The genetic code and transcription

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Chapter 14

• The genetic code and transcription
• Introduction of translation (continuation in
  chapter 15)
• Skip experiments skip pages 353-359

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(No Transcript)
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Triplet code

• 64 codons to specify the 20 amino acids
• Code is degenerate
• Stop and start codons
• Mutations

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Degenerate code 64 codons and 20 amino acids
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Wobble hypothesis

• hydrogen bonding between the codon and anticodon
  at the third position is subject to modified
  base-pairing rules
• May not adhere specifically to the established
  base-pairing rules

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Codons

• First amino acid
• Methionine or N-formyl methionine
• N-formyl methionine in bacteria
• Formyl group or the entire N-formyl methionine
  removed from the final protein
• AUG is the only codon to encode for methionine.
• When AUG appears internally in mRNA, an
  unformylated methionine is inserted into the
  protein.

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Stop codons

• UAG, UAA, and UGA
• Do not specify any amino acid
• Nonsense mutations
• Change the normal sequence to one of the stop
  codons and protein synthesis stops
• Frameshift may result in reaching a stop codon

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Types of mutations

• Point mutation
• Missense mutation
• Nonsense mutation
• Silent mutations
• Transition
• Transversion
• Frameshift mutations

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Types of mutations
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The genetic code is nearly universal
mitochondria is different
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Different initiation points create overlapping
genes

• Single mRNA may have multiple initiation points
  for translation produce different proteins

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RNA polymerase is used in transcription

• RNA polymerase binds to the promoter
• Makes RNA from a DNA template
• No primer is required
• Uses ribonucleotides instead of
  deoxyribonucleotides

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DNA is transcribed into RNA
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RNA polymerase from E. coli

• contains the subunits a2, ß, ß', and s
• The s subunit is responsible for promoter
  recognition
• s subunit dissociates after synthesis begins (8-9
  nucleotides)

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E. coli promoters

• have two consensus sequences important for RNA
  polymerase binding

TATAAT, positioned at 10 Pribnow box TTGACA,
positioned at -35 Closer the sequence is to
consensus, the stronger the promoter is

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Promoter sequence

• Two important consensus sequences
• -10 TATAAT (similar to eukaryotic -25 TATAAAA)
• -35 TTGACA
• Sequences on either side of -10 and -35 or in
  between are relatively unimportant in promoter
  recognition
• Where to start transcribing
• 1 sequence (10 bases upstream from the -10)
• Which strand will be read
• Template strand (3 to 5 on the DNA)
• Which direction the RNA polymerase will move
• 5 to 3

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Promoter sequence is on the nontemplate strand of
DNA
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Problem

• DNA
• 5CCGTGGACCTACGTACTATTATAGCTAGCCCTAGATTGGCCTGTCAAC
  GCGG3
• 3GGCACCTGGATGCATGATAATATCGATCGGGATCTAACCGACAGTTGC
  GCC5
• Where is the promoter sequence located?
• Which is the template strand?
• Which direction will transcription occur?
• Where is the start site for transcription?

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Steps in transcription

• Initiation?Elongation?
• Termination
• Initiation RNAP binds the promoter
• Elongation
• RNA nucleotides added at the 3 end of growing
  RNA strand (5?3 direction)
• Termination reach a specific sequence and new
  RNA molecule is released

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Steps in transcription
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Termination

• Terminator where RNA polymerase disassociates
  from DNA
• Sequence on DNA transcribed into RNA
• Common sequence on the DNA is an inverted repeat
  (prokaryotes)
• RNA creates a hairpin secondary structure
• form a stem-loop structure followed by a string
  of Us
• A-U base pairs needed to destabilize the DNA-RNA
  interaction and is needed to end transcription

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RNA transcribes off the template strand

• DNA
• 5TGCTGGTCGACTG3 (1)
• 3ACGCCCAGCTGAC5 (2)
• RNA
• 5UGCGGGUCGACUG3
• DNA strand 1 nontemplate
• DNA strand 2 template

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Can you write a sequence in the DNA or RNA that
will form a stem loop structure?
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Rho dependent termination

• Protein that binds to RNA and moves down to RNA
  polymerase-DNA complex
• A downstream portion of the newly synthesized RNA
  forms a stem loop
• Causes RNA polymerase to disassociate from DNA
• Rho breaks the H-bonds between the DNA-RNA hybrid
  and the RNA dissociates

(from Genomes 2)
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Polycistronic mRNA

• Unlike Eukaryotes, Prokaryotes have Polycistronic
  mRNA
• Group of cotranscribed genes
• One promoter and one terminator
• Operon