The Genetic CodeThe Genetic CodeChapter 15Chapter 15

胡攸乔 胡攸乔 200431060050200431060050

The central dogmaThe central dogma

Translation key words

tRNA(transfer RNAs): the special adaptor molecules mediate the translation of genetic

Codons: Three consecutive nucleotides

The total number is 64 while the amino acids number is 22

OutlineOutline

The Code Is DegenerateThe Code Is Degenerate

Three Rules Govern the Genetic CodeThree Rules Govern the Genetic Code

Suppressor Mutations Can Reside in Suppressor Mutations Can Reside in the Same or a Different Genethe Same or a Different Gene

The Code is Nearly UniversalThe Code is Nearly Universal

Topic 1 The Code Is DegenerateTopic 1 The Code Is Degenerate

conceptsconcepts Many amino acides are specifiMany amino acides are specifi

ed by more than one codon,ed by more than one codon, the phenomenon called the phenomenon called dedegeneracygeneracy..

codons specifying the same acodons specifying the same amino acid are mino acid are synonymssynonyms

The Code Is DegenerateThe Code Is Degenerate

1 When the first two nucletides are identical, the third n1 When the first two nucletides are identical, the third nucleotide can be either cytosine or uracil and the coucleotide can be either cytosine or uracil and the codon will still code for the same amino acid, C and U,don will still code for the same amino acid, C and U,A and G can be interchangeableA and G can be interchangeable

22 Not all degeneracy is based on equivalence of the firNot all degeneracy is based on equivalence of the first two nucleotidesst two nucleotides

leucine UUA 、 UUG 、 CUU 、 CUC 、 CUA 、 CUG

3 There can be great variation in the AT/GC ratios in the DNA of vario3 There can be great variation in the AT/GC ratios in the DNA of various organisms without correspondingly large changes in the relus organisms without correspondingly large changes in the relative proportion of amino acids in their proteinsative proportion of amino acids in their proteins

Perceiving Order in the Makeup of the CodePerceiving Order in the Makeup of the Code

1 The code evolved in such a way as to minimize 1 The code evolved in such a way as to minimize the deleterious effects of mutationsthe deleterious effects of mutations

Examples: a: Mutation in the first position of a codon will often give a simiExamples: a: Mutation in the first position of a codon will often give a similar amino acid.lar amino acid.

b:Codons with pyrimidines in the second position specify mostly b:Codons with pyrimidines in the second position specify mostly hydrophobic amino acids,with purines in the second position corresphydrophobic amino acids,with purines in the second position correspond mostly to polar amino acids.ond mostly to polar amino acids.

c:Change in the third position rarely will a different amino acid be c:Change in the third position rarely will a different amino acid be specified,even a transversion mutation specified,even a transversion mutation ..

Perceiving Order in the Makeup of the CodePerceiving Order in the Makeup of the Code

22 degeneracy degeneracy may function as a safety mechanism to minimize emay function as a safety mechanism to minimize errors in reading of such condons.rrors in reading of such condons.

4.4. Whenever the first two positions of a codon are both Whenever the first two positions of a codon are both occupied by G or C, each of the four nucleotides in toccupied by G or C, each of the four nucleotides in the third position specifies the same amino acid.he third position specifies the same amino acid.

5.5. On the other hand, Whenever the first two positions On the other hand, Whenever the first two positions of the codon are both occupied by A or U, the identitof the codon are both occupied by A or U, the identity of the third nucleotide does make a difference.y of the third nucleotide does make a difference.

Since G:C pairs are stronger than A:U onesSince G:C pairs are stronger than A:U ones

Wobble in the AnticodonWobble in the Anticodon

Problem If there is a specific tRNA for every codon , at least 61different tRNAs would exist.) while the truth is Some tRNA could recognize several different codonsInosine is found in the anticodon loop as a fifth base

Wobble in the AnticodonWobble in the Anticodon

In 1966, Francis Crick In 1966, Francis Crick Wobble conceptWobble conceptThe base at the 5’ end of the The base at the 5’ end of the anticodon is no as spatically anticodon is no as spatically confined as the other two allconfined as the other two allowing it to form hydrogen boowing it to form hydrogen bonds with any of several based nds with any of several based located at the 3’ end of a colocated at the 3’ end of a codon, with pairing restrict to tdon, with pairing restrict to those on the right tablehose on the right table

Base in Anticodon Base in Codon

GG

CC

AA

UU

II

U or CU or CGGUUA or GA or GA,U,or CA,U,or C

We can see the 5’ end of anticodon is freer to wobble than is the fully stackerd base at the 3’end of the anticodon

Wobble base pairing ribose-ribose distance are close to those of A:U G:C base pairs

Three Codons Direct Chain TerminatioThree Codons Direct Chain Terminationn

UAA,UAG,UGA UAA,UAG,UGA are read not by special tRNA, but are read not by special tRNA, but by specific proteins known as release factors(Rby specific proteins known as release factors(RF1 and RF2 in bacteria and eRF1 in eukaryotes).F1 and RF2 in bacteria and eRF1 in eukaryotes).

Release factors enter the A site of the ribosome aRelease factors enter the A site of the ribosome and trigger hydrolysis of the peptidyl-tRNA occund trigger hydrolysis of the peptidyl-tRNA occupying the P site, resulting in the release of the pying the P site, resulting in the release of the newlysynthesized protein.newlysynthesized protein.

How the Code Was CrackedHow the Code Was CrackedOne of the great achievement in the molecular biology( turn to chapter 2 for more details)

The use of artificial mRNAs and the availability of cell-free systems for carrying out protein synthesisSeeking for more details in this web site http://nobelprize.org/educational_games/medicine/gene-code/history.html

Stimulation of Amino Acid Incorporation Stimulation of Amino Acid Incorporation by Synthetic mRNAsby Synthetic mRNAs

Polynucleotide phosphorylase reactionPolynucleotide phosphorylase reaction[XMP]n + XDP [XMP]n+1 + P

Extracts from E. coli cells can incorporate amino acids into proteins.After several minutes the synthesis came to a stop because the degradation of mRNA. The addition of fresh mRNA to extracts caused an immediate resumption of synthesis.This led the scientist an opportunity to elucidate the nature of the code using synthetic RNA

Poly-U Code for PolyphenylalaninePoly-U Code for Polyphenylalanine

A Clever ExperimentA Clever Experiment They carried out a series o They carried out a series of experiments to see what amino acid a particulaf experiments to see what amino acid a particular nucleotide template gave rise to. Strands of ter nucleotide template gave rise to. Strands of template with a known combination of nucleotidemplate with a known combination of nucleotides were run in the "cell-free" system.s were run in the "cell-free" system.They made a very simple nucleic acid, composeThey made a very simple nucleic acid, composed of a chain of only one single, repeated letter – td of a chain of only one single, repeated letter – the nucleotide uracil, or U. Using this nucleic acid,he nucleotide uracil, or U. Using this nucleic acid, the system produced a protein that also contain the system produced a protein that also contained a single letter, but now written in the protein ed a single letter, but now written in the protein language: the amino acid F, phenylalanine. By slanguage: the amino acid F, phenylalanine. By showing that a strand of U triplets was indeed the howing that a strand of U triplets was indeed the template for the amino acid phenylalanine they template for the amino acid phenylalanine they cracked the first letter of the code.cracked the first letter of the code.This was the result Nirenberg presented in MoscThis was the result Nirenberg presented in Moscow. While he was at the conference he got a phoow. While he was at the conference he got a phone call from Matthaei (still working at the lab bane call from Matthaei (still working at the lab back home) who told him that CCC was probably tck home) who told him that CCC was probably the template for the amino acid proline, P.he template for the amino acid proline, P.

The experiment which usThe experiment which used uracil (U) as a template ed uracil (U) as a template produced a protein entirelproduced a protein entirely made up of the amino aciy made up of the amino acid phenylalanine (F). The fird phenylalanine (F). The first letter of the genetic code st letter of the genetic code was hence identified.was hence identified.

Mixed Copolymers Allowed Additional CodoMixed Copolymers Allowed Additional Codon Assignmentsn Assignments

Poly-AC contain 8 codons: CCC, CCA, CAC, ACC, CAA, ACA, AAC, and AAA.They code for Asp, Glu, His, Thr & Pro (CCC), Lys (AAA). The proportions of the 8 codons incorporated into polypeptide products depend on the A/C ratio

Transfer RNA Binding to Defined TrinuTransfer RNA Binding to Defined Trinucleotide Codonscleotide Codons

Specific amino-acyl-tRNA can bind to ribosome-mRNA complexesThe addition of trinucleotide results in corresponding amino-acyl-tRNA attachment.

The addition of trinucleotide results in co

rresponding amino-acyl-tRNA attachment.tri- and tetra-nucleotides could be polytri- and tetra-nucleotides could be polymerized into polymers with repeating semerized into polymers with repeating sequences that could be used in cell-free quences that could be used in cell-free iin vitron vitro translation assays . translation assays .In the case of trinucleotides, three polypIn the case of trinucleotides, three polypeptides will be synthesized, each of whiceptides will be synthesized, each of which is a homopolymer of a single amino ach is a homopolymer of a single amino acidid

Topic 2 Three Rules Govern The Topic 2 Three Rules Govern The Genetic CodeGenetic Code

1.1. Codons are read in a Codons are read in a 5’ to 35’ to 3’ direction.’ direction.

2.2. Codons are Codons are nonoverlappingnonoverlapping and the messag and the message contains no gaps.e contains no gaps.

3.3. The message is translated in a The message is translated in a fixed reading ffixed reading framerame, which is set by the , which is set by the initiation codoninitiation codon..

Three Kinds of Point Mutations Alter Three Kinds of Point Mutations Alter the Genetic Codethe Genetic Code

missense mutation:missense mutation:anan alteration that change alteration that changes a condon specific for one amino acid to a cods a condon specific for one amino acid to a codon specific for another amino acid .on specific for another amino acid .nonsense/stop mutationnonsense/stop mutation: : an alteration causian alteration causing a change to a chain-termination codon.ng a change to a chain-termination codon.Frameshift mutationFrameshift mutation:: insertions or deletions insertions or deletions of one or a smal number of base pairs that alteof one or a smal number of base pairs that alter the reading frame.r the reading frame.

Three Kinds of Point MutationsThree Kinds of Point Mutations

Genetic Proof that the Code is Read in Genetic Proof that the Code is Read in Units of ThreeUnits of Three

Experiment by Francis Crick, Sydney BreExperiment by Francis Crick, Sydney Brenner and coworkers involving nner and coworkers involving bacteriophage T4 shows that

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

5’-GCU GCU AGC UGC AUG CUG CAU GCU GCU GCU-3’

Ala Ala Ser Cys Met Leu His Ala Ala Ala

Topic 3 Suppressor Mutations can Topic 3 Suppressor Mutations can Reside in The Same or a Different GenReside in The Same or a Different Gen

ConceptsConcepts Reverse mutations:Reverse mutations: change an altered nucleotide schange an altered nucleotide s

equence back to its original arrangement.equence back to its original arrangement. Suppressor mutations:Suppressor mutations: suppress the change due to suppress the change due to

mutation at site A by producing an additional genetmutation at site A by producing an additional genetic change at site B.ic change at site B.

Intragenic suppressionIntragenic suppression occurring within the same gene as the original mutation,but at a different site missense mutation

Intergenic suppression Intergenic suppression occurring in another gene

Suppressor genes:genes that cause suppression of mutations in other genes.

Suppressor Mutations can Reside in Suppressor Mutations can Reside in The Same or a Different GeneThe Same or a Different Gene

Suppression of frameshift

Intergenic Suppression Involves Mutant tRNAsIntergenic Suppression Involves Mutant tRNAs

Suppressor genes do not act by changing the nucleotide Suppressor genes do not act by changing the nucleotide sequence of a mutant gen.Instead, they change the wsequence of a mutant gen.Instead, they change the way the mRNA template is read.ay the mRNA template is read.

nonsense mutationsnonsense mutations :A mutation in the anticod :A mutation in the anticodon of tRNA that alters the anticodon so it is noon of tRNA that alters the anticodon so it is now complementary to a nonsense codon allowiw complementary to a nonsense codon allowing the tRNA to insert its cognate amino acid at ng the tRNA to insert its cognate amino acid at this nonsense codon during translation. this nonsense codon during translation.

Intergenic Suppression Involves Mutant tRNIntergenic Suppression Involves Mutant tRNAsAs

If a mutation occurs in the DIf a mutation occurs in the DNA that changes the AAG NA that changes the AAG codon in the mRNA to UAcodon in the mRNA to UAG, the UAG codon will be rG, the UAG codon will be read as a stop signal and tead as a stop signal and the translation product wilhe translation product will be a truncated (short) usl be a truncated (short) usually nonfunctional polypually nonfunctional polypeptide. eptide.

Nonsense Suppressors also Read Normal Termination Signals

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

E. coli can tolerate the misreading of the UAG stop condon UAG but not UAA because UAG is used infrequently as a chain-terminating codon at the end of open

Proving the Vality of the Genetic CodeProving the Vality of the Genetic Code

A classic and instructive experiment in 1A classic and instructive experiment in 1966 helped to validate the genetic code.966 helped to validate the genetic code.

NH2—Lys Ser Pro Ser Leu Asn Alu—COOH

5’ —AAA AGU CCA UCA CUU AAU GC—3’

5’ —AAA GUC CAU CAC UUA AUG GC—3’

NH2--:Lys Val His His Leu Met Ala—COOH

Topic4 The Code is Nearly UniversTopic4 The Code is Nearly Universalal

Mitochondrial tRNAs are unusual in the way that they decode mitochondrial messages.Only 22 tRNAs are present in mammalian mitochondria. The U in the 5’ wobble position of a tRNA is capable of recognizing all four bases in the 3’ of the codon.

The Code is Nearly UniversalThe Code is Nearly Universal

The Code is Nearly UniversalThe Code is Nearly Universal