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Chapter 32 Slides

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  • 1. Chapter 32 The Genetic Code to accompany Biochemistry, 2/e by Reginald Garrett and Charles Grisham All rights reserved. Requests for permission to make copies of any part of the work should be mailed to: Permissions Department, Harcourt Brace & Company, 6277 Sea Harbor Drive, Orlando, Florida 32887-6777
  • 2. Outline
    • 32.1 Elucidating the Genetic Code
    • 32.2 The Nature of the Genetic Code
    • 32.3 The Second Genetic Code
    • 32.4 Codon-Anticodon Pairing, Third-Base Degeneracy and the Wobble Hypothesis
    • 32.5 Codon Usage
    • 32.6 Nonsense Suppression
  • 3. Translating the Message
    • How does the sequence of mRNA translate into the sequence of a protein?
    • What is the genetic code?
    • How do you translate the "four-letter code" of mRNA into the "20-letter code" of proteins?
    • And what are the mechanics like? There is no obvious chemical affinity between the purine and pyrimidine bases and the amino acids that make protein.
    • As a "way out" of this dilemma, Crick proposed "adapter molecules" - they are tRNAs !
  • 4.  
  • 5. The Collinearity of Gene and Protein Structures
    • Watson and Crick's structure for DNA, together with Sanger's demonstration that protein sequences were unique and specific, made it seem likely that DNA sequence specified protein sequence
    • Yanofsky provided better evidence in 1964: he showed that the relative distances between mutations in DNA were proportional to the distances between amino acid sunstitutions in E. coli tryptophan synthase
  • 6. Elucidating the Genetic Code
    • A triplet code is required: 4 3 = 64, but 4 2 = 16 - not enough for 20 amino acids
    • But is the code overlapping ?
    • See Figure 32.2
    • And is the code punctuated ?
  • 7.  
  • 8. The Nature of the Genetic Code
    • A group of three bases codes for one amino acid
    • The code is not overlapping
    • The base sequence is read from a fixed starting point, with no punctuation
    • The code is degenerate (in most cases, each amino acid can be designated by any of several triplets
  • 9. Biochemists Break the Code
    • Assignment of "codons" to their respective amino acids was achieved by in vitro biochemistry
    • Marshall Nirenberg and Heinrich Matthaei showed that poly-U produced polyphenylalanine in a cell-free solution from E. coli
    • Poly-A gave polylysine
    • Poly-C gave polyproline
    • Poly-G gave polyglycine
    • But what of others?
  • 10. Getting at the Rest of the Code
    • Work with nucleotide copolymers (poly (A,C), etc.), revealed some of the codes
    • But Marshall Nirenberg and Philip Leder cracked the entire code in 1964
    • They showed that trinucleotides bound to ribosomes could direct the binding of specific aminoacyl-tRNAs (See Figure 31.6)
    • By using C-14 labelled amino acids with all the possible trinucleotide codes, they elucidated all 64 correspondences in the code (Table 32.3)
    • Read also about Khorana's experiment
  • 11.  
  • 12. Features of the Genetic Code
    • All the codons have meaning : 61 specify amino acids, and the other 3 are "nonsense" or "stop" codons
    • The code is unambiguous - only one amino acid is indicated by each of the 61 codons
    • The code is degenerate - except for Trp and Met , each amino acid is coded by two or more codons
    • Codons representing the same or similar amino acids are similar in sequence
    • 2nd base pyrimidine: usually nonpolar amino acid
    • 2nd base purine: usually polar or charged aa
  • 13. AA Activation for Prot. Synth.
    • The Aminoacyl-tRNA Synthetases
    • Codons are recognized by aminoacyl-tRNAs
    • Base pairing must allow the tRNA to bring its particular amino acid to the ribosome
    • But aminoacyl-tRNAs do something else: activate the amino acid for transfer to peptide
    • Aminoacyl-tRNA synthetases do the critical job - linking the right amino acid with "cognate" tRNA
    • Two levels of specificity - one in forming the aminoacyl adenylate and one in linking to tRNA
  • 14.  
  • 15. Aminoacyl-tRNA Synthetases
    • Mechanism and specificity
    • Deacylase activity "edits" and hydrolyzes misacylated aminoacyl-tRNAs
    • Despite common function, the synthetases are a diverse collection of enzymes
    • Four different quaternary structures:  ,  2 ,  4 and  2  2
    • Subunits from 334 to more than 1000 residues
    • Two different mechanisms (See Figure 32.5)
  • 16.  
  • 17.  
  • 18.  
  • 19. Recognition of tRNAs
    • by the aminoacyl-tRNA synthetases
    • Anticodon region is not the only recognition site
    • The "inside of the L" and other regions of the tRNA molecule are also important
    • Read pages 1080-1082 on specificity of several aminoacyl-tRNA synthetases
  • 20.  
  • 21.  
  • 22.  
  • 23.  
  • 24.  
  • 25. Third-Base Degeneracy
    • and the Wobble Hypothesis
    • Codon-anticodon pairing is the crucial feature of the "reading of the code"
    • But what accounts for "degeneracy": are there 61 different anticodons, or can you get by with fewer than 61, due to lack of specificity at the third position?
    • Crick's Wobble Hypothesis argues for the second possibility - the first base of the anticodon (which matches the 3rd base of the codon) is referred to as the "wobble position"
  • 26.  
  • 27. The Wobble Hypothesis
    • The first two bases of the codon make normal (canonical) H-bond pairs with the 2nd and 3rd bases of the anticodon
    • At the remaining position, less stringent rules apply and non-canonical pairing may occur
    • The rules: first base U can recognize A or G, first base G can recognize U or C, and first base I can recognize U, C or A (I comes from deamination of A)
    • Advantage of wobble: dissociation of tRNA from mRNA is faster and protein synthesis too