Moderator : Dr. M. S. Somannavar

Presenter : Jay prakash sah

Genetic code

Salient features

Mutation

Types of mutation

Mutagens

Clinical correlations

Crick, Brenner et al. experiment - Demonstratethat codons consist of three DNA bases

Marshall Nirenberg and Heinrich J. Matthaei in1961 reported the first three letter code words foreach amino acid

H. Gobind Khorana- Genetic code dictionary andprevious results confirmed

Dictionary that identifies the correspondencebetween a sequence of nucleotide bases and asequence of amino acids

Codon- triplet sequence of nucleotides on themRNA ( A G,C & U)

64 codon

Triplet codons

Non-overlapping

Non-punctuated

Degenerate

Unambiguous

Universal

Wobbling phenomenon

Terminator codon

Initiator codon

Consecutive sequence of three bases onmRNA

Ex:

UUU -Phenylalanine

AUACGAGUC

A U A C G A G U C

1 2 3

overlapping code

A U A C G A G U C

Consecutive or continuous

A particular codon always codes for the sameamino acid

Ex:

UGG - Tryptophan

Same codons are used to code for the same amino acid in all the living organisms.

I

Amber(UAA),Ochre(UAG) & Opal codons(UGA)

UGA - Selenocysteine(Sec)-21(Glutathione peroxidase)

UAG - Pyrolysine-22(Methyl transferase)

(Methanosarcina barkeri)

UGA & UAA- Glutamine( paramecium)

UGA – Trp (mycoplasma)

Garret

AUG - codon for methionine.

In few proteins,

GUG

Change in nucleotide sequence of DNA

Out of every 106 cell divisions-1 mutation occurs

May occur in somatic cells ( aren’t passedoffspring)

May occur in gametes (eggs & sperm) and passedto offspring

Substitution of one base pair by another .

Transition :

purine to purine

Transversions :

purine to pyrimidine

satyanarayana

Mutations: SubstitutionsSubstitution mutation

GGTCACCTCACGCCA

↓

CCAGUGGAGUGCGGU

↓

Pro-Arg-Glu-Cys-Gly

Normal gene

GGTCTCCTCACGCCA

↓

CCAGAGGAGUGCGGU

Codons

↓

Pro-Glu-Glu-Cys-Gly

Amino acids

Mutation Codon Change to DNA

sense strand

Change in

Amino Acid

S (sickle cell

anaemia)

6 GAG to GUG Glu to Val

C (cooley’s

syndrome)

6 GAG to AAG Glu to Lys

GSan Jose 7 GAG to GGG Glu to Gly

E 26 GAG to AAG Glu to Lys

MSaskatoon 63 CAT to TAT His to Tyr

MMilwauki 67 GTG to GAG Val to Glu

OArabia 121 GAA to GTA Glu to Val

One or more base pairs are inserted in or deleted from the DNA .

I . Single base additions ,

Normal gene

GGTCTCCTCACGCCA

↓

CCAGAGGAGUGCGGU

Codons

↓

Pro-Glu-Glu-Cys-Gly

Amino acids

Addition mutation

GGTGCTCCTCACGCCA

↓

CCACGAGGAGUGCGGU

↓

Pro-Arg-Gly-Val-Arg

II. Trinucleotide expansion

Huntingtons’s chorea

CAG repeated 30 to 300 times

III. Duplication.

Ex: Duchene Muscular Dystrophy

i) Large gene deletion

Ex: Alpha-thalassemia

i) Deletion of codon

Ex: Cystic fibrosis

i) Deletion of a single base

Consequences of point mutation

Normal gene

GGTCTCCTCACGCCA

↓

CCAGAGGAGUGCGGU

Codons

↓

Pro-Glu-Glu-Cys-Gly

Amino acids

Substitution mutation

GGTCTTCTCACGCCA

↓

CCAGAAGAGUGCGGU

↓

Pro-Glu-Glu-Cys-Gly

1. SILENT MUTATION

A. Acceptable

Eg: Normal Hemoglobin A molecule ,

67th amino acid in beta chain

GUU(Val)

GCU(Ala)

Hb sydney (functionally normal)

Eg: HbS or sickle cell Haemoglobin

beta chain - 6th position

GAG(glutamine)

GUG( Valine).

Hbs leads to sickle cell anemia.

Incompatible with normal life

Eg : HbM

Distal histidine of alpha chain.

leads to premature termination of the protein

Eg: Thalassemia

Codon 17 of the β-chain

Non-SenceNormal gene

GGTCTCCTCACGCCA

↓

CCAGAGGAGUGCGGU

Codons

↓

Pro-Glu-Glu-Cys-Gly

Amino acids

Substitution mutation

GGTCTCCTCACTCCA

↓

CCAGAAGAGUGAGGU

↓

Pro-Glu-Glu-STOP

Insertion or deletion of base in a gene results

in an altered reading frame of the mRNA

A ‘garbled’ protein - produced.

Normal mRNA AUG UCU UGC AAA……..

Normal Protein Met Ser Cys Lys …….

Deleted U mRNA AUG CUU GCA AA…..

garbled Protein Met Leu Ala ……..

1. physical agents

i) UV light

ii) Ionising radiation e.g. X-ray.

iii) visible light

iv) Heat

2. chemical agents

i) 5-Bromouracil

ii) 2-Aminopurine

iii) Nitrous acid

iv) Acridine dyes.

Spontaneous tautomeric shifts in the basescontribute to replication errors

Ex: Thymine (keto form) shifts to enol form ,which pairs with guanine

Double strand DNA breaks

It penetrates the whole body –

cause both somatic and Germ line mutations

Mutagenic component of sunlight

Can not penetrate beyond the outer layer ofthe skin and - unable cause germ linemutations.

only causes sunburn and skin cancer mainlythrough the formation of pyrimidine dimers

1. Base analog

Bromouracil (structural analog of Thymine)

Enzyme of nucleotide synthesis and DNA synthesistreat Bromouracil as thymine and incorporate itinto DNA , where it pairs with adenine

Attach alkyl groups to nitrogen or oxygen atomsin the bases.

Ex:

Methyl bromide ( used as grain fumigent)

Ethylene oxide (used for sterilization of surgical instruments)

Planar fused ring structures –

Insert themselves between the stacked DNA bases.

Salmonella Typhimurium (his-) are selected

Mutagenesis – indicated by his+ phenotype.

The compound to be tested is mixed withbacteria and introduced into histidine deficientmedium.

Reverse mutation.

Number of colonies -proportional to quantity ofmutagen.

Ethyl methanasulphonate

Spontaneous revertantcolonies

1.Single strand conformation polymorphism(SSCP technique)

2.Heteroduplex Analysis

3.Conformation sensitive gel electrophoresis

4.Protein truncation test(PTT)

5.Denaturing HPLC

CLINICAL CORRELATION

HbS

Sickle cell disease:

Due to missense mutation.

Changes from A to U

GAA or GAG (Glu)

GUA or GUG (Val)

Hemoglobin C disease:

Due to missense mutation.

Changes from G to A

GAA or GAG (Glu)

AAA or AAG (Lys)

In Hb Mckees Rocks

145 ( beta chain)

UAU or UAC (Tyrosine)

UAA or UAG(terminator codon )

Shortening of the beta chain from its normal 146residue to 144 residues

cause overproduction of red blood cells

>36

>200

100-1000

Encoding genes on chromosome 16 (shortarm)

Each cell has 4 copies of the alpha globingene

◦ Loss of ONE gene silent carrier◦ Loss of TWO genes thalassemia minor (trait)◦ Loss of THREE genes Hemoglobin H

◦ Loss of FOUR genes Hemoglobin Barts

Encoding genes on chromosome 11 (short arm)

Each cell contains 2 copies of beta globin gene

“Loss” of ONE gene thalassemia minor (trait)

“Loss” of BOTH gene Thalassemia major

Hb α-codon(142)

Amino acid(142)

α-globin length(residues)

A UAA 141

Contant spring CAA Glutamine 172

Icaria AAA Lysine 172

Seal Rock GAA Glutamate 172

Koya Dora UCA Serine 172

Deletion of phe residue at position 508 inCFTR Gene (chromosome 7)

Causes improper folding of protein

Defective chloride transport ( pancreas,lungtestis & sweat glands)

Harper’s Review of Biochemistry

Lehniger’s principle of Biochemistry

Lippincott’s Illustrated Review of Biochemistry

Text Book of Biochemistry with clinical correlations- Devlin TM

Text Book of Biochemistry by Vasudevan

Text book of biochemistry, satyanarayana

Principle of biochemistry, William H. simmons.

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