The Genetic Code

1. The Genetic Code Chapter 15 胡攸乔 200431060050

2. The 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

4. Outline • The Code Is Degenerate • Three Rules Govern the Genetic Code • Suppressor Mutations Can Reside in the Same or a Different Gene • The Code is Nearly Universal

5. Topic 1 The Code Is Degenerate • concepts Many amino acides are specified by more than one codon, the phenomenon called degeneracy. codons specifying the same amino acid are synonyms

7. The Code Is Degenerate 1 When the first two nucletides are identical, the third nucleotide can be either cytosine or uracil and the codon will still code for the same amino acid, C and U,A and G can be interchangeable • Not all degeneracy is based on equivalence of the first two nucleotides leucine UUA、UUG、CUU、CUC、CUA、CUG 3 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 acids in their proteins

8. Perceiving Order in the Makeup of the Code 1 The code evolved in such a way as to minimize the deleterious effects of mutations Examples: a: Mutation in the first position of a codon will often give a similar amino acid. b:Codons with pyrimidines in the second position specify mostly hydrophobic amino acids,with purines in the second position correspond mostly to polar amino acids. c:Change in the third position rarely will a different amino acid be specified,even a transversion mutation.

9. Perceiving Order in the Makeup of the Code 2 degeneracy may function as a safety mechanism to minimize errors in reading of such condons. • Whenever the first two positions of a codon are both occupied by G or C, each of the four nucleotides in the third position specifies the same amino acid. • On the other hand, Whenever the first two positions of the codon are both occupied by A or U, the identity of the third nucleotide does make a difference. Since G:C pairs are stronger than A:U ones

10. Wobble 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 codons • Inosine is found in the anticodon loop as a fifth base

11. Wobble in the Anticodon In 1966, Francis Crick • Wobble concept • The base at the 5’ end of the anticodon is no as spatically confined as the other two allowing it to form hydrogen bonds with any of several based located at the 3’ end of a codon, with pairing restrict to those on the right table Base in Anticodon Base in Codon

12. 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

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

14. Three Codons Direct Chain Termination UAA,UAG,UGA are read not by special tRNA, but by specific proteins known as release factors(RF1 and RF2 in bacteria and eRF1 in eukaryotes). Release factors enter the A site of the ribosome and trigger hydrolysis of the peptidyl-tRNA occupying the P site, resulting in the release of the newlysynthesized protein.

16. How the Code Was Cracked • One 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 synthesis • Seeking for more details in this web site http://nobelprize.org/educational_games/medicine/gene-code/history.html

17. Stimulation of Amino Acid Incorporation by Synthetic mRNAs Polynucleotide 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

18. Poly-U Code for Polyphenylalanine • A Clever Experiment They carried out a series of experiments to see what amino acid a particular nucleotide template gave rise to. Strands of template with a known combination of nucleotides were run in the "cell-free" system. • They made a very simple nucleic acid, composed of a chain of only one single, repeated letter – the nucleotide uracil, or U. Using this nucleic acid, the system produced a protein that also contained a single letter, but now written in the protein language: the amino acid F, phenylalanine. By showing that a strand of U triplets was indeed the template for the amino acid phenylalanine they cracked the first letter of the code. • This was the result Nirenberg presented in Moscow. While he was at the conference he got a phone call from Matthaei (still working at the lab back home) who told him that CCC was probably the template for the amino acid proline, P. The experiment which used uracil (U) as a template produced a protein entirely made up of the amino acid phenylalanine (F). The first letter of the genetic code was hence identified.

19. Mixed Copolymers Allowed Additional Codon 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

20. Transfer RNA Binding to Defined Trinucleotide Codons • Specific amino-acyl-tRNA can bind to ribosome-mRNA complexes • The addition of trinucleotide results in corresponding amino-acyl-tRNA attachment.

21. The addition of trinucleotide results in corresponding amino-acyl-tRNA attachment. • tri- and tetra-nucleotides could be polymerized into polymers with repeating sequences that could be used in cell-free in vitro translation assays . • In the case of trinucleotides, three polypeptides will be synthesized, each of which is a homopolymer of a single amino acid

22. Topic 2 Three Rules Govern The Genetic Code • Codons are read in a 5’ to 3’ direction. • Codons are nonoverlapping and the message contains no gaps. • The message is translated in a fixed reading frame, which is set by the initiation codon.

23. Three Kinds of Point Mutations Alter the Genetic Code • missense mutation:an alteration that changes a condon specific for one amino acid to a codon specific for another amino acid . • nonsense/stop mutation: an alteration causing a change to a chain-termination codon. • Frameshift mutation: insertions or deletions of one or a smal number of base pairs that alter the reading frame.

24. Three Kinds of Point Mutations

25. Ala Ala Ser Cys Met Leu His Ala Ala Ala 5’-GCU GCU AGC UGC AUG CUG CAU GCU GCU GCU-3’ Genetic Proof that the Code is Read in Units of Three • Experiment by Francis Crick, Sydney Brenner 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.

26. Topic 3 Suppressor Mutations can Reside in The Same or a Different Gen • Concepts • Reverse mutations:change an altered nucleotide sequence back to its original arrangement. • Suppressor mutations:suppress the change due to mutation at site A by producing an additional genetic change at site B. Intragenic suppressionoccurring within the same gene as the original mutation,but at a different site missense mutation Intergenic suppression occurring in another gene

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

28. Suppression of frameshift Suppressor Mutations can Reside in The Same or a Different Gene

29. Intergenic Suppression Involves Mutant tRNAs Suppressor genes do not act by changing the nucleotide sequence of a mutant gen.Instead, they change the way the mRNA template is read. nonsense mutations :A mutation in the anticodon of tRNA that alters the anticodon so it is now complementary to a nonsense codon allowing the tRNA to insert its cognate amino acid at this nonsense codon during translation.

30. Intergenic Suppression Involves Mutant tRNAs If a mutation occurs in the DNA that changes the AAG codon in the mRNA to UAG, the UAG codon will be read as a stop signal and the translation product will be a truncated (short) usually nonfunctional polypeptide.

31. Nonsense Suppressors also Read Normal Termination Signals • The act of nonsense suppression can be viewed as a competition between the suppressor tRNA 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

32. Proving the Vality of the Genetic Code • A classic and instructive experiment in 1966 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

33. Topic4 The Code is Nearly Universal • 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.

34. The Code is Nearly Universal

35. The Code is Nearly Universal