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Translation. Protein Synthesis: Ch 17 From : Kevin Brown – University of Florida. Protein Synthesis. Translation from mRNA Codons to amino acids http ://www.youtube.com/watch?v=suN- sV0cT6c. mRNA transcript organization. 1) Open reading frame: codes for protein
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Translation Protein Synthesis: Ch 17 From : Kevin Brown – University of Florida
Protein Synthesis • Translation from mRNA • Codons to amino acids http://www.youtube.com/watch?v=suN-sV0cT6c
mRNA transcript organization • 1) Open reading frame: codes for protein • 5’end has translational start site, 3’ end has translational stop site • 2) 5’ UTR – untranslated region (upstream of start site) • 3) 3’ UTR – untranslated region (downstream of stop site)
Codons • Codons are read from 5’ – 3’ in triplets • Genetic code • 61 codons for amino acids, 20 amino acids • Amino acid sequence is determined by the complementary base pairing between mRNA codons and tRNA anticodons
Translation – tRNA molecule • tRNA – Transfer RNA – transfers amino acids from cytoplasm to ribosome, where the amino acids get added onto a growing polypeptide chain. • Anticodon – triplet code found on tRNA, that is complementary to a specific mRNA codon. • Made of a single RNA strand that folds back on itself to form a 3 D structure – L-shaped.
tRNA • 64 codons • 45 tRNA • Why? • Last base on tRNA anticodon is flexible – so some tRNA anticodons can recognize more than one codon
Wobble Non standard base pairing at 3’ end of codon – flexibility Many tRNAs have inosine base at 5’ end of anticodon- Inosine can pair with A, U, or G
tRNA bind to specific amino acid Aminoacyl-tRNAsynthetase (enzyme) • AminoacyltRNAsynthetase joins a particular amino acid to a tRNA through covalent bonding • There are 20 of these enzymes Amino acid P Adenosine P P P Adenosine P P i ATP P i P i tRNA P Adenosine AMP Aminoacyl tRNA(“charged tRNA”)
Start signals • Translational start signal • 5’ AUG 3’ , which codes for Methionine • Which AUG is the start signal? • Bacteria – often have Shine-Delgarno sequence upstream from AUG
Eukaryotes – no Shine Delagarno, but a loose sequence – Kozak sequence
rRNA - ribosomal RNA • Ribosome is made up of rRNA & proteins • Ribosome – the site of protein synthesis • Eukaryotes: Large subunit – 60 S, Small subunit – 40 S • Bacteria: Large subunit – 70S , Small subunit – 30 S
A site (Aminoacyl-tRNA binding site) Ribosome Schematic P site (Peptidyl-tRNAbinding site) Exit tunnel E site (Exit site) E A P Largesubunit mRNAbinding site Smallsubunit
Protein Synthesis - Initiation Largeribosomalsubunit U 3 5 C A P site Met Met 3 5 A G U P i InitiatortRNA GTP GDP E A mRNA 5 5 3 3 Start codon Translation initiation complex mRNA binding site
Growing polypeptide Amino end Next aminoacid to beadded topolypeptidechain E tRNA mRNA 3 Codons 5
Amino end ofpolypeptide Protein synthesis - Elongation E 3 mRNA Ribosome ready fornext aminoacyl tRNA Asite Psite 1) Codon recognition 5 GTP GDP P i E E P A A P GDP P i 3) Translocation GTP 2) Peptide bond formation E A P
P Protein synthesis - Termination Releasefactor Freepolypeptide 5 3 3 3 GTP 2 5 5 2 GDP 2 Stop codon i (UAG, UAA, or UGA)
Translation animation • http://www.youtube.com/watch?v=Ikq9AcBcohA
Energy usage during Translation • tRNA Charging – ATP to AMP + PPi for each amino acid covalently bonded to tRNA • Initiation – 1 ATP to ADP + Pi and 1 GTP to GDP + Pi hydrolyzed during formation of initiation complex • Elongation – 2 GTP hydrolyzed per amino acid • 1 for codon recognition • 1 for energy for translocation of tRNA from A to P • Termination – 1 GTP to GDP + Pi hydrolyzed in termination
Completing the protein • Folding (may be helped w/chaperone protein) • Post-translational modifications: • Chemical modifications – add groups • Remove amino acids from leading end of polypeptide chain • Cleavage of protein
Some polypeptides get targeted to specific locations • All ribosomes start free • If polypeptide gives signal, ribosome binds to ER. • Signal peptide is at leading (NH3) end of polypeptide • Signal recognition particle (SRP) – recognized signal peptide and brings into receptor protein in ER • Protein either is in ER lumen, or may end up as part of membrane
Mutations • Changes in genetic information of cell (or virus) • Small scale – few nucleotides: • Substitution, insertion, deletion • Frameshift are most damaging • Large scale – chromosomal changes
Translation • http://www.dnalc.org/view/15501-Translation-RNA-to-protein-3D-animation-with-basic-narration.html