1. Today 14.2 & 14.4 Transcription and Translation http://highered.mheducation.com/sites/00725 07470/student_view0/chapter3/animation__p rotein_synthesis__quiz_3_.html

2. Gene Expression Revisited

3. Figure 14.6 UUU Second mRNA base UUC UUA UUG UCU UCC UCA UCG UAU UAC UAA UAG UGU UGC UGA UGG CUU CUC CUA CUG CCU CCC CCA CCG CAU CAC CAA CAG CGU CGC CGA CGG AUU AUC AUA AUG ACU ACC ACA ACG AAU AAC AAA AAG AGU AGC AGA AGG GUU GUC GUA GUG GCU GCC GCA GCG GAU GAC GAA GAG GGU GGC GGA GGG First mRNA base (5 end of codon) U C A G U C A G U C A G U C A G U C A G U C A G Phe Leu Ser Tyr Cys Trp Met or start Stop Arg Gln His Pro Leu Val Ala Asp Glu Gly I Ie Thr Lys Asn Arg Ser Third mRNA base (3 end of codon)

4. RNA polymerases assemble polynucleotides in the 5 to 3 direction However, RNA polymerases can start a chain without a primer

5. Figure 14.8-3 Transcription unit RNA polymerase Promoter Template strand of DNA Start point Termination Completed RNA transcript RNA transcript Unwound DNA Rewound DNA RNA transcript Direction of transcription (“downstream”) Initiation Elongation 3 5 3 5 3 5 3 5 3 5 3 5 3 5 3 5 3 5 3 5 3 2 1

6. Initiation Promoters Transcription Factors Transcription Initiation Complex

8. Figure 14.9 Transcription factors TATA box Promoter Nontemplate strand Start point Transcription initiation complex forms. Transcription initiation complex DNA RNA transcript A eukaryotic promoter Several transcription factors bind to DNA. 3 5 5 3 3 5 3 5 3 5 3 2 1 Template strand Transcription factors RNA polymerase II 3 5 3 5 TATAAAA ATATTTT

9. Elongation Many RNA polymerases can be acting at one time No primer needed RNA Polymerase can only add to 3’ end of new strand

10. Figure 14.10 Nontemplate strand of DNA Direction of transcription RNA polymerase 3 5 3 5 RNA nucleotides Template strand of DNA Newly made RNA 3 end 5 U C U G A A A A A A A A A A T T T T T T T C C C CC C C G G G U

11. Accurate translation requires two steps – First: a correct match between a tRNA and an amino acid, done by the enzyme aminoacyl-tRNA synthetase – Second: a correct match between the tRNA anticodon and an mRNA codon Flexible pairing at the third base of a codon is called wobble and allows some tRNAs to bind to more than one codon

12. Figure 14.16-1 Tyrosyl-tRNA synthetase Tyrosine (Tyr) (amino acid) Amino acid and tRNA enter active site. Tyr-tRNA Complementary tRNA anticodon 1 UAA

13. Figure 14.16-2 Tyrosyl-tRNA synthetase Tyrosine (Tyr) (amino acid) Amino acid and tRNA enter active site. Tyr-tRNA Complementary tRNA anticodon Using ATP, synthetase catalyzes covalent bonding. AMP  2 ATP 21 UAA P i

14. Figure 14.16-3 Tyrosyl-tRNA synthetase Tyrosine (Tyr) (amino acid) Amino acid and tRNA enter active site. Tyr-tRNA Complementary tRNA anticodon Aminoacyl tRNA released. Using ATP, synthetase catalyzes covalent bonding. AMP  2 ATP 2 3 1 UAA P i

15. Figure 14.17b PEA Large subunit Small subunit P site (Peptidyl-tRNA binding site) Exit tunnel E site (Exit site) mRNA binding site (b) Schematic model showing binding sites A site (Aminoacyl- tRNA binding site)

16. A ribosome has three binding sites for tRNA – The P site holds the tRNA that carries the growing polypeptide chain – The A site holds the tRNA that carries the next amino acid to be added to the chain – The E site is the exit site, where discharged tRNAs leave the ribosome

17. Building a Polypeptide The three stages of translation 1.Initiation 2.Elongation 3.Termination All three stages require protein “factors” that aid in the translation process

18. Ribosome Association and Initiation of Translation The initiation stage of translation brings together mRNA, a tRNA with the first amino acid, and the two ribosomal subunits A small ribosomal subunit binds with mRNA and a special initiator tRNA Then the small subunit moves along the mRNA until it reaches the start codon (AUG) Animation: Translation Introduction

19. A Small ribosomal subunit binds to mRNA. GTP P site U A mRNA 5 3 Met P i mRNA binding site Start codon Small ribosomal subunit Initiator tRNA 5 3 5 3 Large ribosomal subunit completes the initiation complex. U G C 5 3 Translation initiation complex Large ribosomal subunit A E Met GDP  12 Proteins called initiation factors bring all these components together

20. Elongation of the Polypeptide Chain During elongation, amino acids are added one by one to the previous amino acid at the C-terminus of the growing chain Each addition involves proteins called elongation factors and occurs in three steps: codon recognition, peptide bond formation, and translocation Translation proceeds along the mRNA in a 5 to 3 direction

21. Figure 14.19-1 Amino end of polypeptide mRNA P site P i 5 3 E GTP  A site GDP Codon recognition E P A 1

22. Figure 14.19-2 Amino end of polypeptide mRNA P site P i 5 3 E GTP  A site GDP Peptide bond formation Codon recognition E P A E P A 2 1

23. Figure 14.19-3 Amino end of polypeptide mRNA Ribosome ready for next aminoacyl tRNA P site P i 5 3 E GTP  A site GDP Peptide bond formation Codon recognition Translocation E P A E P A P i GTP  GDP E P A 3 2 1

24. Polypeptide synthesis always begins in the cytosol Synthesis finishes in the cytosol unless the polypeptide signals the ribosome to attach to the ER Polypeptides destined for the ER or for secretion are marked by a signal peptide

25. A signal-recognition particle (SRP) binds to the signal peptide The SRP brings the signal peptide and its ribosome to the ER

26. Figure 14.21 Polypeptide synthesis begins. SRP binds to signal peptide. SRP binds to receptor protein. SRP detaches and polypeptide synthesis resumes. Signal- cleaving enzyme cuts off signal peptide. Completed polypeptide folds into final conformation. Signal peptide removed ER membrane Signal peptide Protein SRP receptor protein Ribosome mRNA CYTOSOL ER LUMEM SRP Translocation complex 12 3 4 5 6