Chapter 15

The Genetic Code

The Central Dogma

Which of the triplet codons are responsible for specifying which amino acid and what are the rules that govern their use?

TopicⅠ

The Code is Degenerate

Many amino acids are specified by more than one

codons,a phenomenon called degeneracy.

61 of the 64 triplets specify an amino acid,which the remaining three triplets being chain-terminating signals.

Degeneracy

Codons specifying the same amino acid are synonyms.Often,the degeneracy is based on eqivalence of the first two nucleotides.

The frequent third-place eqivalence of the four NTPs explains how there can be great variation in the AT/GC ratios in the DNA of various organisms without correspondingly large changes in the relative proportion of amino acid in their proteins.

Perceiving Order in the Makeup of the Code

Mutations in the first two positions of a codon will often give a very similar amino acid.

Mutations in the third position rarely will a different amino acid be specified.

If the first two positions are both occupied by G or C,each of the four NTPs specifies the same amino acid.But if the first two positions are both occupied by A or U,the identity of the third NTP does make a difference.

Codon-Anticodon pairing of two tRNA Leu molecules

Wobble in the Anticodon

An anticodon base is not one of the 4 regular ones,but a fifth base,inosine.

Not all combination are possible.The pairing permitted by the wobble rules are those that give ribose-ribose distances close to that of the standard A:U or G:C base pair.

The 3D structure and the stacking interaction between codon-anticodon explains why wobble is not seen in the 5’ of the code but in the 3’.

Pairing Combinations with the Wobble Concept

Pairing Combinations with the Wobble Concept

Base in 5’ Anticodon Base in 3’ CodonBase in 5’ Anticodon Base in 3’ Codon

G U or CC GA UU A or GI A, U, or C

G U or CC GA UU A or GI A, U, or C

Wobble base pairing.

Structure of yeast tRNA(phe)

How the Code is Cracked

The genetic code is cracked through the study of protein synthesis in cell-free extracts.

Addition of new mRNA to an extract depleted of its original messenger component results in the production of new proteins whose amino acid sequences are determined by the externally added mRNA.

The synthetic templates are created using the enzyme polynucleotide phosphorylase,which catalyzes the reaction:

[XMP]n+XDP→[XMP]n+1+Pi

The first step in cracking the genetic code comes when the synthetic poly-U is found to code specifically for poly-phe.

Use of other polyribonucleotides allowed assignment of codons for the various amino acid.

Determination of the exact order of nucleotides in codons subsequently comes from a study of specific trinucleotide-tRNA-ribosome interaction and the use of regular copolymers as messengers.

copolymercopolymer Codons Recognize

d

Codons Recognize

d

Amino Acids Incorporated or

Polypeptide Made

Amino Acids Incorporated or

Polypeptide Made

Codon Assignmen

t

Codon Assignmen

t (CU)” CUC|UCU|CUC… Leucine 5’-CUC-3’

Serine UCU

(UG)” UGU|GUG|UGU… Cystine UGU

Valine GUG

(AC)” ACA|CAC|ACA… Threonine ACA

Histidine CAC

(AG)” AGA|GAG|AGA… Arginine AGA

Glutamine GAG

(AUC)” AUC|AUC|AUC… Polyisoleucine 5’-AUC-3’

(CU)” CUC|UCU|CUC… Leucine 5’-CUC-3’

Serine UCU

(UG)” UGU|GUG|UGU… Cystine UGU

Valine GUG

(AC)” ACA|CAC|ACA… Threonine ACA

Histidine CAC

(AG)” AGA|GAG|AGA… Arginine AGA

Glutamine GAG

(AUC)” AUC|AUC|AUC… Polyisoleucine 5’-AUC-3’

Assignment of Codons Using Repeating Copolymers Built from Two or Three Nucleotides.

TopicⅡ

Three Rules Govern The Genetic Code

What are the Rules

The first rule holds that codons are read in a 5’ to 3’ direction.

The second rule is that codons are nonoverlapping and the message contains no gaps.

The final rule is that the message is translated in fixed reading frame,which is set by the initiation codon.

Read from 5’ to 3’ Direction.

Translation Starts at an Initiation Codon. For Prokaryotes is AUG Codon.

Three Kinds of Point Mutation Alter the Genetic Code

An alteration that changes a codon specific for one amino acid to a codon specific for another amino acid is called a missence mutation.

A more drastic effect results from an alteration causing a change to a chain-termination codon,which is known as a nonsence or stop mutation.

The third kind of point mutation is a frameshift mutation.

In some cases the effect of missence,nonsence,and frameshift mutations can be partially suppressed by extragenic suppressors.

For example,mutant tRNAs read stop codons generated by nonsence mutations as if they were codons for a specific amino acid.

Genetic Proof that the Code is Read in Units of Three

★Aclassic experiment involving bacteriophage T4

★Because the gene could tolerate three insertions but not one or two, the genetic code must be read in units of three.

TopicⅢ

Suppressor Mutations Can Reside in the Same or a Different Gene

The effects of harmful mutations can be reversed by a second genetic change-reverse(back) mutation,which change an altered nucleotide sequence back to its original arrangement.

But more often,the mutations occurring at different locations on the chromosome that suppress the change due to a mutation at A site by producing an additional genetic change at site B.

Such suppressor mutations fall into two main categories:

Occurring within the same gene as the original mutation,but at a different site in this gene is called intragenic suppression.

Those occurring in another gene is called intergenic suppression.

Genes that cause suppression of mutations in other genes are called suppressor genes.

Intergenic Suppression Involves Mutant tRNAs

Mutant tRNA genes are those suppress the effects of nonsense,missense,and frameshift mutations in protein-coding genes.

The mutant tRNAs acts to suppress the nonsense codon in mRNA.(Figure 15-7)

Nonsense Suppressor also Read Normal Termination Signals

The act of nonsense suppression can be viewed as a competition between the suppressor tRNA and the release factor.

When a stop codon comes into the ribosomal A site,either read-though or polypeptide chain termination will occur,depending on which arrives first.

Proving the Validity of the Genetic Code

The code is cracked by means of biochemical ways involving the use of cell-free systems for carrying out protein synthesis.So how do we know definitively that the code is true in living cells?

In phage T4,there is a gene encodes a cell-wall degrading enzyme called lysozyme.

The experimental strategy is to compare the amino acid sequence of the doubly mutant protein with that of wild-type lysozyme.

The solution verifies the validity of the genetic code.

TopicⅣ

The Code is Nearly Universal

The results of large-scale sequencing of genomes have largely confirmed the expected universality of the genetic code.

Sequences of the regions known to specify proteins have revealed the following differences between the standard and mitochondrial genetic codes:

UGA is not a stop signal but codes for tryptophan;

Inter methionine is encoded by both AUG and AUA;

In mammalian mitochondria,AGA and AGG are not arginine codons but specify chain termination;

In fruit fly mitochondria,AGA and AGG are also not arginine codons but specify serine.

Genetic Code of Mammalian mitochondria

Only 22 tRNAs are present in mammalian mitochondria,whereas a minimum of 32 tRNAs are required to decode the “universal” code according to the wobble rules.

Thus,there are tRNAs which have in the 5’ position of their anticodons a U residue.The U residue is able to engage in paring with any of the four nucleotides in the third codon position.

The universality of the code helps us human to create the field of genetic engineering by making it possible to express cloned copies of genes encoding useful protein products in surrogate host organisms,such as the production of human insulin in bacteria.

Thank you!

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郭帆