Lectures 19 & 19: Protein Synthesis Flashcards
How many possible start codons are there?
1
How many possible stop codons are there?
3
What is the polarity of tln?
5’ –> 3’
What is a silent mutation?
Does not affect AA produced
What are the components of tRNA? What do they do?
3’ end - binds AA which will be added
anticodon loop - associates with mRNA
then also D loop, T loop, 5’ end
What is the wobble hypothesis?
first two bases of codon-anticodon interaction are constrained by normal Watson-Crick base-pairing
Requirements for H-bonding at third base are less stringent
What enzyme catalyzes the transfer of a specific AA to the 3’-OH of specific tRNAs (charges tRNA)?
aminoacyl-tRNA synthetases
What is the overall aminoacyl-tRNA synthetase rxn?
ATP + AA + tRNA –> aminoacyl tRNA* –> AMP + PPi
What are the two steps of the rxn which charges tRNA?
- activation of AA by rxn w/ ATP to form aminoacyl adenylate
- rxn of activated AA with 3’-OH of tRNA to form aminoacyl-tRNA
How is uncharged vs charged tRNA with Ser written?
uncharged = tRNAser cahrged = ser-tRNAser
What are the subunits of bacterial ribosomes?
50S + 30S
70S
What are the subunits of eukaryotic ribosomes?
60S + 40S
80S
How many tRNA molecules can one ribosome bind?
three
e, p, a
what are the three tRNA binding sites on ribosome called?
E = exit P = peptidyl A = aminoacyl
What happens at the A (aminoacyl) tRNA binding site of a ribosome?
attachment of incoming aminoacyl-tRNA
What happens at the P (Peptidyl) tRNA binding site of a ribosome?
attachment of pepidyl tRNA
What happens at the E (exit) tRNA binding site of a ribosome?
harbors spent tRNA
releases it
What happens during tln initiation?
- mRNA binds, aligned in correct reading frame
- initiator aminoacyl tRNA binds
- ribosome assembles from small and large subunits
What happens during tln elongation?
- aminoacyl tRNA binds, checks codon-anticodon match
- new peptide bond is formed
- growing chain is translocated from A site to P site
- mRNA is pulled along - next codon exposed to A site
What happens during tln termination?
- release factors bind GTP - bind stop codon in A site
- peptidyl tRNA in P site is hydrolyzed - release peptide chain, leave tRNA in P site
- GTP hydrolysis –> release tRNA, release factors, mRNA; ribosome dissociates
How does ribosome know which Met codon is the start codon in prokaryotes?
Shine-Dalgarno sequence upstream
How does ribosome know which Met codon is the start codon in eukaryotes?
5’ cap (7-methyl guanosine residue)
Is mRNA linear or circular during translation in euk and prok?
prok - linear
euk - circular (poly-A tail interacts with 5’ cap)
which kingdom has polycistronic vs monocystronic tln?
prok = poly euk = mono
What does Polycistronic mean?
once ribosome attaches to Shine-Dalgarno sequence, multiple proteins can be made from same mRNA strand
What does monocistronic mean?
single protein is translated from single mRNA
what are initiation factors?
proteins which help assemble and disassemble transient complexes during tln
-not permanent parts of ribosome
What are the prok initiation factors?
IF1, IF2, IF3
How many initiation factors do euk have? Which is similar to one of the prok IFs?
> 10
eIF-2 ~ IF2
What is the role of GTP in tln?
catalyze
conf change in ribosomes to accelerate rxns
What is the role of GTP hydrolysis in tln?
Makes rxns irreversible
What do the prok elongation factors do?
EF-Tu brings each aatRNA in, checks match
EF-G helps shift mRNA and tRNA
How many release factors do prok have?
Euk?
prok - 3 (RF 1,2,3)
euk - 1 (eRF)
How many elongation factors do prok have? Euk?
prok - 3 (EF 1,2,3)
euk - 2 (eEf 1,2)
Why doesn’t peptide elongation require nrg?
aa-tRNA is charged
What do the two active sites of amino acyl sythetases do?
one recognizes correct tRNA and attaches AA
Other proofreads - recognizes and removes incorrectly attached AA
Which steps of tln require energy?
synthesis of aa-tRNA initiation elongation release polypeptide proofreading
what does phosphorylation of eIF-2 do?
decreases rate of protein synthesis
What does eIF-2 do?
Carries GTP and initiator tRNA to ribosome
How does the regulation of globin synthesis in response to heme availability exemplify tln regulation?
- heme is absent –> eIF2 is phosphorylated by HCI = inactive
- heme is present –> phosphate is removed, tln occurs
How is the 5’-cap involved in regulating euk tln?
- protects from nucleases
- distinguishes btw mRNA and types of tRNA
- eIFs recognize cap
How is the 5’-UTR involved in regulating euk tln?
sequences for translational efficiency
How is the 3’-UTR involved in regulating euk tln?
signal sequences so mRNA is translated in specific places in cell
How is the poly-A tail involved in regulating euk tln?
- stabilizes - important for lifespan of mRNA
- may catalyze formation of large ribosomal subunit
What feature of mRNA plays a huge role in tln regulation?
lifetime
what is a key regulatory molecule in tln of the ferritin/transferrin receptor?
aconitase
In what two ways does aconitase affect tln of the ferritin/transferrin receptor?
binds IRE 5’-UTR of ferritin mRNA –> blocks tln initiation
binds IRE 3’-UTR of ferritin mRNA –> stabilizes mRNA against degradation
what is the function of ferritin? What happens to the tln of ferritin if [iron] increases?
intracellular iron storage protein
increase [iron] increase tln of ferritin
what is the function of the transferrin receptor? What happens to the tln of transferrin receptor if [iron] increases?
imports iron
increase [iron] decrease tln of transferrin receptor
What is endogenous regulation of proteins synthesis?
siRNA pr miRNA can down-regulate tln by inducing mRNA degradation
What is exogenous regulation of protein synthesis?
dsRNA gets processed to siRNA –> degradation of mRNA
In what two ways can interferons work?
-inhibition of translation (makes protein kinase - phosphorylation of eEf-2) mRNA degradation (produces endoribonuclease --> mRNA non-functional)
How is an interferon produced?
- virus produces dsRNA = signal to host that there is a viral infection
- host cells secrete interferons (IFNs)
- IFNs bind to other cells, induce expression of enzymes which will interfere with protein synthesis
