stand for
Svedbergs
which statements describe transcription factors
- binds specific sequence of DNA
- binds only at promoter regions
- changes affinity for RNA pol
- changer affinity for DNA pol
- is correctly positioned on DNA by anticodon base-pairing
- and 3. are correct
which statements correctly describe co-activators
- binds specific sequence of DNA
- binds only at promoter regions
- changes affinity for RNA pol
- changes affinity for DNA pol
- is correctly positioned on the DNA by anticodon basepairing
- is correct
what type of protein secondary structure in transcription factors can read out the identity of nucleotides
alpha-helix
which region of the base (A,T,G,C) is used by transcription factors to chemically distinguish the identity of the nucleotides
major groove
T or F: LacI disrupts W-C basepairing when binding to the operator
false
what is the signal recognition particle
cytoplasmic protein complex
what is the function of the signal recognition particle
bind the N-terminal end of growing polypeptides for membrane translocation
imagine a mutation in the lac operator such that lacI is unable to bind
which of the following would occur
1. in the presence of lactose, no mRNA would be produced
2. in the presence of lactose, some mRNA would be produced
3. in the absence of lactose, no mRNA would be produced
4. in the absence of lactose, some mRNA would be produced
- and 4. are correct
a mutation in a eukaryotic gene induces an extra 80 AA in the translated product
what type of mRNA likely occurred
non-stop mRNA
a mutation in a eukaryotic gene induces an extra 80 AA in the translated product
what is the most likely identity of the last AA
lysine
imagine a mutation in BACTERIA such that the ribosome reaches the end of the mRNA without recruiting RF1 or RF2
what is the name of the structural mimic that can rescue this type of mutation
tmRNA
imagine a mutation in BACTERIA such that the ribosome reaches the end of the mRNA without recruiting RF1 or RF2
briefly describe how the structural mimic tmRNA is able to salvage the ribosome
the tRNA mimic portion loads into the A-site and catalyzes the peptide transfer to an alanine. the other mRNA like pieces of the tmRNA are then read by charged tRNAs as per usual
T or F: enhancers only increase transcriptional activity of genes that are located thousands of basepairs away in the genome
false
imagine a mutation in the bacterial L4 protein involved in regulation of ribosomal protein production. this mutation renders the L4 protein unable to bind to the mRNA
what would occur??
the amount of ribosomal proteins would increase
of the 7 points of regulation of protein expression, which is most heavily regulated by the cell
transcription initiation
why is transcription initiation most heavily regulated by the cell
so that no energy is wasted downstream in mRNA or protein production
what are 4 different mechanisms that are used by cells to alter protein activity post-translationally
phosphorylation, dimerization, translocation, ubiquitination
cystic fibrosis is caused by defects in the CFTR protein (uses ATP to move Cl ions across plasma membrane) and if it doesnt work fluid builds up, resulting in disease.
how could a mutation in the first 20 AA induce disease if it does not affect Cl pumping ability
the mutation could prevent trafficking to the plasma membrane
which of the following statements are true
- every cell in the human body has the same DNA sequence
- every cell in the human body has the same histone mark patterns
- every cell in the human body transcribed the same mRNAs
- every cell in the human body translates the same proteins
- is true
imagine a mutation in yeast that prevents binding of Gal4p from the gal promoter sequence
how would this mutation affect the expression of the GAL genes
they would never be expressed because the activator could never recruit the RNA polymerase to that location
how would a mutation that prevents expression of recA affect the expression of the SOS genes in E. coli
it would prevent SOS gene expression in the presence of widespread damage
what are all the possible codons for Phe
5’ UUU 3’
5’ UUC 3’
what are all the possible anticodons for Phe
5’ AAA 3’
5’ GAA 3’
for an organism that only has one tRNA for Phe, what would the anticodon likely be
5’ GAA 3’
what modification on which AA helps E. coli distinguish between an initiator tRNA and a regular tRNA
formyl group on methionine
design a forward primer for the ORF with 6 nucleotides and a PstI cutsite
DNA sequence:
5’ CAACCTAATGCACCCCGATTCCATAAAGTAAAACCTTCAACTC 3’
5’ CTGCAG ATG CAC 3’
design a reverse primer for the ORF with 6 nucleotides and a EcoRV cutsite
DNA sequence:
5’ CAACCTAATGCACCCCGATTCCATAAAGTAAAACCTTCAACTC 3’
3’ AAG TAA GATATC 3’
(want in 5’ –> 3’ direction)
5’ GATATC TTA CTT 3’
