Translation Flashcards
whats a codon. How many?
triplet of DNA that encodes a specific aa. 64. 3 dont encode a specific aa. 20 aa encoded in our genetic code.
Some aa have more than one codon which is referred to as _____
degeneracy; doesnt mean its inaccurate
Codons DO NOT
overlap; no spaces bet them. Have to be in frame in order to produce protein with right aa sequence.
What are the stop codons?
UAA, UGA, UAG
Whats AUG, UUU, CCC, GGG, AAA,
AUG; Met: start codon UUU; Phe CCC; Pro GGG; Gly AAA; Lys
whats the ORF
open reading frame; the inframe seq bet the initiation codon and the termination codon thats building the protein.
What does 5’ end of mRNA correspond to? What about 3’ end?
N terminal of the protein. 3’ end of mRNA corresponds to the C terminal of the protein.
Protein synthesis begins at the amino terminal end of the nascent proteins.How did they figure out which way proteins get synthesized?
Used radioactive leucine in reticulocytes (immature RBC) and followed globin synthesis. THey would add in leucine and extract the protein at diff time points after they added radio label and see what part of newly made proteins was being labeled. Early carboxy terminus was being labeled which means amino terminus was already being made. The longer they left label in thre the more of protein was being radio labeled.
RNA cannot interact directly with __ ___ to specify their order
amino acids; tRNA does this via tRNA amino acyl synthases; serve as adaptor molecules specific for each aa that bind to specific codons on RNA templates.
Replication has the most proofreading properties associated with it bc:
only have 1 DNA genome which gets transferred to daughter cells. Want it to stay constant
what are features of tRNA
- at least 32 tRNA
- all diff for 20 aa
- some can recognize more than 1 codon. (wobble rules of genetic code)
- mitochondria has own set of tRNAs
- Always have G at 5’ end
- Always have ACC at 3’ end
- 8 or more bases chemically modiefied
In tRNA, the D arm contains_____. The T psudo C arm contains the ___,___,____. The anticodon arm contains the _____. The amino arm contains the ____ and binds the aa.
dihydrouridine; ribothymidine, psuedouridine, and cytosine.
Anticodon.
CCA
The anticodon bp’s with the ____ in the mRNA. The tRNA and the mRNA bp ____ to each other. The first base of the codon is paired with the ____ base of the ___
codon; antiparralel; third; anticodon.
Whats the wobble hypothesis
- First two bases of a codon always form strong strandard (W-C) bp’s with the corresponding bases of tRNA & confer most of the specificity.
- The first base of some anticodons det the number of codons read by a given tRNA.
- When an aa is specified by sev diff codons, those codons that differ in either of the first two bases req diff tRNAs
- a min of 32 diff tRNAs are req to translate all 61 codons
If there is a U it can bind to ___ or G. If there is a G it can bind with C or ___IF there is an I (which means base is hypoxanthine) it can bind to ____ ____ or C
A. U
A, U
What are the 3 sites on ribosome
- Peptidyl site
- Aminoacyl (A) site
- Exit (E) site
Diff in eukaryotic and prokaryotic ribosome is in their ____ value. But their S values do not add up!
sedimentation and density
what are the 5 stages of protein synthesis
- Activation of aa
- initiation
- elongationn
- termination and ribosome recycling
- folding and postranslational processing
what is stage 1. what catalyzes this reaction?
AA are attached via ester bonds to the 3’ OH of the corresponding tRNA; this process is called CHARGING the tRNA via amino acyl-tRNA synthethase. This step is energy needy and is irreversible!
what is the enzyme bound intermediate formed by reaction of ATP and aa? what gets transferred to corresponding specific tRNA?
aminoacyl adenylate (aminoacyl AMP); aminoacyl group
how do aminoacyl tRNA synthestases recognize correct tRNAs?
- some rec. anticodon itself
- some rec. 10/more bases in tRNA
- some rec one specific base pair!
- structural features seem to be impt
What does the aminoactyl tRNA form a complex with?
molecule called EF-Tu (elongation factor) that binds one GTP molecule
what is stage 2?
Initiation: all organisms have two tRNAs for methionine (initiation and within polypeptide chain)
What is the starting aa in bacteria
N - formylmethionine
In bacteria, initiation req 7 components:
- 30s ribosomal subunit
- mRNA
- initiating fMet tRNA
- 3 initiation factors: IF-1, 2, &3
- GTP
- 50s ribosomal subunit
- Mg++
What does the Shine Dalgarno seq in bacteria do?
Places P site of ribosome complex right over the AUG of the start site.
What do IF-1 and 3 in bacteria do?
IF-1 binds and blocks the A site. IF-3 helps with positioning 30s ribosome on Shine Dalgarno seq.
what is step 2 of initiation in bacteria
30s ribosomal subunit, mRNA, IF-1, and 3 is joined by IF-2 and fMet-tRNA. the tRNA binds to P site, NOT a site.
what is step 3 of initiation in bacteria
30s subunit binds to 50s ribosomal unit. GTP bound by IF-2 gets hydrolyzed to GDP. The other IF leave. The complete ribosome has charged initiatior tRNA in complete P site and an empty A site. This complex is officially called the initiation complex.
In eukaryotes there are ___ initiation factors. We have a ____sequence rec by 40s subunit. Complete ___ribosome assembled using the other IFs and initiating Met-tRNA
- One binds to 5’ cap of mRNA and complex formed with 40s subunit. Kozak; has AUG embedded in it. 80s
what is stage 3 of protein synthesis for eukaryotes? how does it work?
Elongation: stepwise addition of aa to polypeptide chain by formation of peptide bonds. In both pro and eukaryotes there are 10-100 ribosomes found bound to each mRNA molecule.
In bacteria elongation requires 3 steps:
- Initiation complex (for first peptide bond)
- Next aminoacyl -tRNA specified by next codon
- Elongation factors EF-Tu, EF-Ts, EF-G
what is the enzyme that catalyzed peptide bond formation between the aa’s
peptidyl transferase; this enzyme is a ribozyme (23s rRNA) which gets its energy by the high energy ester bond bet aa and tRNA
In elongation stage in bacteria the initiating fMet is transferred from its ____ to the amino group of the second aa in the ____ site. A dipeptidyl tRNA is now base paired to the second codon in the ___ sit of the 30s subunit. The spent initiation tRNA is base paired to the AUG in the P site on the 30s subunit
tRNA; A; A;
Once peptide bond forms it changes conformation with the ___s but not the 30s. The deacylated inititiation tRNA moves into the E site of 50s and new dipeptidyl trNA moves to P site on 50s.
50
In translocation of elongation, the ribosome moves along the mRNA one codon towards the 3’ end. Movement is mediated by elongation factor___. Ribsome movement shifts the ____tRNA from the ___ site on the 30s subunit to the P site. The spent tRNA shifts from the ___ site on the 30s to the E site. The ___ site is now empty and ready for next aa -tRNA
EF-G. GTP. dipeptidyl
A. P. A.
T/F A and E sites cannot be occupied simultaneously.
True
Why use two steps to move tRNA?
Prevents premature release of peptidyl tRNA and enhances processivity.
Bacteria have ___releasing factors to terminate protein synthesis. RF 1 is for ___ and UAA. RF 2 is for ___ and UAA. No one know what the third one is for. They induce peptidyl transferase to transfer _____ to water resulting in release of peptide chain from ribosome. In eukaryotes there is only ___ releasing factor
3; UAG; UGA; peptide chain. 1
Transcription and translation are tightly coupled in bacteria. This allows protein synthesis to take place before ___ is completed and degraded
mRNA
Protein synthesis is expensive. There are at least __ high energy bonds expended per peptide bond. Energy is used for ____. Used to maintain proper alignment of codon, tRNA and growing end of polypeptide chain
- specificity
Overall error rate for protein synthesis is:
1:10,000 aa
what are inhibitors of translation
antibiotics and toxins:
- tetracycline; bind procaryote A site; block aa tRNa binding which is useful for treating periodontal disease
- chloramphenicol: blocks peptidyl transfer in prokaryotes and mitochondria; useful for treating rickettsial infections
- streptomycin: binds s12 of 30s ribosome.
- aminoglycosides: block 50s ribosome binding to initiation complex
- cycloheximide: blocks eukaryote 80s peptidyl transferase
- diptheria toxin: catalyzes ADP-ribosylation of eEF2, inactivating it
- ricin: extremely toxic protein from castor bean; inactivates eukaryote 60s subunit by depurinating A residue of 28S rRNA.
