０４级生物科学 2 组 张瑶心 ２００４３１２５０００４ MOLECULAR BIOLOGY Chapter 15 The Genetic Code

Molecular Biology of the Gene, 5/E --- Watson et al. (2004) Part I: Chemistry and Genetics Part II: Maintenance of the Genome Part III: Expression of the Genome Part IV: Regulation Part V: Methods

Chapter 15 The Genetic Code Part III: Expression of the Genome This part concerned with one of the greatest challenges in understanding the gene － how the gene is expressed.

Chapter 15 The Genetic Code Part III: Expression of the Genome Ch 12: Mechanisms of transcription Ch 13: RNA splicing Ch 14: Translation Ch 15: The genetic code

Chapter 15 The Genetic Code

Topic1: The code is degenerate Topic2: Three rules govern genetic code Topic3: Suppressor mutations can reside in the same or a different gene Topic4: The code is nearly universal

Topic1: The code is degenerate There are 64 permutations of codons. One of the most striking features of code : Each of the 61 triplets can specify an amino acid, the remaining 3 triplets are chain-terminating signals.

Topic1: The code is degenerate Degeneracy( 简并性 ): A phenomenon that Many amino acids are specified by more than one codon. Synonyms( 同义密码子 ): Codons specifying the same amino acid. 64 kind of condons 20 kind of amino acids more than one codon can specify the same amino acid

Topic1: The code is degenerate Order in the Makeup of the Code 1.The code evolved in such a way as to minimize the deleterious effects of mutations. eg : 1. Mutations in the first position of a codon will often give a similar (if not the same) amino acid. 2. A chage in the second position of a codon will usually replace one amino acid with a very similar one. 3. If a codon suffers a transition mutation in the third position, rarely will a different amino acid be specified.

Topic1: The code is degenerate 2. 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. Whenever the first two positions of a codon are both occupied by A or U, the identity of the third nucleotide does make a difference. REASON : G:C bps are stronger than A:U bps, mismatches in pairing the third codon base are often tolerated if the first two positions make strong G:C bps. Order in the Makeup of the Code

Topic1: The code is degenerate Wobble concept ( 摆动理论 ): Wobble concept ( 摆动理论 ): The base at the 5’ end of the anticodon is not as spatially confined as the other two, allowing it to form hydrogen bonds with any of several bases located at the 3’ end of a codon 1. One tRNA anticodon can recognize several different codons. 2. Anticodon have 5 kind of bases: I (inosine 次黄嘌呤 ), G, C, A, U. observations: Explainations:

Topic1: The code is degenerate Figure 15-1 Codon-anticodon pairing of two tRNA Leu moleculars Figure 15-1 Codon-anticodon pairing of two tRNA Leu moleculars The structure of the tRNA

Topic1: The code is degenerate Base in 5 ’ Anticodon Base in 3 ’ Codon G U or C C G A U U A or G I A, U, or C G U or C C G A U U A or G I A, U, or C Table 15-2 Pairing Combinations with the Wobble Concept The pairings permitted by the wobble rules are those that give ribose-ribose distances close to that of the standard A:U or G:C bps.

Topic1: The code is degenerate Figure 15-2 Wobble base pairing Figure 15-2 Wobble base pairing The ribose-ribose distances for the wobble pairs are close to those of A:U or G:C base pairs

Topic1: The code is degenerate Wobble Concept The molecular mechanism of Wobble Concept In tRNA, the 3 anticodon bases -as well as the following 3’ bases in the anticodon loop- all point in roughly the same direction, with their exact conformations largely determined by stacking interaction s between the flat surfaces of the bases( 由碱基平面 之间的堆积作用所决定 ).

Topic1: The code is degenerate The first 5’ anticodon base is at the end of the stack of bases and is perhaps less restricted in its movement than the other two anticodon bases- hence, wobble in the third (3’)position of the codon.

Topic1: The code is degenerate Figure 15-3 Structure of yeast tRNA(Phe) The adjacent base is always a bulky modified purine residue. The adjacent base

Topic1: The code is degenerate There are 3 codons ( UAA, UAG, UGA ) do not correspond to any amino acid. Instead, they signify chain termination. These chain-termination codons are read not by special tRNA but by specific proteins known as release factors (RF). 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 newly synthesized protein.

Topic1: The code is degenerate ( 略 ) The process of cracking the genetic code ( 略 ) Page (chapter 15) Page 35 (chapter 2) 1. Stimulation of amino acid incorporation by synthetic mRNAs 2. Poly-U codes for polyphenylalanine 3. Mixed copolymers allowed additional codon assignments 4. Transfer RNA binding to defined trinucleotide codons 5. Codon assignments from repeating copolymers

Topic1: The code is degenerate Figure 15-4 Polynucleotide phosphorylase reaction How the RNA is synthesized? [XMP]n + XDP = [XMP]n+1 + P How the RNA is synthesized? [XMP]n + XDP = [XMP]n+1 + P

Topic1: The code is degenerate Figure 15-5 Preparing oligo-ribonucleotides

1.Codons are read in a 5’ to 3’ direction. Topic2: Three rules govern genetic code 2.Codons are nonoverlapping and the message contains no gaps. ( This means that successive codons are represented by adjacent trinucleotides in register.)

Topic2: Three rules govern genetic code 3.The message is translated in a fixed reading frame, which is set by the initiation codon. (Translation stars at an initiation codon, which is located at the 5’ end of the protein-coding sequence. Because codons are nonoverlapping and consist of three consecutive nucleotides, a stretch of nucleotides could be translated in principle in any of three reading frames. It is the initiation codon that dictates which of the three possible reading frames is used.)

Topic2: Three rules govern genetic code Three kinds of point mutations alter the genetic code. 1. Missense Mutation ( 错义突变 ): An alteration that changes a codon specific for one amino acid to a codon specific for another amino acid. 2. Nonsense Mutation ( 无义突变 )/ Stop Mutation ( 终止 突变 ): When it arises in the middle of a genetic message, an incomplete polypeptide is released from the ribosome owing to premature chain termination. ( The size of the incomplete polypeptide chain depends on the location of the nonsense mutation.) A more drastic effect result from an alternation causing a change to a chain-termination codon.

Topic2: Three rules govern genetic code 3. Frameshift mutation ( 移码突变 ): Insertions or deletions of one or a small number of base pairs that alter the reading frame １ １. The insertion (or deletion) of a single base drastically alters the coding capacity of the message not only at the site of the insertion but for the remainder of the messenger as well. 2. Likewise, the insertion (or deletion) of two bases would have the effect of throwing the entire coding sequence, at and downstream of the insertions, into a different reading frame.

Topic2: Three rules govern genetic code 3. The insertion (or deletion) of two bases will obviously alter the stretch of message, at and between the three inserted bases. But because the code is read in units of three, messenger RNA downstream of the three inserted bases will be in its proper reading frame and hence, completely unaltered.

Topic3: Suppressor mutations can reside in the same or a different gene Reverse (back) Mutation ( 逆转突变 ) : An alternation which change an altered nucleotide sequence back to its original arrangement. Suppressor Mutation ( 抑制突变 ) ： An alternation occurring at different locations on the chromosome that suppress the change due to a mutation at site A by producing an additional genetic change at site B. The effects of harmful mutations can be reversed by a second genetic change in the following two ways:

Topic3: Suppressor mutations can reside in the same or a different gene Two type of suppressor mutation : 1. Intragenic suppression ( 基因内抑制 ) ： Suppressor mutation occurring within the same gene as the original mutation, but at a different site in this gene. 2. Intergenic suppression ( 基因间抑制 ) ： Suppressor mutation occurring in different gene to the original mutation. Suppressor genes ( 抑制基因 ): Genes that cause suppression of mutations in other genes.

Topic3: Suppressor mutations can reside in the same or a different gene Both of the two types of suppression work by causing the production of good (or partially good) copies of the protein made inactive by the original harmful mutation. EG: If the first mutation caused the production of inactive copies of one of the enzymes involved in making arginine, then the suppressor mutation allows arginine to be made by restoring the synthesis of some good copies of the same enzyme. The mechanisms by which intergenic and intragenic suppressor mutations cause the resumption of the synthesis of good proteins are completely different.

Topic3: Suppressor mutations can reside in the same or a different gene Intragenic suppression ( 基因内抑制 ) EG 1: missense mutation Its effect can sometimes be reversed through an additional missense mutation in the same gene. In such cases, the original loss of enzymatic activity is due to an altered three-dimensional configuration resulting from the presence of an incorrect amino acid in the encoded protein sequence. An second missense mutation in the same gene can bring back biological activity if it somehow restores the original configuration around the functional part of molecule.

Topic3: Suppressor mutations can reside in the same or a different gene Figure 15-6 Suppression of frameshift mutations Intragenic suppression ( 基因内抑制 ) EG 2: frameshift mutation

Topic3: Suppressor mutations can reside in the same or a different gene Intergenic suppression involves mutant tRNAs. Suppressor genes do not act by changing the nucleotide sequence of a mutant gene. Instead, they change the way the mRNA template is read. EG: Mutant tRNA suppress the effect of nonsense mutations in protein-coding genes. There 3 genes that suppress the UAG codon (one of the 3 stop codons). Each of them inserts one different amino acid at the nonsense position. In each of the three UAG suppressor mutants, the anticondon of a tRNA species specific for one of these amino acids has altered.

Topic3: Suppressor mutations can reside in the same or a different gene Figure 15-7 a

Topic3: Suppressor mutations can reside in the same or a different gene Figure 15-7 b

Topic3: Suppressor mutations can reside in the same or a different gene The codons corresponding to the mutant tRNA can be read normally as well. Two different mechanisms. (Page ) 1. UAG stop codon There are 3 separate genes code for the tRNA (Tyr) independently. The two minor tyrosine tRNA species act to suppress the nonsense codon in mRNA. The major tyrosine tRNA species act to read the UGA stop codon normally. 2. UGA stop codon A mutant form of tRNA (Trp) can recognize and suppress the UGA stop condon while retaining its capacity to read UGG (Trp) codons.

Topic3: Suppressor mutations can reside in the same or a different gene The suppression of nonsense mutation 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-through or polypeptide chain termination will occur, depending on which (the mutant tRNA or the release factor ) arrives first.

Topic4: The code is nearly universal In certain subcellular organelles, the genetic code is in fact slightly different from the standard code. The genetic code is universal or conservative In most of the cells. BUT … … Sequences of the regions known to specify proteins have revealed the differences between the standard and mitochondrial genetic codes

Topic4: The code is nearly universal Table 15-6 Genetic Code of Mammalian Mitochondria Table 15-6 Genetic Code of Mammalian Mitochondria

Topic4: The code is nearly universal Exceptions to the “universal” code are not limited to mitochondria but also found in several prokaryotic genomes and in the nuclear genomes for certain eukaryotes.

S u m m a r y 1. In the ”universal” genetic code used by every organism from bacteria to humans,61 codons signify specific amino acids; the remaining 3 are chain-termination codons. 2. A given tRNA sometimes specifically recognize several codons. This ability arises from wobble in the base at the 5’ end of the anticodon. 3. The genetic code is subject to 3 principal rules. 4.There are 3 kinds of point mutations which can alter the genetic code: Missense Mutation, Nonsense Mutation (Stop Mutation) and Frameshift mutation. 5. A slightly different genetic code is utilized in mitochondria and principal genomes of certain prokaryotes and protazoa.