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From Gene to Protein

From Gene to Protein . Gene Expression. Process by which DNA directs the synthesis of a protein 2 stages transcription translation All organisms One gene  one protein. 1. Transcription of DNA. Gene Composed of DNA 1000s of base pairs long Sequence of G, A, T, C is a code.

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From Gene to Protein

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  1. From Gene to Protein

  2. Gene Expression • Process by which DNA directs the synthesis of a protein • 2 stages transcription translation • All organisms One gene  one protein

  3. 1. Transcription of DNA • Gene • Composed of DNA • 1000s of base pairs long • Sequence of G, A, T, C is a code ACATACATGTAGCAACGAAACCTGAACATGCGAACATTGACACAACAACAAUGUAGCCA A portion of the insulin gene

  4. A gene is transcribed into mRNA • One DNA strand is a template • Follow base complementation using G, A, U, C Which strand is the template strand?

  5. • This mRNA is referred to as a “transcript” • Single stranded • Uracil nucleotide (not thymine) • ONLY gene of interest is transcribed • mRNA will leave the nucleus for translation step

  6. mRNA (transcript) leaves nucleus • Translated to protein in cytoplasm • What organelle translates mRNA to protein? • How many amino acids? • How many amino acids long is a protein? • How could 10 identical protein molecules be translated from 1 mRNA? • What happens to mRNA when the cell has finished with it? • Can the cell transcribe more than one gene at a time?

  7. 2. Translation of mRNA • Nucleotides (G, A, U, C) translated to amino acids • Ribosomes assemble amino acids to form polypeptide • What is the difference between a polypeptide and a protein? • What is the length difference between a chromosome and an mRNA transcript? • Why do researchers use frog embryos to study translation?

  8. Differences between prokaryotes and eukaryotes • Proks • No nucleus • Translation can begin while transcription is still in progress

  9. Euks • Nuclear envelope • Transcript is modified before leaving nucleus • Primary transcript mRNA  mRNA • Translation in cytoplasm

  10. Central Dogma DNA  mRNA  protein (F. Crick 1956)

  11. The genetic code Triplet code 43 = • # combinations of 3 using G, A, T, C • Can code for all 20 aa • Each triplet codes for a certain amino acid

  12. Codon table (1960’s Nirenberg and Khorana) • 3 stop codons • 1 start codon • Redundant • Nearly universal

  13. Which strand of DNA is the template? Identify the codons (triplets) Code is non-overlapping Compare the mRNA to the non-template strand How many codons are needed for a 20 amino acid long protein? How many nucleotides?

  14. The reading frame AUG UGG GUU GGC UCA Amino acids? met –trp- phe-gly-ser

  15. Transcription in more detail • Initiation of transcription A. RNA polymerase binds to template strand of DNA  upstream ----------------------------------------------downstream 

  16. The promoter is upstream from the start site for transcription

  17. B. Transcription factors (proteins) are required for RNA pol to bind

  18. C. mRNA begins to be synthesized • (RNA pol does not need a primer)

  19. 2. Elongation of transcript • RNA pol synthesizes 5’  3’ • More than one transcript made

  20. 3. Termination RNA pol detaches

  21. Translation summary

  22. Translation in more detail •  Cytoplasm • The players: • Transfer RNA tRNA • Ribosomes • mRNA • Amino acids

  23. 1. tRNA 2o structure is a cloverleaf CONCEPT CHECK How many bases is a tRNA? How many bases is an anticodon? Where is the amino acid site with respect to the anticodon? Why does the tRNA look to be double stranded? What does the “t” mean in tRNA?

  24. A charged tRNA carries an amino acid • Contains an anticodon • Complementary to a codon Note the CCA

  25. 2. Ribosome • rRNA + protein • Large and small subunits

  26. 3 binding sites on ribosome • P site holds tRNA • A site for tRNA with next amino acid • E site allows tRNA to leave • Note: large and small • subunits

  27. Ribosomes adds each aa from tRNA to growing polypeptide chain up to 100,000s per cell Activity overview of protein synthesis

  28. Stages of translation • Initiation • Ribosome scans mRNA for AUG start • tRNA brings met

  29. 2. Elongation Amino acids added to chain Peptide bonds between amino acids

  30. 3. Termination stop codon Polypeptide released

  31. Polyribosomes • One mRNA translated by many ribosomes

  32. Wobble hypothesis • How many sense codons? • Stop codons? • BUT……………only 45 tRNAs • Some tRNAs recognize more than one codon

  33. 3rd position in mRNA codon “wobbles” Us and Cs can be matched with G in anticodon Rules of base pairing relaxed in the 3rd position

  34. After translation  ER  Golgi Post – translational modification

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