1. From gene to protein

2. Our plan: Overview gene expression Walk through the process –Review structure and function of DNA –Transcription –Translation Gene expression and mutations

3. Gene expression- the process by which DNA directs the synthesis of proteins Overview DNA mRNAProtein TranscriptionTranslation

4. Overview: Cell as a city DNA: Blueprint for the city Nucleus: City Hall Nuclear envelope: Fence around City Hall

5. Overview: Cell as a city DNA mRNAProtein TranscriptionTranslation The blueprint cannot leave City Hall Photocopies of the blueprint can be taken out into the city Architects and builders translate the blue print into the city’s infrastructure The DNA remains in the nucleus Transcription generates mobile RNA transcripts using DNA as a template The RNA sequence can be translated into a protein

6. Overview DNA mRNAProtein TranscriptionTranslation

7. DNA Hydrogen bond 3 end 5 end 3.4 nm 0.34 nm 3 end 5 end (b) Partial chemical structure (a) Key features of DNA structure 1 nm Genes- discrete units of hereditary information consisting of a specific nucleotide sequence of DNA

8. Overview DNA mRNAProtein TranscriptionTranslation

9. Transcription Transcription-DNA guides the production of RNA Takes place in three phases: Initiation Elongation Termination

10. Transcription Transcription-DNA guides the production of RNA Takes place in three phases: Initiation Elongation Termination RNA polymerase binds to a promoter in the DNA (contains a signal sequence and at start point) The DNA is separated and unwound Transcription begins

11. - A eukaryotic promoter includes a TATA box 3 1 2 3 Promoter TATA box Start point Template DNA strand 5 3 5 Transcription factors Several transcription factors must bind to the DNA before RNA polymerase II can do so. 5 53 3 Additional transcription factors bind to the DNA along with RNA polymerase II, forming the transcription initiation complex. RNA polymerase II Transcription factors 5 5 5 3 3 RNA transcript Transcription initiation complex

12. Transcription Transcription-DNA guides the production of RNA Takes place in three phases: Initiation Elongation Termination Elongation RNA polymerase Nontemplate strand of DNA RNA nucleotides 3 end Direction of transcription (“downstream”) Template strand of DNA Newly made RNA 3 5 5 RNA polymerase moves along the template strand Untwists the helix Adds complimentary RNA nucleotides to the 3’ end of the chain

13. Transcription Transcription-DNA guides the production of RNA Takes place in three phases: Initiation Elongation Termination Transcription terminates after a special sequence is transcribed Termination sequence (proks-I.e. hairpins) Polyadenylation sequence (euks) The transcript is cut and released

14. Transcription Eukaryotic cells modify RNA before it enters the cytoplasm Both ends of the transcript are processed Some sections are cleaved and those remaining are spliced together

15. Transcription Eukaryotic cells modify RNA before it enters the cytoplasm Both ends of the transcript are processed Some sections are cleaved and those remaining are spliced together 5’ end receives a 5’ cap 3’ end receives a poly-A tail Protein-coding segment Polyadenylation signal 3 3 UTR5 UTR 5 5 Cap Start codon Stop codon Poly-A tail G PPPAAUAAA AAA …

16. Transcription Eukaryotic cells modify RNA before it enters the cytoplasm Both ends of the transcript are processed Some sections are cleaved and those remaining are spliced together Both: Facilitate the export of the mRNA Protect mRNA from degradation Help ribosomes attach Protein-coding segment Polyadenylation signal 3 3 UTR5 UTR 5 5 Cap Start codon Stop codon Poly-A tail G PPPAAUAAA AAA … 5’ end receives a 5’ cap 3’ end receives a poly-A tail

17. Transcription Eukaryotic cells modify RNA before it enters the cytoplasm Both ends of the transcript are processed Some sections are cleaved and those remaining are spliced together Introns (non-coding regions) are cut Exons (coding regions) are spliced together Pre-mRNA mRNA Coding segment Introns cut out and exons spliced together 5 Cap Exon Intron 5 1 30 31104 ExonIntron 105 Exon 146 3 Poly-A tail 5 Cap 5 UTR3 UTR 1 146

18. Transcription Question: What would be the sequence of RNA generated from the following DNA template strand? DNA: 3’-A T C C G T-5’

19. Transcription Question: What would be the sequence of RNA generated from the following DNA template strand? DNA: 3’-A T C C G T-5’ mRNA:5’-U A G G C A-3’

20. Overview DNA mRNA Protein TranscriptionTranslation

21. Proteins are made from polypeptide polymers, which are made from amino acid monomers Antibody protein Protein from flu virus

22. Translation Proteins are made from polypeptide polymers, which are made from amino acid monomers Antibody protein Protein from flu virus

23. Translation Proteins are made from polypeptide polymers, which are made from amino acid monomers Antibody protein Protein from flu virus How is the information in RNA transformed into an amino acid?

24. Translation-The genetic code Only four nucleotides in RNA and 20 amino acids (the genetic code is not like Chinese) More like English, different permutations of letters build meaningful words –Only 16 two-letter combinations of nucleotides (4 2 ) –64 possibilities if three-nucleotide combinations code for an amino acid

25. Translation-The genetic code Only four nucleotides in RNA and 20 amino acids (the genetic code is not like Chinese) More like English, different permutations of letters build meaningful words –Only 16 two-letter combinations of nucleotides (4 2 ) –64 possibilities if three-nucleotide combinations code for an amino acid

26. Translation-The genetic code Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon) Genetic instructions for a polypeptide are written in RNA as a series of non- overlapping three- nucleotide “words” (codons)

27. Translation-The genetic code Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon) Genetic instructions for a polypeptide are written in RNA as a series of non- overlapping three- nucleotide “words” (codons) Ie. 5’-AAG-3’=lysine

28. Translation-The genetic code Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon) Genetic instructions for a polypeptide are written in RNA as a series of non- overlapping three- nucleotide “words” (codons) Ie. 5’-AAG-3’=lysine 61 code for amino acids

29. Translation-The genetic code Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon) Genetic instructions for a polypeptide are written in RNA as a series of non- overlapping three- nucleotide “words” (codons) Ie. 5’-AAG-3’=lysine 61 code for amino acids 3 stop codons

30. Translation-The genetic code Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon) Genetic instructions for a polypeptide are written in RNA as a series of non- overlapping three- nucleotide “words” (codons) Ie. 5’-AAG-3’=lysine 61 code for amino acids 3 stop codons AUG=Methionine or start

31. Translation-The genetic code Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon) Genetic instructions for a polypeptide are written in RNA as a series of non- overlapping three- nucleotide “words” (codons) Ie. 5’-AAG-3’=lysine 61 code for amino acids 3 stop codons AUG=Methionine or start The code is redundant, but not ambiguous

32. Translation-The genetic code Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon) Genetic instructions for a polypeptide are written in RNA as a series of non- overlapping three- nucleotide “words” (codons) Ie. 5’-AAG-3’=lysine 61 code for amino acids 3 stop codons AUG=Methionine or start The code is redundant, but not ambiguous This code is nearly universal

33. Amino acid attachment site Hydrogen bonds Anticodon 3 5 Translation Just as the architects and builders translate the copied blueprints into the city’s infrastructure, tRNA translates mRNA codons into amino acids tRNA Consists of a single strand of RNA Anticodon on one end (can bind with an mRNA codon) Corresponding amino acid on upper portion

34. Translation Three stages: Initiation Elongation Termination

35. Translation Three stages: Initiation Elongation Termination 3 3 5 5 U U A A C G Met GTP GDP Initiator tRNA mRNA 5 3 Start codon mRNA binding site Small ribosomal subunit 5 P site Translation initiation complex 3 EA Met Large ribosomal subunit Initiation begins when mRNA binds to the small sub-unit of a ribosome

36. Translation Three stages: Initiation Elongation Termination 3 3 5 5 U U A A C G Met GTP GDP Initiator tRNA mRNA 5 3 Start codon mRNA binding site Small ribosomal subunit 5 P site Translation initiation complex 3 EA Met Large ribosomal subunit Initiation begins when mRNA binds to the small sub-unit of a ribosome Charged tRNA binds to the start codon

37. Translation Three stages: Initiation Elongation Termination 3 3 5 5 U U A A C G Met GTP GDP Initiator tRNA mRNA 5 3 Start codon mRNA binding site Small ribosomal subunit 5 P site Translation initiation complex 3 EA Met Large ribosomal subunit Initiation begins when mRNA binds to the small sub-unit of a ribosome Charged tRNA binds to the start codon Large ribosomal sub-unit binds Cluster is called the initiation complex

38. Translation Three stages: Initiation Elongation Termination Amino end of polypeptide mRNA 5 3 E P site A site GTP GDP E P A E PA GTP Ribosome ready for next aminoacyl tRNA E P A New tRNAs come in and bind at A site as their complimentary condon is made available

39. Translation Three stages: Initiation Elongation Termination Amino end of polypeptide mRNA 5 3 E P site A site GTP GDP E P A E PA GTP Ribosome ready for next aminoacyl tRNA E P A New tRNAs come in and bind at A site as their complimentary condon is made available New amino acids is bonded to the growing chain

40. Translation Three stages: Initiation Elongation Termination Amino end of polypeptide mRNA 5 3 E P site A site GTP GDP E P A E PA GTP Ribosome ready for next aminoacyl tRNA E P A New tRNAs come in and bind at A site as their complimentary condon is made available New amino acids is bonded to the growing chain Complex moves to free the A site

41. Translation Three stages: Initiation Elongation Termination Amino end of polypeptide mRNA 5 3 E P site A site GTP GDP E P A E PA GTP Ribosome ready for next aminoacyl tRNA E P A New tRNAs come in and bind at A site as their complimentary condon is made available New amino acids is bonded to the growing chain Complex moves to free the A site tRNA that shifted to E site exits

42. Translation Three stages: Initiation Elongation Termination Amino end of polypeptide mRNA 5 3 E P site A site GTP GDP E P A E PA GTP Ribosome ready for next aminoacyl tRNA E P A New tRNAs come in and bind at A site as their complimentary condon is made available New amino acids is bonded to the growing chain Complex moves to free the A site tRNA that shifted to E site exits New tRNA binds at the A site

43. Translation Three stages: Initiation Elongation Termination Termination occurs when a ribosome encounters a stop codon Release factors bind mRNA and polypeptide are released Release factor 3 5 Stop codon (UAG, UAA, or UGA) 5 3 2 Free polypeptide 2 GDP GTP 5 3

44. Transcription Question: What polypeptide sequence would be generated from the following DNA template strand? DNA: 3’-T T C A G T-5’ Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon)

45. Transcription Question: What polypeptide sequence would be generated from the following DNA template strand? DNA: 3’-T T C A G T-5’ RNA: 5’-A A G U C A-3’ Peptide sequence: Lysine, serine Second mRNA base First mRNA base (5 end of codon) Third mRNA base (3 end of codon)

46. Summary DNA mRNA Protein TranscriptionTranslation

47. Point mutations Impacts of genetic mutation on gene expression Point mutations- change in a single base pair –Base-pair substitution Silent-no change in polypeptide Missense (substituted amino acid) Nonsense (early stop codon)

48. Point mutations Impacts of genetic mutation on gene expression Point mutations- change in a single base pair –Insertion and deletion Addition or loss of base pairs Causes a frame shift

49. You should understand: The process of gene expression The impact of point mutations on gene expression DNA mRNA Protein TranscriptionTranslation