Protein Synthesis

1. Protein Synthesis

2. What is the central dogma of biology?

3. Transcription: Initiation

4. Transcription: Initiation • RNA polymerase attaches to a “promoter” region in front (“upstream”) of a gene • Eukaryotes: RNA polymerase requires multiple “Transcription factor” proteins to be able to bind to the promoter • Promoters have characteristic DNA sequences (ex:TATA Box in eukaryotes – 5’ TATAAA 3’)

5. Transcription: Elongation

6. Transcription: Elongation • RNA production occurs in a 5’ to 3’ direction • “Template Strand” of DNA is the one that the RNA transcript is produced off of • The “Coding Strand” of the DNA will have the same sequence as the RNA transcript…EXCEPT?

7. Transcription: Termination

8. Transcription: Termination • mRNA transcript production continues until the end of the transcription unit is reached • Multiple mechanisms of termination exist

9. Post-Transcriptional mRNA processing (EUKARYOTES ONLY!) • Exons are the “coding” regions of the DNA and Introns are the “noncoding” regions of the DNA

10. How is the mRNA transcript modified? • The introns must be removed and the exons must be spliced together prior to the mRNA leaving the nucleus • **This is required to produce a functional protein

11. How does alternative splicing contribute to the production of various proteins? • Having multiple exons in a gene allows eukaryotes to make multiple functioning proteins from one gene

12. Ribosomes • Site of Protein Synthesis • “non-membrane” bound organelle • All cells have them • Composed of two subunits • Has 3 sites: • A site – where amino acids enter the ribosome • P site – where the growing polypeptide chain is • E site – where empty tRNA molecules leave

13. tRNA • Transfer RNA molecules • brings amino acids to the ribosome • Amino-acyl synthase enzyme catalyzes reaction to attach amino acid to tRNA • When carrying amino acid, “charged tRNA” • Anticodon region interacts with mRNA

14. The Genetic Code

15. The Genetic Code • Universal across ALL domains of Life! • Triplet Code: mRNA is read in units of three bases (codons) • There are 64 possible codons (for 20 amino acids) • The code is redundant and unambiguous • The code has “start” and “stop” codons

16. Translation: Initiation

17. Translation: Initiation • mRNA attaches to the small ribosomal subunit • Methionine is brought to the start codon (AUG) by the methionine tRNA • Start Codon is located in the P-site • tRNA binding is mediated by its “anti-codon” • “Translation Initiation Complex”

18. Translation: Elongation

19. Translation: Elongation • The next codon determines the next amino acid to be brought to the ribosome • Incoming, “Charged” tRNA enters at the A-site • Growing polypeptide chain is transferred to the new tRNA molecule – Peptide Bond Formed • Ribosome translocates (shifts) tRNA molecule with chain to P-site; previous (now “uncharged”) tRNA moves to E-site • Next codon available in A-site for new t-RNA

20. Translation: Termination

21. Translation: Termination • When a stop codon (UAG, UAA, or UGA) is encountered, a release factor binds to the A-site • The polypeptide chain is released • The ribosome disassembles

22. How does prokaryotic protein synthesis compare to eukaryotic?

23. How does prokaryotic compare to eukaryotic protein synthesis? • Transcription: Prokaryotes can bind DIRECTLY to the promoter region (no transcription factors required) • No Post-transcription modification of RNA • Prokaryotes have no nucleus so Transcription and Translation are coupled (occur together) – RNA is transcribed and translated at the same time

24. What is a polyribosome? How do prokaryotic and eukaryotic organisms benefit from them? • Many ribosomes translating the same RNA transcript • Enables simultaneous translation of one transcript