Gene Expression: From DNA to Protein

1. Chapter 17From Gene to Protein

2. 17.1 – Genes specify proteins via transcription & translation • Gene Expression • DNA directs the synthesis of proteins (or RNA) • Includes translation & transcription • Proteins are the links between genotype and phenotype • One gene-one polypeptide hypothesis • Each gene codes for a polypeptide • Can be a protein or part of a protein

3. Messenger RNA (mRNA) • Produced during transcription • Carries the genetic message of DNA to the protein making machinery of the cell (ribosome) • In Eukaryotes • Transcription results in pre-mRNA, which undergoes RNA processing to yield the final mRNA • In Prokaryotes • Transcription directly makes mRNA • Transcription & Translation occur at the same time

5. Transcription • Synthesis of RNA using DNA as a template • Occurs in the nucleus • Only one strand of DNA is transcribed (called template strand) • The mRNA produced is a complementary strand • The mRNA base triplets are called codons • Written in the 5’ to 3’ direction

6. The genetic code is redundant • More than one codon codes for the 20 Amino Acids • Read based on a consistent reading frame • Groups of 3 must be read in the correct groupings in order for translation to be successful

7. All 64 codons were deciphered by the mid-1960s • Of the 64 triplets, 61 code for amino acids; 3 triplets are “stop” signals to end translation

8. Translation • Production of a polypeptide chain using mRNA • Occurs at the ribosomes • The instructions for the PP chain are written as a triplet code

10. The genetic code is nearly universal, shared by the simplest bacteria to the most complex animals • Genes can be transcribed and translated after being transplanted from one species to another

12. 17.2 – Transcription is the DNA-directed synthesis of RNA • RNA polymerase • Enzyme that separates the two DNA strands • Connects the RNA nucleotides as they base-pair • Can add RNA nucleotides only to the 3’ end so it elongates in the 5’ to 3’ direction • Uracil replaces thymine

13. Promoter • DNA sequence that RNA polymerase attaches • Terminator • DNA sequence that signals the end of transcription

14. Transcription unit • Entire stretch of DNA that is transcribed into RNA • May code for a polypeptide or an RNA such tRNA or rRNA

15. 3 stages of transcription • 1) Initiation • 2) Elongation • 3) Termination

16. 1) Initiation • In bacteria • RNA polymerase recognizes & binds to the promoter • In Eukaryotes • RNA polymerase II cannot bind to the promoter without supporting help from proteins known as transcription factors • Transcription Factors • Assist the binding of RNA polymerase to the promoter, & the initiation of transcription

17. Transcription initiation complex • The whole complex of RNA polymerase II & transcription factors • A promoter called a TATA box is crucial in forming the initiation complex in eukaryotes

19. 2) Elongation • RNA polymerase moves along the DNA (untwists the double helix) • 10 to 20 bases at a time • RNA nucleotides are continually added to the 3’ end of the growing chain • 40 nucleotides per second • As the complex moves down the DNA strand, the double helix re-forms with the new RNA molecule straggling away from the DNA template

21. 3) Termination • RNA transcript is released & the polymerase detaches upon transcribing a terminator sequence in the DNA

23. 17.3 – Eukaryotic cells modify RNA after transcription • Modifications to RNA after transcription: • Adding a 5’ cap & a poly-A tail • Facilitate the export of mRNA from the nucleus • Help protect mRNA from degradation by enzymes • Facilitate the attachment of the mRNA to the ribosome

24. RNA splicing (in Eukaryotic Cells) • Large portions of the newly made RNA strand are removed – called INTRONS • The ones left behind are called EXONS & are spliced together by a spliceosome

26. Special RNA called small nuclear RNA (snRNA) aid the spliceosomes • Play a role by catalyzing the excision of the introns & joining the exons • When RNA is an enzyme it is called a RIBOZYME

27. 17.4 – Translation is the RNA-directed synthesis of a polypeptide • Utilizes mRNA, tRNA, & rRNA • tRNA • Transfers AA from a pool of AA in the cytoplasm to a ribosome • The ribosome accepts the AA & adds it into a growing PP chain • Each tRNA is specific for an AA • One one end, it binds to the AA & the other end has a triplet called an anticodon which allows it to pair with a codon on an mRNA

29. Codon – mRNA triplet (there are 64) • mRNa is read codon by codon & one AA is added to the chain for each codon read • The rules for base-pairing between the third base of a codon & the corresponding tRNA anticodon are not as strict as DNA & mRNA so it is called a wobble

30. rRNA complexes with proteins • Forms the 2 subunits that form ribosomes • Ribosomes have 3 binding sites for tRNA • P-site – holds the tRNA that carries the growing PP chain • A-site – holds the tRNA that carries the AA that will be added next • E-site – exit site for tRNA

32. 3 stages of Translation • 1) Initiation • 2) Elongation • 3) Termination

33. Initiation • A) a small ribosomal subunit binds to mRNA in a way that the first codon of the mRNA strand (AUG) is placed in the proper position • B) tRNA with the anticodon UAC (carries the AA methionine), hydrogen bonds to the first codon (proteins called initiation factors aid) • C) Large subunit of ribosome attaches • Allows the tRNA with methionine to attach to the P-site • The A-site will now be available for the next tRNA with the 2nd AA

35. 2) Elongation • A) Codon Recognition • The codon in the A-site is matched by the incoming tRNA anticodon • B) Peptide bond formation • The incoming AA in the A-site forms a peptide bond with the existing chain of AA held in the P-site • Catalyzed by an rRNA (ribozyme) • C) Translocation • Occurs when tRNA in the A-site is moved to the P-site & the tRNA in the P-site is moved to the E-site • A-site is now clean and is ready for another AA

37. 3) Termination • A stop codon in the mRNA is reached & translation stops • A protein called release factor binds to the stop codon & the PP is freed from the ribosome • PP’s then will fold to assume their specific shape • May be modified further to make them functional • The destination of the protein is determined by the sequence of about 20 AA’s at the leading end of the PP chain (signal peptide)

39. 17.5 – Point mutations can affect structure & function • Point mutation are alterations of just one base pair – 2 basic types: • 1) Base-pair substitutions • The replacement of one nucleotide & its complementary base pair in the DNA with another pair of nucleotides • Missense – enable the codon to still code for an AA although it may not be the correct one • Nonsense – change a regular AA codon into a stop codon

42. 2) Insertions & deletions • Additions & loses of nucleotide pairs in genes • If they interfere with the codon groupings they can cause a frameshift mutation • Causes the mRNA to be read incorrectly

45. Mutagens • Substances or forces that interact with DNA in ways that cause mutations • X-rays & chemicals