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Protein Synthesis

Protein Synthesis. DNA at work. If DNA = recipe book Proteins = courses of a meal. Recipes for all polypeptides are encoded by DNA mRNA is a copy of that recipe (DNA sequence) mRNA (recipes) travel to ribosomes for translation into polypeptides (proteins). Early developments.

dorothy-harding
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Protein Synthesis

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  1. Protein Synthesis DNA at work

  2. If DNA = recipe bookProteins = courses of a meal • Recipes for all polypeptides are encoded by DNA • mRNA is a copy of that recipe (DNA sequence) • mRNA (recipes) travel to ribosomes for translation into polypeptides (proteins)

  3. Early developments • 1909: A. Garrod suggests that “genes” create phenotypes via enzymes • Genes: heritable units of DNA • Phenotype: observable characteristic • People who lack particular enzymes have disease phenotypes (metabolic incompetence)

  4. Early developments • 1940’s:Beadle & Tatum; Neurospora crassa (mold) produce thousands of offspring; some cannot grow on traditional food source = nutritional mutants • Could these mutants lack an enzyme?

  5. Early developments • They do! • It’s often one dysfunctional enzyme per mutant, and one dysfunctional gene • One gene-one enzyme hypothesis • One gene-one protein • One protein-one polypeptide

  6. Protein recipe is written in genetic code (genes) • Genes lie along DNA • What are chromosomes? • Genes are linear sequences of nucleotides • One, three-nucleotide sequence = codon

  7. Genetic code & codons • Each codon codes for a particular Amino Acid • Each gene has many codons in it • Codons also exist for “start translating” and “stop translating”

  8. Genetic code & codons • Redundant – multiple codons specify same AA • Unambiguous - NO codon specifies more than one AA • Ancient – ALL organisms have same genetic code • AUG = Methionine whether you’re a redwood or a fruitfly

  9. How RNA is made • RNA polymerase adds RNA nucleotides to DNA template • RNA molecule peels away from DNA strand

  10. How RNA is made • Initiation: RNA polymerase binds to a promoter(specific nucleotide sequence) • Elongation: Polymerase adds complementary nucleotides to DNA template; RNA peels away, DNA reconnects

  11. How RNA is made • Termination: RNA polymerase reaches “terminator sequence”. • RNA polymerase detaches; mRNA detaches

  12. Further processing • Addition of caps (G) & tails (poly A) by RNA polymerase • Allow recognition by ribosomes (Cap, Tail) • Protect RNA from RNase attack (Cap) • Protect RNA from exonuclease attack (Tail) • Allow export by transporter molecules

  13. Further processing • Introns spliced out • Intervening sequences; NOT transcribed into polypeptide • Exons joined • Coding regions of DNA that are transcribed into Amino Acids

  14. tRNA brings appropriate AA • tRNA is “cook’s helper” • Brings individual ingredients (AA) to make the recipe (protein) • Binds appropriate AA (in cytoplasm) • Recognizes the mRNA codon that specifies its AA • Complementary nucleotide sequence (Anticodon) for recognition

  15. tRNA binding sites • Anticodons & AA attachment sites are themselves a string of three nucleotides • One enzyme attaches each AA to any of its possible tRNA transporters

  16. Ribosomes & Translation • rRNA plus proteins • 2 rRNA subunits • Bind mRNA • Bind tRNA with attached Amino Acids

  17. Ribosomes • Small subunit binds mRNA • Large subunit, with tRNA binding sites, attaches to small subunit + mRNA

  18. Translation • Initiation • mRNA binds to small subunit. • Initiator tRNA binds to start codon,always AUG -> first AA of all polypeptides is always Met

  19. Translation* • Elongation • Large subunit binds to small -> functional ribosome • Initiator tRNA attaches to P site of ribosome. Holds growing polypeptide. Next tRNA attaches to A site

  20. Translation • Elongation • Codon recognition: tRNA anticodon binds to mRNA codon in the A site • Peptide bond formation: Polypeptide detaches from tRNA in P site & binds to AA & tRNA in A site

  21. Translation • Elongation • Translocation: tRNA in P site detaches, A site tRNA & mRNA move, as unit, into P site. New tRNA attaches to A site. • Termination • Stopcodon is reached; no AA is added; polypeptide releases & subunits dissociate

  22. DNA – RNA - Protein • Gene expression

  23. Mutations • Any change in nucleotide sequence • Substitutions • Insertions • Deletions • Many alternative phenotypes result from single nucleotide changes

  24. Point Mutations • Substitution: • A single base pair is changed. • Synonymous (silent):results in NO AA change…why not? • Nonsynonymous: results in single AA change • These are less likely to be deleterious. WHY?

  25. Example* • Hemoglobin mutations • HbE: Codon position 26; Replace GLU w/ LYS; reduced Hb production. Hemoglobin instability at low O2 • HbC: Position 6; Replace GLU w/ LYS; RBC’s become rigid & crystalize • HbS: Position 6; Replace GLU w/ VAL; At low O2, Hb polymerizes & RBC’s collapse

  26. Point Mutations • Indels: insertions/ deletions • A single nucleotide is inserted or deleted • Far more likely to be deleterious because these shift the reading frame (triplet grouping)

  27. Sources of mutation* • Mutagenesis: Production of mutations • Spontaneous mutations: • Errors in replication coupled with subsequent errors in proofreading • Errors in chromosome (DNA) separation during cell division • Mutagens: Physical or chemical agents • X-rays, UV light (high energy photons)

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