Structure & concept of gene,Structure & concept of gene,One gene one enzyme One gene one enzyme

hypothesishypothesis,,Genetic CodeGenetic Code

PROTEIN SYNTHESISPROTEIN SYNTHESIS,,Regulation of gene Regulation of gene

expressionexpression

Dr. Madhumita BhattacharjeeAssiatant ProfessorBotany Deptt.P.G.G.C.G. -11,Chandigarh

Definitions of the gene

• The gene is the unit of genetic information that controls a specific aspect of the phenotype.

• The gene is the unit of genetic information that specifies the synthesis of one polypeptide.

1942: George Beadle and Edward Tatum

Studied relationships between genes and enzymes in the haploid fungus Neurospora crassa (orange bread mold).

Discovered that genes act by regulating definite chemical events.

One Gene-One Enzyme Hypothesis

Each gene controls synthesis/activity of a single enzyme.

“one gene-one polypeptide”

1958: George Beadle (Cal Tech) & Edward Tatum (Rockefeller Institute)

Beadle and Tatum (1942)--One Gene, One Enzyme

• Bread mold Neurospora can normally grow on minimal media, because it can synthesize most essential metabolites.

• If this biosynthesis is under genetic control, then mutants in those genes would require additional metabolites in their media.

• This was tested by irradiating Neurospora spores and screening the cells they produced for additional nutritional requirements (auxotrophs).

Beadle and Tatum proposed: “One Gene-One Enzyme Hypothesis”

However, it quickly became apparent that…

1. More than one gene can control each step in a pathway (enzymes can be composed of two or more polypeptide chains, each coded by a separate gene).

2. Many biochemical pathways are branched.

“One Gene-One Enzyme Hypothesis”

“One Gene-One Polypeptide Hypothesis”

Modern Concept of Gene

• Until 1940, the gene was considered as the basic unit of genetic information as defined by three criteria.- Cistron:The unit of function, controlling

the inheritance of one “character” or phenotypic attribute.

– Recon : The unit of recombination– Muton:The unit of mutation.

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Genetic code:

Def. Genetic code is the nucleotide base sequence on DNA ( and subsequently on mRNA by transcription) which will be translated into a sequence of amino acids of the protein to be synthesized.The code is composed of codonsCodon is composed of 3 bases ( e.g. ACG or UAG). Each codon is translated into one amino acid.

The 4 nucleotide bases (A,G,C and U) in mRNA are used to produce the three base codons. There are therefore, 64 codons code for the 20 amino acids, and since each codon code for only one amino acids this means that, there are more than one cone for the same amino acid.

How to translate a codon (see table):This table or dictionary can be used to translate any codon sequence.Each triplet is read from 5′ → 3′ direction so the first base is 5′ base, followed by the middle base then the last base which is 3′ base.

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Examples: 5′- A UG- 3′ codes for methionine 5′- UCU- 3′ codes for serine 5′ - CCA- 3′ codes for proline

Termination (stop or nonsense) codons:Three of the 64 codons; UAA, UAG, UGA do not code for any amino acid. They are termination codes which when one of them appear in mRNA sequence, it indicates finishing of protein synthesis.

Characters of the genetic code: 1- Specificity: the genetic code is specific, that is a specific codon always code for the same amino acid.2- Universality: the genetic code is universal, that is, the same codon is used in all living organisms, procaryotics and eucaryotics.3- Degeneracy: the genetic code is degenerate i.e. although each codon corresponds to a single amino acid,one amino acid may have more than one codons. e.g arginine has 6 different codons

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Gene mutation (altering the nucleotide sequence):

1- Point mutation: changing in a single nucleotide base on the mRNA can lead to any of the following 3 results:

i- Silent mutation: i.e. the codon containg the changed base may code for the same amino acid. For example, in serine codon UCA, if A is changed to U giving the codon UCU, it still code for serine. See table.

ii- Missense mutation: the codon containing the changed base may code for a different amino acid. For example, if the serine codon UCA is changed to be CCA ( U is replaced by C), it will code for proline not serine leading to insertion of incorrect amino acid into polypeptide chain.

iii- Non sense mutation: the codon containing the changed base may become a termination codon. For example, serine codon UCA becomes UAA if C is changed to A. UAA is a stop codon leading to terminationof translation at that point.

How your cell makes very How your cell makes very important proteinsimportant proteins

• The production (synthesis) of proteinsproteins.

• 3 phases3 phases:

1.1. TranscriptionTranscription

2.2. RNA processingRNA processing

3.3. TranslationTranslation

• DNA DNA RNA RNA ProteinProtein

DNA DNA RNA RNA ProteinProtein

Nuclearmembrane

TranscriptionTranscription

RNA ProcessingRNA Processing

TranslationTranslation

DNA

Pre-mRNA

mRNA

Ribosome

Protein

Eukaryotic Eukaryotic CellCell

Before making proteins, Your Before making proteins, Your cell must first make RNAcell must first make RNA

• How does RNARNA (ribonucleic acid) (ribonucleic acid) differ from DNA (deoxyribonucleic acid)DNA (deoxyribonucleic acid)?

RNARNA differs from DNADNA

1. RNARNA has a sugar ribosesugar ribose

DNADNA has a sugar deoxyribosesugar deoxyribose

2. RNARNA contains uracil (U)uracil (U)

DNADNA has thymine (T)thymine (T)

3. RNARNA molecule is single-strandedsingle-stranded

DNADNA is double-strandeddouble-stranded

1. Transcription1. Transcription

• Then moves along one of the DNA strandsDNA strands and links RNARNA nucleotides together.

Nuclearmembrane

TranscriptionTranscription

RNA ProcessingRNA Processing

TranslationTranslation

DNA

Pre-mRNA

mRNA

Ribosome

Protein

Eukaryotic Eukaryotic CellCell

1. Transcription OR1. Transcription OR RNA production RNA production

• RNA molecules are produced by copying part of DNA into a complementary sequence of RNA

• This process is started and controlled by an enzyme called RNA polymerase.

1. Transcription1. Transcription

DNADNA

pre-mRNApre-mRNA

RNA PolymeraseRNA Polymerase

Types of RNATypes of RNA

• Three types ofThree types of RNARNA:

A.A. messenger RNA (mRNA)messenger RNA (mRNA)

B.B. transfer RNA (tRNA)transfer RNA (tRNA)

C.C. ribosome RNA (rRNA)ribosome RNA (rRNA)

• All types ofAll types of RNARNAproduced in theproduced in the nucleus!nucleus!

mRNA

• Carries instructions from DNA to the ribosome.

• Tells the ribosome what kind of protein to make

A. Messenger RNA (mRNA)A. Messenger RNA (mRNA)

methionine glycine serine isoleucine glycine alanine stopcodon

proteinprotein

A U G G G C U C C A U C G G C G C A U A AmRNAmRNA

startcodon

Primary structure of a proteinPrimary structure of a protein

aa1 aa2 aa3 aa4 aa5 aa6

peptide bonds

codon 2 codon 3 codon 4 codon 5 codon 6 codon 7codon 1

rRNA

• Part of the structure of a ribosome

• Helps in protein production

tRNA

• Bring right amino acid to make the right protein according to mRNA instructions

B. Transfer RNA (tRNA)B. Transfer RNA (tRNA)

amino acidamino acidattachment siteattachment site

U A C

anticodonanticodon

methionine amino acidamino acid

RNA ProcessingRNA Processing

Nuclearmembrane

TranscriptionTranscription

RNA ProcessingRNA Processing

TranslationTranslation

DNA

Pre-mRNA

mRNA

Ribosome

Protein

Eukaryotic Eukaryotic CellCell

RNA Processing RNA Processing ((Post Transcriptional Changes)Post Transcriptional Changes)

• IntronsIntrons are pulled out and exonsexons come together.

• End product is a mature RNA mature RNA moleculemolecule that leaves the nucleusnucleus & move to the cytoplasm.cytoplasm.

RNA SplicingRNA Splicing

pre-RNA molecule

intron

intronexon exon exon

exon exon exon

Mature RNA moleculeMature RNA molecule

exon exon exon

intron intron

splicesome splicesome

RibosomesRibosomes

PSite

ASite

Largesubunit

Small subunit

mRNAmRNA

A U G C U A C U U C G

3. Translation - making 3. Translation - making proteinsproteins

Nuclearmembrane

TranscriptionTranscription

RNA ProcessingRNA Processing

TranslationTranslation

DNA

Pre-mRNA

mRNA

Ribosome

Protein

Eukaryotic Eukaryotic CellCell

3. Translation3. Translation

• Three parts:

1. initiationinitiation: start codon (AUG)

2. elongationelongation:

3. terminationtermination: stop codon (UAG)

3. Translation3. Translation

PSite

ASite

Largesubunit

Small subunit

mRNAmRNA

A U G C U A C U U C G

InitiationInitiation

mRNAmRNA

A U G C U A C U U C G

2-tRNA

G

aa2

A U

A

1-tRNA

U A C

aa1

anticodon

hydrogenbonds codon

mRNAmRNA

A U G C U A C U U C G

1-tRNA 2-tRNA

U A C G

aa1 aa2

A UA

anticodon

hydrogenbonds codon

peptide bond

3-tRNA

G A A

aa3

ElongationElongation

mRNAmRNA

A U G C U A C U U C G

1-tRNA

2-tRNA

U A C

G

aa1

aa2

A UA

peptide bond

3-tRNA

G A A

aa3

Ribosomes move over one codon

(leaves)

mRNAmRNA

A U G C U A C U U C G

2-tRNA

G

aa1

aa2

A UA

peptide bonds

3-tRNA

G A A

aa3

4-tRNA

G C U

aa4

A C U

mRNAmRNA

A U G C U A C U U C G

2-tRNA

G

aa1aa2

A U

A

peptide bonds

3-tRNA

G A A

aa3

4-tRNA

G C U

aa4

A C U

(leaves)

Ribosomes move over one codon

mRNAmRNA

G C U A C U U C G

aa1aa2

A

peptide bonds

3-tRNA

G A A

aa3

4-tRNA

G C U

aa4

A C U

U G A

5-tRNA

aa5

mRNAmRNA

G C U A C U U C G

aa1aa2

A

peptide bonds

3-tRNA

G A A

aa3

4-tRNA

G C U

aa4

A C U

U G A

5-tRNA

aa5

Ribosomes move over one codon

mRNAmRNA

A C A U G U

aa1

aa2

U

primaryprimarystructurestructureof a proteinof a protein

aa3

200-tRNA

aa4

U A G

aa5

C U

aa200

aa199

terminatorterminator or stopor stop codoncodon

TerminationTermination

End ProductEnd Product• The end products of protein synthesis is

a primary structure of a proteinprimary structure of a protein.

• A sequence of amino acid amino acid bonded together by peptide bondspeptide bonds.

aa1

aa2 aa3 aa4aa5

aa200

aa199

Regulation of Gene Expression

The control of gene expression

• Each cell in the human contains all the genetic material for the growth and development of a human

• Some of these genes will be need to be expressed all the time called Constitutive genes

• These are the genes that are involved in of vital biochemical processes such as respiration

• Other genes are not expressed all the time• They are switched on an off at need called Non-

constitutive genes

Operons

• An operon is a group of genes that are transcribed at the same time.

• They usually control an important biochemical process.

Jacob, Monod & Lwoff

Inducible Genes - Operon Model

• Definition: Genes whose expression is turned on by the presence of some substance– Lactose induces expression of the lac

genes

• Catabolic pathways

Lactose Operon

• Structural genes– lac z, lac y, & lac a– P-Promoter– Polycistronic

mRNA

• R-Regulatory gene– Repressor

• Operator• Inducer - lactose

i

Operon

RegulatoryGene

p o z y a DNA

m-RNA

-GalactosidasePermease

Transacetylase

Protein

Lactose Operon

• Inducer -- lactose– Absence

• Active repressor• No expression

i p o z y a

No lac mRNA

Absence of lactose

Active

i p o z y a

-Galactosidase Permease Transacetylase

Presence of lactose

Inactive

– Presence• Inactivation of

repressor• Expression

1. When lactose is absent • A repressor protein is continuously synthesised. It sits on

a sequence of DNA just in front of the lac operon, the Operator site

• The repressor protein blocks the Promoter site where the RNA polymerase settles before it starts transcribing

Regulator gene

lac operonOperator site

z y aDNA

I O

Repressor protein

RNA polymeraseBlocked

© 2007 Paul Billiet ODWS

2. When lactose is present • A small amount of a sugar allolactose is formed within

the bacterial cell. This fits onto the repressor protein at another active site (allosteric site)

• This causes the repressor protein to change its shape (a conformational change). It can no longer sit on the operator site. RNA polymerase can now reach its promoter site

Promotor site

z y aDNA

I O

Repressible Genes - Operon Model

• Definition: Genes whose expression is turned off by the presence of some substance (co-repressor)– Tryptophan represses the trp genes

• Co-repressor is typically the end product of the pathway

Tryptophan Operon• Structural genes

– trp E, trpD, trpC trpB & trpA

– Common promoter• Regulatory Gene

– Apo-Repressor• Inactive

• Operator• Co-repressor

– Tryptophan

R

Operon

RegulatoryGene

P O E D C

5 Proteins

B AL

Inactive repressor (apo-repressor)

Tryptophan Operon

• Co-repressor -- tryptophan– Absence of tryptophan

• Gene expression

R P O E D C

5 Proteins

B AL

Inactive repressor (apo-repressor)

Absence of Tryptophan

R P O E D C

No trp mRNA

B AL

Presence of Tryptophan

Inactive repressor (apo-repressor)

Trp(co-repressor)

– Presence of tryptophan

• Activates repressor– No gene expression

• Negative control