the genetic code each amino acid is specified by a triplet of

The genetic code

• Each amino acid is specified by a triplet of nucleotides, known as a codon.

The Genetic Code

• TTT Phe TCT Ser TAT Tyr TGT Cys• TTC Phe TCC Ser TAC Tyr TGC Cys• TTA Leu TCA Ser TAA Och TGA Umb• TTG Leu TCG Ser TAG Amb TGG Trp

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• CTT Leu CCT Pro CAT His CGT Arg• CTC Leu CCC Pro CAC His CGC Arg• CTA Leu CCA Pro CAA Gln CGA Arg• CTG Leu CCG Pro CAG Gln CGG Arg

•

• ATT Ile ACT Thr AAT Asn AGT Ser• ATC Ile ACC Thr AAC Asn AGC Ser• ATA Ile ACA Thr AAA Lys AGA Arg• ATG Met ACG Thr AAG Lys AGG Arg

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• GTT Val GCT Ala GAT Asp GGT Gly• GTC Val GCC Ala GAC Asp GGC Gly• GTA Val GCA Ala GAA Glu GGA Gly• GTG Val GCG Ala GAG Glu GGG Gly

The Genetic Code


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ATG MetSingle methionine

codon acts as initiator

The Genetic Code


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TAA Och

TGA Umb TAG Amb Three nonsense codons

act as stop signals

The Genetic Code


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Some amino acids (e.g.

leucine) have up to six codons

The Genetic Code


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Some aminoacids (e.g.proline) have fourcodons

The Genetic Code


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Some aminoacids (e.g.

glutamine) have twocodons

The Genetic Code


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Tryptophan andmethionine

have one codoneach

The Genetic Code


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The last nucleotidein a codon

is often irrelevant

ACT Thr ACC Thr ACA Thr ACG Thr

The Genetic Code


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When the last nucleotide

does matter, it is usually

only important whether it is a purine or pyrimidine

CAT His CAC His CAA Gln CAG Gln

The Genetic Code

A nucleotide consists of a ribose sugar bonded to phosphoric acid, with a nitrogen base of either a pyrimidine (cytosine or thymine) or purine (adenine or guanine) as a side chain. A base called Uracil replaces all thymine bases in mRNA.

The Genetic Code


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• GTT Val GCT Ala GAT Asp GGT Gly• GTC Val GCC Ala GAC Asp GGC Gly• GTA Val GCA Ala GAA Glu GGA Gly• GTG Val GCG Ala GAG Glu GGG GlyWith methionine

and tryptophan, the exact base

matters

ATT Ile ATC Ile ATA Ile

ATG Met

The Genetic Code

Recommended supplementary reading

Chatty, readable account of how Crick and Brenner solved the mystery of the genetic code. This is not a textbook. It is Francis Crick’s autobiographical

answer to James Watson’s book The Double Helix, which describes the search for the structure of

DNA, and in which Watson notes the dictionary definition of a crick as “a pain in the neck”.

Crick, F. What Mad Pursuit? 1989

(James Cameron-Gifford Library Q143.C7, George Green Library QH506.CRI)

The Genetic Code

How was the code deciphered?

Most of the work to show the general form of the genetic code was done by Francis Crick and Sidney Brenner.

Crick Brenner

They started off with George Gamow’s

arguments based on simple school

arithmetic to show that the code was probably

a triplet code.

Crick

Brenner


Why must the code be in triplets?

There are only four nucleotides, therefore a singlet code (i.e. a code in which each nucleotide

specifies an amino acid) could only encode four amino acids.

However, there are twenty amino acids found in most proteins. Therefore, the code cannot be

singlet in nature.

If the code were doublet, then there would be four possible

nucleotides in the first position and four in the second. This gives:

4 4 = 42 = 16 codons

Still too few to encode 20 amino acids.

G

A

T

C

GATC

GATC

GATC

GATC


If the code were triplet, then there would be four possible nucleotides in the first position, four in

the second and four in the third. This gives:

4 4 4 = 43 = 64 permutations

This is too many to encode 20 amino acids but the code could work if either some permutations are not used or if more than one encodes each amino

acid (or both).


Type of code Number of permutationsSinglet 41 = 4 Doublet 42 = 16 Triplet 43 = 64 Quadruplet 44 = 256 Pentuplet 45 = 1024

Only the triplet code really looks feasible


There are also different ways that the code can be read:

• It can be punctuated or unpunctuated.

• If it is unpunctuated it can be overlapping or non-overlapping.


An overlapping code

GTCACCCATGGAGGTATCT

1

2

3

4

Once the first codon is set (e.g. GTC), the next one can only be one of four (TCA, TCG, TCT or TCC). This is a disadvantage.

A non-overlapping unpunctuated codeGTCACCCATGGAGGTATCT

1 2 3 4 5

1 2 3 4 5

1 2 3 4 5

There are three ways to read this type of code, referred to as “reading frames”. This makes this type of code non-ideal.

GTCACCCATGGAGGTATCT

1 2 3 4 5

Here, one nucleotide (A) is used as a punctuation mark. This code has several advantages:1. The reading frame is set by the punctuation.2. Because only three nucleotides are used in codons, the number of coding permutations available is 33 = 27 amino acids

A non-overlapping punctuated code

Is the code really overlapping?GTCACCCATGGAGGTATCT

1

2

3

4

Once the first amino acid is set, the next one can only be one of four. Therefore, certain amino acids could never be next to each other.

This can be tested by experimentation

• Francis Crick and Sidney Brenner did “nearest neighbour” analysis on real proteins.

• They found that any amino acid could be next to any other one. Therefore, the code cannot be overlapping.

Is the code really overlapping?

Is the code punctuated?

• Francis Crick and Sidney Brenner went on to analyse a particular type of mutant that is induced by intercalating agents (e.g. acridine dye).

• Intercalating agents will insert themselves between the base pairs of DNA. These can stretch the base pairs apart during replication and cause an extra nucleotide to be inserted or one to be left out.

• They found a gene (the rII gene) that has special properties. It can tolerate several wrong codons in the early part of the coding sequence and still make an active protein as long as the later part of the coding sequence is correct.


• The mutations caused by intercalating agents fall into two classes, 1 and 2. Both cause a mutant phenotype in the rII gene.

1

2Mutant phenotype

Mutant phenotype


• Double mutants (two mutations in one gene) also cause a mutant phenotype in the rII gene.

1

2

1

2


Mutant phenotype

Mutant phenotype

• When the double mutant has two different kinds of mutation, they suppress each other and you get a non-mutant phenotype in the rII gene.

Wild type (non-mutant) phenotype

12

• Remember that the mutations caused by acridine dyes result from the loss or gain of one nucleotide.

• They cause the reading frame to change and are called frame-shift mutations.

• The fact that they can arise means that there must be reading frames and that means that the code in unpunctuated.

How does this work?

GTCACCCATGGAGGTATCT 1 2 3 4 5

Original code

GTCTACCCATGGAGGTATC

1 2 3 4 5

Code with frame shift mutationAll codons after the inserted nucleotide are wrong (some may be stop codons).

Two wrongs can make a right


Original code

GTCTACCATGGAGGTATCT

1 2 3 4 5

Code with different frame shift mutations After second mutation, codons back in original frame.

Is the code triplet?

• Crick and Brenner went on to show that three frame shift mutations of the first type (insertion) or three of the second type (deletion) in the rII gene could also give a wild-type phenotype.

• This could only happen if the code was triplet. If the code was quadruplet then you would have to add or delete four nucleotides to reset the reading frame.

Three wrongs can make a right


Original code

GTCTACTCACATGGAGGTA

1 2 3 4 5 6

Code with three similar frame shift mutations An extra codon is inserted and a few codons are wrong, then all of the rest are OK.

How was the code “cracked”?

• We must first consider how genetic information is used by the cell.

• In higher organisms (eukaryotes) the DNA is in the nucleus and the protein is made in the cytoplasm there must be an intermediate.

• Messenger RNA (mRNA) moves from the nucleus to the cytoplasm and carries the genetic code.

The Central Dogma

• Francis Crick proposed the idea that genetic information moves in one direction and called this

the central dogma of molecular genetics.

replication

DNA RNA Protein transcription translation


• Cells can be broken open and the elements needed for protein synthesis can be isolated. When RNA is added, the protein encoded by that RNA is made.

• Artificial RNA can be made in the test tube and added to this system.


• Cells can be broken open and the elements needed for protein synthesis can be isolated. When RNA is added, the protein encoded by that RNA is made.

• http://profiles.nlm.nih.gov/JJ/Views/Exhibit/documents/codeoflife.html

• Artificial RNA can be made in the test tube and added to this system. This work was done by Marshall Nirenberg and Har Gobind Khorana

• http://www.ucs.mun.ca/~c64dcp/Khorana.html

Nirenberg

Khorana


• Nirenberg made simple RNA with the sequence:

UUUUUUUUUUUUUUUUUUUUU

• When he put this into the test tube, he found that the protein made was a string of one type of amino acid, phenylalanine, joined together.

Therefore the codon UUU (or TTT in DNA),

encodes phenylalanine.


Similarly, RNA with the sequence:CCCCCCCCCCCCCCCCCCCCCC encodes a protein that is all proline.

AAAAAAAAAAAAAAAAAAAAencodes a protein that is all lysine.


• Khorana made less simple RNA with the sequence:

UGUGUGUGUGUGUGUGUGUGU

• When he put this into the test tube, he found that the protein made was a string of two alternating amino acids, valine and cysteine.


TGT = CysGTG = Val

• By successively more sophisticated experiments of this type, the amino acids specified by most of the 61 amino acid encoding triplets were identified.

• Final confirmation required experiments with another type of RNA, transfer RNA (tRNA), which is the subject of the next lecture.


the genetic code each amino acid is specified by a triplet of

Documents

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amino acids

nonsense codons act

phosphoric acid

aminoacids e

isa purine orpyrimidineca

usuallyonly importantwhether