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Biochemistry 2/e - Garrett & Grisham

Copyright © 1999 by Harcourt Brace & Company

Chapter 32

The Genetic Codeto accompany

Biochemistry, 2/e

byReginald 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, 6277Sea Harbor Drive, Orlando, Florida 32887-6777





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





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!





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 thatDNA sequence specified protein sequence

• Yanofsky provided better evidence in 1964: he

showed that the relative distances between

mutations in DNA were proportional to the distancesbetween amino acid sunstitutions in E. coli

tryptophan synthase





Elucidating the Genetic Code

• A triplet code is required: 43 = 64, but

42 = 16 - not enough for 20 amino

acids• But is the code overlapping?

• See Figure 32.2

• And is the code punctuated?





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





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?





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 thepossible trinucleotide codes, they elucidated all 64

correspondences in the code (Table 32.3)

• Read also about Khorana's experiment





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





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





Aminoacyl-tRNA Synthetases

Mechanism and specificity

• Deacylase activity "edits" and hydrolyzes

misacylated aminoacyl-tRNAs

• Despite common function, the synthetases area diverse collection of enzymes

• Four different quaternary structures: , 2, 4

and 22 • Subunits from 334 to more than 1000 residues

• Two different mechanisms (See Figure 32.5)





/ G & G





Bi h i t 2/ G tt & G i h





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






Biochemistr 2/e Garrett & Grisham





Biochemistry 2/e Garrett & Grisham





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 secondpossibility - the first base of the anticodon (which

matches the 3rd base of the codon) is referred to

as the "wobble position"





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

wooble hypotestic nd protein synthesis

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