CHAPTER 15 The Genetic Code CHAPTER 15 The Genetic Code DNARNA Protein Replication TranscriptionTranslation

1.Genetic information transfer from polynucleotide chain into polypeptide chain. 2.Take place in ribosomes. 3.tRNAs recognize codons. 1.Genetic information transfer from polynucleotide chain into polypeptide chain. 2.Take place in ribosomes. 3.tRNAs recognize codons.

Which codons are responsible for specifying which amino acids? What are the rules that gover their use? Which codons are responsible for specifying which amino acids? What are the rules that gover their use? Questions

Outline The code is degenerate Three rules govern the genetic code Suppressor mutations can reside in the same or different gene The code is nearly universal The code is degenerate Three rules govern the genetic code Suppressor mutations can reside in the same or different gene The code is nearly universal

Topic 1 THE CODE IS DEGENERATE Topic 1 THE CODE IS DEGENERATE

Many amino acids are specified by more than one codon- degeneracy. Codons specifying the same amino acid are called synonyms. Many amino acids are specified by more than one codon- degeneracy. Codons specifying the same amino acid are called synonyms.

Often, when the first two nucleotides are identical, the third nucleotide can be either C or U without changing the code. A and G at the third position are interchangeable as well.

Transition in the third position of a codon specifies a same amino acid. Transversion in this position changes the amino acid about half the time.

CUG CUC Codon-anticodon pairing of two tRNA Leu molecules

AT/GC ratios proportion of amino acids Code degeneracy explains how there can be great variation in the AT/GC ratios in the DNA of various organisms without large changes in the proportion of amino acids in their proteins.

Perceiving Order in the Makeup of the Code 1.The genetic code evolved in such a way as to minimize the deleterious effects of mutations. 2.Code degeneracy may serve as a safety mechanism to minimize errors in the reading of codons.

1.Mutation in the first position of a codon will often give a similar amino acid. 2.The second position of a codon: Pyrimidines-hydrophobic amino acids Purines-polar amino acids 3.A transition mutation in the third position,rarely will a different amino acid. Consistency 1

If the first two positions are both occupied by G or C, each of the four nucleotides in the third position specifies the same amino acid. Consistency 2

Wobble in the Anticodon Question: Is there a specific tRNA for every codon? (If it was true, at least 61 different tRNAs would exist.) Some tRNA could recognize several different codons Inosine is present in the anticodon loop as a fifth base

Inosine inosine adenine Inosine arises through enzymatic modification of adenine

Wobble Concept 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 more than one bases located at the 3’ end of a codon.

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 Pairing Combinations with the Wobble Concept

The Wobble Rules The pairings permitted are those give ribose-ribose distances close to that of the standard A:U or G:C base pairs. The ribose-ribose distances: Purine-purine: too long Pyrimidine-pyrimidine: too short

Wobble base pairing:The ribose-ribose distances for the wobble pairs are close to those of A:U or G:C base pairs

Critical Thinking The wobble concept predicted that at least three tRNAs exist for the six serine codons (UCU, UCC, UCA, UCG, AGU, and AGC). Why?

Why wobble is allowed at the 5’ anticodon The 3-D structure of tRNA shows that the stacking interactions between the flat surfaces of the 3 anticodon bases + 2 followed bases position the first (5’) anticodon base at the end of the stack, thus less restricted in its movements. The 3’ base appears in the middle of the stack, resulting in the restriction of its movements.

The adjacent base The adjacent base is always a bulky modified purine residue.

Three Codons Direct Chain Termination Three codons, UAA, UAG, and UGA signify chain termination. They are not read by tRNAs but by proteins called release factors (RF1 and RF2 in bacteria and eRF1 in eukaryotes).

How the Code Was Cracked See Chapter 2, Page 35: Establishing the Genetic Code The use of artificial mRNAs and the availability of cell-free systems for carrying out protein synthesis began to make it possible to crack the code

Stimulation of Amino Acid Incorporation by Synthetic mRNAs 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. 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.

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

Experimental Results: UUU codes for phenylalanine. CCC codes for proline. AAA codes for lysine. The guanine residues in poly-G firmly hydrogen bond to each other and form multistranded triple helices that do not bind to ribosomes. UUU codes for phenylalanine. CCC codes for proline. AAA codes for lysine. The guanine residues in poly-G firmly hydrogen bond to each other and form multistranded triple helices that do not bind to ribosomes.

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 Such experiment can determine the composition of the codons, but not the order of the three nucleotides. The proportions of the 8 codons incorporated into polypeptide products depend on the A/C ratio Such experiment can determine the composition of the codons, but not the order of the three nucleotides.

Transfer RNA Binding to Defined Trinucleotide Codons A method to order the nucleotides within some of the codons. Specific amino-acyl-tRNA can bind to ribosome-mRNA complexes. The addition of trinucleotide results in corresponding amino- acyl-tRNA attachment. A method to order the nucleotides within some of the codons. Specific amino-acyl-tRNA can bind to ribosome-mRNA complexes. The addition of trinucleotide results in corresponding amino- acyl-tRNA attachment.

Codon Assignments from Repeating Copolymers Organic chemical and enzymatic techniques were used to prepare synthetic polyribonucleotides with known repeating sequences.

Preparing oligo-ribonucleotides

copolymer Codons Recognized Amino Acids Incorporated or Polypeptide Made Codon Assignment (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’

Topic 2 THREE RULES GOVERN THE GENETIC CODE Topic 2 THREE RULES GOVERN THE GENETIC CODE

Three Rules 1.Codons are read in a 5’ to 3’ direction. 2.Codons are nonoverlapping and the message contains no gaps. 3.The message is translated in a fixed reading frame which is set by the initiation codon. 1.Codons are read in a 5’ to 3’ direction. 2.Codons are nonoverlapping and the message contains no gaps. 3.The message is translated in a fixed reading frame which is set by the initiation codon.

Three Kinds of Point Mutations Alter the Genetic Code 1. Missense mutation: An alternation that changes a codon specific for one amino acid to a codon specific for another amino acid.

2. Nonsense or stop mutation: An alternation causing a change to a chain-termination codon.

3. Frameshift mutation: Insertions or deletions of one or a small number of base pairs that alter the reading frame.

Genetic Proof that the Code Is Read in Units of Three A classic 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. A classic 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.

Topic 3 SUPPRESSOR MUTATIONS CAN RESIDE IN THE SAME OR A DIFFERENT GENE Topic 3 SUPPRESSOR MUTATIONS CAN RESIDE IN THE SAME OR A DIFFERENT GENE

1.Reverse (back) mutations: change an altered nucleotide sequence back to its original arrangement. Reverse the harmful mutations by a second genetic change

2.Suppressor mutations: suppress the change due to mutation at site A by producing an additional genetic change at site B. (1) Intragenic suppression (2) Intergenic suppression 2.Suppressor mutations: suppress the change due to mutation at site A by producing an additional genetic change at site B. (1) Intragenic suppression (2) Intergenic suppression

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

Suppressor mutations work by producing good (or partially good) copies of the protein that are made inactive by the original harmful mutation.

Suppression of frameshift mutations

Intergenic Suppression Involves Mutant tRNAs Mutant tRNA genes suppress the effects of nonsense mutations in protein-coding genes. They act by reading a stop codon as if it were a signal for a specific amino acid. Mutant tRNA genes suppress the effects of nonsense mutations in protein-coding genes. They act by reading a stop codon as if it were a signal for a specific amino acid.

Figure 15-7 a

Nonsense Suppressors also Read Normal Termination Signals The act of nonsense suppression is a competition between the suppressor tRNA and the release factor. In E. coli, Suppression of UAG codons is efficient, and suppression of UAA codon average is inefficient. Why??. The act of nonsense suppression is a competition between the suppressor tRNA and the release factor. In E. coli, Suppression of UAG codons is efficient, and suppression of UAA codon average is inefficient. Why??.

Topic 4 THE CODE IS NEARLY UNIVERSAL Topic 4 THE CODE IS NEARLY UNIVERSAL

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

Allow us to directly compare the protein coding sequences among all organisms. Make it possible to express cloned copies of genes encoding useful protein in different host organism. Example: Human insulin ecpression in bacteria) Allow us to directly compare the protein coding sequences among all organisms. Make it possible to express cloned copies of genes encoding useful protein in different host organism. Example: Human insulin ecpression in bacteria) Benefits of the universal codes

However, in certain subcellular organelles, the genetic code is slightly different from the standard code.

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. 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.

Genetic Code of Mammalian Mitochondria

Key points of the chapter 1.“The genetic code is degenerate” What does it mean? What’s the benefits? What’s about the anticodon recognition? How the code was discovered? 2.What are the three roles governing the genetic code? What are the mutations altering genetic code? 1.“The genetic code is degenerate” What does it mean? What’s the benefits? What’s about the anticodon recognition? How the code was discovered? 2.What are the three roles governing the genetic code? What are the mutations altering genetic code?

3.What are suppressor mutations? 4.What are the benefits of the code universality? What’s about the mitochondrial codes and tRNAs? 3.What are suppressor mutations? 4.What are the benefits of the code universality? What’s about the mitochondrial codes and tRNAs?

Thank you~~