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The Genetic Code CHARACTERISTICS OF THE GENETIC CODE: 1. In linear form

Uses the 4 ribonucleotides on mRNA (A, U, G, C)

Sequence is complimentary to nucleotide sequence of DNA

2. In triplet Codon language of mRNA Each word is made of 3

nucleotides specifying 1 amino acid

mRNA and Triplet Code Words

# of codons

Amino acids

1 Met, Trp

2 Asn, Asp,Cys, Gln,Glu,His, Lys, Phe, Tyr

3 Ile

4 Ala, Gly, Pro, Thr, Val

6 Arg, Leu, Ser

codon codon

Methionine Valine

3. Unambigous 1 codon ->1 amino acid only

5. Contains

start signals- initiate translation stop signals- terminate translation

AUG UAA UAG UGA Start Stop (Methionine)

6. Commaless once translaton begins, codons are

read continuously without breaks no internal punctuation

4. Degenerate a given amino acid can be specified

by more than one triplet codon GUU GUA GUC GUG Valine Valine is an example of the degeneracy of the code. All four of these triplets code for valine.

7. Nonoverlapping each nucleotide is part of only 1

codon 8. Universal (nearly)

With minor exceptions, all organisms use a single coding dictionary

Early Studies that Established the Basic

Operational Patterns of the Genetic Code

In the late 1950s, researchers thought that DNA itself might directly encode proteins during their synthesis. Francois Jacob and Jacques Monod

suggested that an RNA molecule was intermediate between DNA and proteins

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Once this mRNA was discovered, the question was how could just 4 bases code for 20 amino acids. Sidney Brenner Argued that the code must be a triplet since three letter words represent the minimal use of four letters to specify 20 amino acids. A code of 4 nucleotides taken 2 at a time 42 = 16 nucleotides A code of 4 nucleotides taken 3 at a time 43 = 64 nucleotides Francis Crick and colleagues Frameshift Mutation

! Occurs as a result of the addition or deletion of one or more nucleotides in the gene.

! Gain or loss of one or two nucleotides will shift the frame of reading.

! When three nucleotides are involved, the frame of reading is reestablished.

! This would not occur if the code is not triplet.

Deciphering the Code Cell- Free Polypeptide Synthesis

! Because the enzyme used in this experiment

does not require a template, each addition of a ribonucleotide is random, based on the relative concentration of the ribonucleoside diphosphates. ! The probability of the insertion of a specific ribonucleotide is proportional to the availability of that molecule, relative to other available ribonucleotides.

! must contain the essential factors for protein synthesis: ribosomes, tRNA, amino acids, and other molecules essential to translation

! amino acids were labeled ! enzyme to be used was isolated bacteria and it functions to degrade RNA " discovered in 1955 by Marianne Grunberg-Manago and Severo Ochoa " in vitro, with high concentrations of ribonucleosides dephosphates, the reaction can be forced to synthesize RNA " unlike, RNA polymerase, it does not require a DNA template

Use of Homopolymers

Steps: 1. In a series of experiments, RNA

polymers were synthesized using just one type of ribonucleotide.

2. Researchers labeled 1 of 20 amino acids added.

3. Each RNA were tested to determine which, if any, amino acids were incorporated into the newly synthesized proteins.

Result:

Mixed Copolymers

! Makes use of RNA heteropolymers- 2 or

more different ribonucleoside diphosphates are added in combination.

! Recall that, Nirenberg and Matthei said that, the probability of the insertion of a specific ribonucleotide is proportional to the availability of that molecule, relative other available ribonucleotides.

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! If they knew the relative proportion of each type of ribonucleoside diphosphate, they could predict the frequency of any particular triplet codon occurring in the synthetic mRNA.

Example:

! Suppose A and C are added at a ratio of 1A:5C

! Then: there is a 1/6 chance for an A and a 5/6

chance for a C ! So:

! Therefore: By examining the percentages of any given amino acid incorporated into the protein synthesized under the direction of this message, a probable base compositions for each amino acid can be proposed.

Triplet Binding Assay ! Developed by Nirenberg and Philip Leder

! The technique took advantage of the observation that ribosomes, when presented with an RNA sequence as short

as three ribonucleotides, will bind to it and attract the correct charged tRNA corresponding to the triplet code.

! The triplet codon in mRNA attracts a

complimentary sequence within the tRNA

Possible Components

Probability of occurrence of any triplet

Possible Triplets

Final%

3A (1/6)3= 1/216=0.4% AAA 0.4

1C:2A (5/6)(1/6)2=5/216=2.3% AAC ACA ,CAA

3x2.3=6.9

2C:1A (5/6)2(1/6)=25/216=11.6% ACC CAC CCA

3x11.6=34.8

3C (5/6)3=125/216=57.9% CCC 57.9

Percentage of amino acids in protein

Probable base- composition Assignments

Lysine

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Repeating Copolymers

! Developed by Gobind Khorona in the early 60s

! Long RNA molecules are synthesized consisting of short sequences repeated many times

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Wobble Hypothesis

! A pattern of degeneracy at the third position ! Discovered by Crick in 1966 ! The first two base pairings in a codon-anticodon interaction is more critical while the

third is more relaxed.