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Protein Synthesis Biology 12 Mr. McIntyre

Protein Synthesis Biology 12 Mr. McIntyre. Translation: From messenger RNA to protein:. The information encoded in the DNA is transferred to messenger RNA and then decoded by the ribosome to produce proteins. 5’-ATGCCTAGGTACCTATGA-3’ 3’-TACGGATCCATGGATACT-5’. DNA. Transcription.

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Protein Synthesis Biology 12 Mr. McIntyre

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  1. Protein Synthesis Biology 12 Mr. McIntyre

  2. Translation: From messenger RNA to protein: The information encoded in the DNA is transferred to messenger RNA and then decoded by the ribosome to produce proteins.

  3. 5’-ATGCCTAGGTACCTATGA-3’ 3’-TACGGATCCATGGATACT-5’ DNA Transcription mRNA 5’-AUGCCUAGGUACCUAUGA-3’ decoded as 5’-AUG CCU AGG UAC CUA UGA-3’ Translation Protein N-MET-PRO-ARG-TYR-LEU-C

  4. Alanine tRNA

  5. Generalized tRNA

  6. Modified Bases Found in tRNAs = UH2

  7. tRNAs are activated by amino-acyl tRNA synthetases

  8. Structure of an amino acyl-tRNA synthetase bound to a tRNA

  9. One mechanism for maintaining high fidelity of protein synthesis is the high fidelity of aa-tRNA synthetases

  10. Amino-acyl tRNA synthetases: One synthetase for each amino acid a single synthetase may recognize multiple tRNAs for the same amino acid Two classes of synthetase. Different 3-dimensional structures Differ in which side of the tRNA they recognize and how they bind ATP Class I - monomeric, acylates the 2’OH on the terminal ribose Arg, Cys , Gln, Glu, Ile, Leu, Met, Trp Tyr, Val Class II - dimeric, acylate the 3’OH on the terminal ribose Ala, Asn, Asp, Gly, His, Lys, Phe, Ser, Pro, Thr

  11. Two levels of control to ensure that the proper amino acid is incorporated into protein: 1) Charging of the proper tRNA

  12. 2) Matching the cognate tRNA to the messenger RNA

  13. Incorporation of amino acids into polypeptide chains I

  14. Incorporation of amino acids into polypeptide chains II

  15. Protein synthesis occurs on ribosomes

  16. Protein synthesis occurs on ribosomes

  17. and mitochondria

  18. Ribosome Assembly The proteins of each ribosomal subunit are organized around rRNA molecules 16S rRNA

  19. Ribosome Assembly: takes place largely in a specialized domain of the nucleus, the nucleolus

  20. In the nucleolus, RNA polymerase I transcribes the rDNA repeats to produce a 45S RNA precursor The 45S precursor is processed and cleaved into mature rRNAs and ribosomal proteins then bind to generate the large and small ribosomal subunits

  21. 23S rRNA secondary structure

  22. 3D organization of the eukaryotic large subunit rRNA

  23. Ribosomal Proteins decorate the surface of the ribosome Large subunit. Grey = rRNA Gold = ribosomal proteins

  24. Ribosomal proteins often have extensions that snake into the core of the rRNA structure Crystal structure of L19 L15 (yellow) positioned in a fragment of the rRNA (red)

  25. The ribosomal proteins are important for maintaining the stability and integrity of the ribosome, but NOT for catalysis ie. the ribosomal RNA acts as a ribozyme

  26. The large and small subunits come together to form the ribosome Mitochondrial or Prokaryotic Eukaryotic 60S subunit 80S ribosome 40S subunit

  27. The association of the large and small subunits creates the structural features on the ribosome that are essential for protein synthesis Three tRNA binding sites: A site = amino-acyl tRNA binding site P site = peptidyl-tRNA binding site E site = exit site

  28. In addition to the APE sites there is an mRNA binding groove that holds onto the message being translated

  29. There is a tunnel through the large subunit that allows the growing polypeptide chain to pass out of the ribosome

  30. Peptide bond formation is catalyzed by the large subunit rRNA

  31. Peptide bond formation is catalyzed by the large subunit rRNA

  32. Incorporation of the correct amino acyl-tRNA is determined by base-pairing interactions between the anticodon of the tRNA and the messenger RNA

  33. =EF-1 Proper reading of the anticodon is the second important quality control step ensuring accurate protein synthesis Elongation factors Introduce a two-step “Kinetic proofreading”

  34. A second elongation factor EF-G or EF-2, drives the translocation of the ribosome along the mRNA Together GTP hydrolysis by EF-1 and EF-2 help drive protein synthesis forward

  35. Termination of translation is triggered by stop codons Release factor enters the A site and triggers hydrolysis the peptidyl-tRNA bond leading to release of the protein.

  36. Release of the protein causes the disassociation of the ribosome into its constituent subunits.

  37. Release Factor is a molecular mimic of a tRNA eRF1 tRNA

  38. Initiation of Translation Initiation is controlled differently in prokaryotic and eukaryotic ribosomes In prokaryotes a single transcript can give rise to multiple proteins

  39. In prokaryotes, specific sequences in the mRNA around the AUG codon, called Shine-Delgarno sequences, are recognized by an intiation complex consisting of a Met amino-acyl tRNA, Initiation Factors (IFs) and the small ribosomal subunit

  40. GTP hydrolysis by IF2 coincident with release of the IFs and binding of the large ribosomal subunit leads to formation of a complete ribosome,on the mRNA and ready to translate.

  41. Eukaryotic mRNAs have a distinct structure at the 5’ end

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