GENETIC GENETIC INFORMATIONINFORMATION

STRUCTURE OF THE NUCLEIC ACIDSSTRUCTURE OF THE NUCLEIC ACIDSNUCLEIC ACIDS ARE POLYMERS OF NUCLEOTIDES

- Deoxyribonucleic acid (DNA)

- Ribonucleic acid (RNA)

NUCLEOTIDE STRUCTUREDNA and RNA, are polymers of nucleoside monophosphates (nucleotides) group.

Each nucleotide consists of :

1. A Pentose sugar :

a. Ribose sugar RNA

b. Deoxyribose sugar DNA

2. A Nitrogenous base :2. A Nitrogenous base :

a. Purines : adenine (A) and guanine (G) found in DNA or

RNA

b. Pyrimidins : Cytosine (C), thymine (T), and uracil (U)

Thymines are found only in DNA

Uracils are found only in RNA

3. Phosphate

NUCLEOTIDES are the subunits of nucleic acid,

Including DNA. Each of these subunits is made up of:

AN ORGANIC BASE

+

A PENTOSE SUGAR

+PHOSPHATE GROUP

Note that the phosphate groupis bonded to the C-5 atom of thepentose sugar

NH2

C H

N C

C C

O N H

O-

-O P O CH2 O

O- C C H H H H C C

OH H

1

23

4

5

Nitrogenous bases of the nucleotides structureNitrogenous bases of the nucleotides structure

Primary polymeric structurePrimary polymeric structure

Structure of a trinucleotide. The bases of the three linked nucleotides are guanine, cytosine, and adenine

A section of the A section of the polynucleotide chain in polynucleotide chain in DNA (on the left) and DNA (on the left) and RNA (on the right). The RNA (on the right). The Shorthand notations are Shorthand notations are shown alongside.shown alongside.

The normal base-pairing arrangement found in DNA.The normal base-pairing arrangement found in DNA.(The dashed lines indicate hydrogen bonds)(The dashed lines indicate hydrogen bonds)

THE SECONDARY STRUCTURE OF DNATHE SECONDARY STRUCTURE OF DNATHE DOUBLE HELIX

1. Two antiparallel strands form a right-handed helix

2. Complementary base pairing

(a ) ( b )

Various diagrammatic ways of representing DNA : (a) showing polarity Various diagrammatic ways of representing DNA : (a) showing polarity and base pairing but no helical twist; (b) showing helical twist and helix and base pairing but no helical twist; (b) showing helical twist and helix parameters but not base pairs; © showing helix and base pairs; (d) parameters but not base pairs; © showing helix and base pairs; (d) space-filling representation showing major and minor grooves.space-filling representation showing major and minor grooves.

* CENTRAL DOGMA

DNA RNA PROTEIN

* GENE = Unit herediter yang berfungsi biologis

A B C

Genes DNA Strand

DNAc

RNAtr

DNAt

DNA polymerase

• DNA is divided into 2 cate-DNA is divided into 2 cate-gories: gories: exonsexons have regions have regions in the mRNA that are used in the mRNA that are used to make proteins, whereas to make proteins, whereas intronsintrons are missing from the are missing from the mRNA transcript and do not mRNA transcript and do not code for proteinscode for proteins

tRNA Even the Genes final product rRNA are not proteins

* Operon : The sequence of bases coding for one or more polypep- tides, together with the operator controlling its expression

*Intron (intervening sequence) : - Sequence coding for RNA that is missing from the final product

* Exon (expressed sequences) : - Region coding for RNA that end up in the final RNA product

* A gene : A DNA segment or sequence that codes for a polipeptide.* A gene : A DNA segment or sequence that codes for a polipeptide.

MOLEKULAR STRUCTUREMOLEKULAR STRUCTUREOF CHROMOSOMES AND GENESOF CHROMOSOMES AND GENES

SB

P

a chromosomecarries many genes

a double helix of DNA has two strands

a single stand of DNAis made of manynucleotides

each nucleotideis made of hreecomponents

a very ong molecule of double stranded DNA Chromosome

Gene

Single strandof DNA

Nucleotides

Base (B)+

Sugar (S)+

Posphate (P)

ORGANIZATION OF EUKARYOTIC GENOMIC DNAORGANIZATION OF EUKARYOTIC GENOMIC DNA

Size of DNA molecules - Human = 3.0 x 109 base pairs / haploid cell

= 6.0 x 109 base pairs / diploid cell- 1 chromosome ~ 7 cm in length- 46 chromosome 2 m- < 10% of DNA codes for a product

Packing of DNA in cells- DNA interacts with basic proteins known as histones- Histones arginine and lysine- Five classes of histones : H1, H2A, H2B, H3 and H4

H2A H2B Ellipsoid H3 H4

- Other types of proteins are also associated with DNA- Nucleosomes : * bead-like structures of DNA

* DNA coils around the surface of the ellipsoid * Complex of histones plus DNA

REPLIKASI DNAREPLIKASI DNA

OBJECTIVES

* Menerangkan replikasi DNA secara tepat* Mengetahui peran masing tipe DNA polimerase* Mengenal macam mutasi dan DNA repair

Basic requirments for DNA synthesis :1. Substrates2. Template3. Primer4. Enzymes

PROKARYOTIC ENZYMES AND PROTEINS PROKARYOTIC ENZYMES AND PROTEINS FOR REPLICATIONFOR REPLICATION

Multiple DNA polymerases with multiple enzymatic activities(1). DNA polymerase I (pol I) a. Function. Pol I functions in the replication of DNA and in the repair of damaged DNA b. Other enzymatic activities. Pol I has two enzymatic activities - besides DNA polymerase activity (1) Proofreading (Figure 1)

Figure 1. 3’ to 5’ Exonuclease (proofreading) activity

(2) Excision-repair (Figure 2). Pol I has a 5’ to 3’ exonuclease activity, called excision-repair activity, that can hydrolytically remove a segment of DNA from the 5’ end of a strand of duplex DNA

(a) One to ten nucleotide segments of DNA can be removedat one time

3’ to 5’

5’ - G T C A T G G 5’ - G T C A T G G … .. … .. .. … ... … .. … .. .. … ... 3’ - C A G T A C C G C T T A G - 5’ 3’ - C A G T A C C G C T T A G - 5’ + dTMP

exonuclease

3’

- T- O

H

(b) This activity is essential for the removal of primers in DNA (b) This activity is essential for the removal of primers in DNA replicationreplication

(c) This activity is essential for the repair of damaged DNA(c) This activity is essential for the repair of damaged DNA

Figure 2. 5’ to 3’ exonuclease (excision-repair) activity.

3’ to 5’

G A A T G- 3’ 5’ - T C -3’ … .. … .. .. .. … 3’ - C A G T A C C G C T T A G - 5’ 3’ - C A G T A C C G C T T A G - 5’

+ 5’ - T G A A - 3’

exonuclease5’ -T-

Schematic diagram of the basic molecular events at the replication Schematic diagram of the basic molecular events at the replication fork of fork of E.ColiE.Coli

1. DnaA protein 1. DnaA protein is required for proper initiation is required for proper initiation of replication at the originof replication at the origin

2. Primase :

- The catalytic partion of a primasome that makes the RNA primer both lagging and leading

strands synthesis

- Primer provide a 3’-hydroxyl group needed to initiate DNA synthesis

3. Primosome

3. Primosome 3. Primosome a complex of proteins that comprises : a complex of proteins that comprises : - Primase

- dna B has the helicase activity

- dna C

- Other proteins (n, n’, n” and I)

4. Helicases :

- Enzymes that catalyze the unwinding of the DNA helix

- Derive energi from ATP

- Provide single-strand templates for replication

- Disrupt the hydrogen bonds

5. DNA gyrase :5. DNA gyrase :

- Unwinds the supercoil DNA by cutting both strands of DNA

- Followed by the helicase activity

6. Single-strand binding protein ( SSBP )

it binds to single-stranded DNA (ssDNA)

Function : - to protect ssDNA from nucleases

7. DNA Polymerases

- Unwinding of parental strands- Unwinding of parental strands

* Helicases unwind the parental DNA * Single-strand binding proteins ( SSB ) prevent from reassociating

protection

- Action of DNA Polymerase E.Coli has three DNA polymerase Pol I Pol II Pol III

- DNA gyrase uncoils DNA

THE REPLICATION PROCESS TAKES PLACES IN FOUR STAGES:THE REPLICATION PROCESS TAKES PLACES IN FOUR STAGES:

1. Unwind the helix by helicases

- With the aid of the DNA gyrase

- Dna B protein

- SSBP

2. DNA is replicated by DNA polymerase III.

The detail of the process :

- Primase ( a special RNA) a short RNA primer (10

nucleotides and compelemtary to the DNA)

- DNA polymerase III synthesizes complementary DNA

beginning at the 3’ end of the RNA primer

- Leading and lagging strand are performed

- Lagging strand replcation is discontinous and the

fragments is called Okazaki fragments

3. Removal the RNA primer 3. Removal the RNA primer filling the gap by DNA filling the gap by DNA polymerase I polymerase I

4. The fragments are joined by the enzyme DNA lygase4. The fragments are joined by the enzyme DNA lygase

EUKARYOTIC REPLICATION

- Semiconcervative and proceeds bidirectionally from many origins.- Replication rate a. Prokaryotes. An E.coli replication fork progresses at pproximately 1000 base pairs per second. b. Eukaryotes. The eukaryotic replication rate is about 10 times slower than the prokaryotic replication rate.- 8 hours to replicate the human genome.- Enzymes and proteins involved in the replication : * Multiple eukaryotic DNA polymerases. 1. DNA polymerase is essential for replication. - Has primase activity - It is required for both leading and lagging strand synthesis

2. DNA polymerase also is essential for replication. It is required for both leading and lagging strand synthesis

a. Pol associated with proliferating cell nuclear antigen (PCNA)

b. After pol has initiated replication, a protein called replication

factor C (RF-C) facilitates the inhibition an replacement of pol with both PCNA and pol in an ATP- dependent manner.

3. DNA polymerase plays no role in replication and acts only in DNA repair synthesis

4. DNA polymerase also is essential for replication, although its exact role is not clearly defined. It behaves similarly to pol and may support some component of lagging strand synthesis.

5. DNA polymerase replicates mitochondrial DNA

Other factors involved in replicationOther factors involved in replication::

1. 5’ to 3’ exonucleases. One is known to be associated with DNA polymerase .

2. 3’ to 5’ exonucleases activity. DNA polymerase , , and have proofreading capability.

3. Ligase. Unlike prokaryotic ligase that use nicotinamide adenine dinucleotide (NAD) as an energy source, eukaryotic ligases use

ATP

4. Helicase

5. An SSBP called replication protein A (RP-A) has been isolated from mammalian cells

6. Topoisomerases. These enzymes build up in advance of replication forks.

Two basic types of eukaryotic topoisomerases.

a. Topoisomerase I is the major topoisomerase used to relieve supercoils

b. Topoisomerase II also is required during replication.

- Synthesis always goes from 5’ to 3’

Phosphate – O – CH2 Base

O

4’

H’ 1’

H H

H H

3’ 2’

OH H

Next nucleotide

will joine here

NUCLEOTIDES are the subunits of nucleic acid,

Including DNA. Each of these subunits is made up of:

AN ORGANIC BASE

+

A PENTOSE SUGAR

+PHOSPHATE GROUP

Note that the phosphate groupis bonded to the C-5 atom of thepentose sugar

NH2

C H

N C

C C

O N H

O-

-O P O CH2 O

O- C C H H H H C C

OH H

1

23

4

5

CHANGES IN GENETIC MATERIAL CHANGES IN GENETIC MATERIAL

There are 2 types of changes: 1. Point mutation gene mutation

mutation 2. Gross mutation chromosome

aberration aberration

Chromosome aberrationChromosome aberration

Abnormal structure or number of chromosomes; includes deficiency, duplication, inversion, translocation, aneuploid, polyploid, or any change from normal

Point mutation/mutationPoint mutation/mutation A mutation caused by the altera- tion of

a single base in DNA:

Substitution of one base-pair for anotherDuplication of single base-pairsDeletion of single base-pairs

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MUTAGENS (MUTAGENS (= mutagenic agents= mutagenic agents) ) 1. PHYSICAL MUTAGENS: SHORT WAVE a. NON-RADIOACTIVE: UV. b. RADIOACTIVE: X, , , -RAYS.2. CHEMICAL MUTAGENS alkylating agents, nitric acid, acridin pig- ment, analog base, gas.3. BIOLOGICAL MUTAGENS:

a. virus b. aging

TYPES OF MUTATIONTYPES OF MUTATION

a. MUTATION ACCORDING TO CHANGE LOCATION

1. Mutation on nucleotides: deletion, substitution, insertion, and dimerization

2. Mutation on splicing

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b. MUTATION IN ACCORDANCE WITH THE FORM OF CHANGES

1. Mis-sense mutation2. Non-sense mutation3. Silence mutation4. Frame-shift mutation

DELETIONDELETION1 base deletion

Codogen: TTC ATG GGA CGT ACodon: AAG UAC CCU GCA UAmino acid: lys - tyr - arg - ala –

Deletion of base C of first codonDeletion of base C of first codonCodogen: TTCodogen: TT

Frame-shift are occurred here Frame-shift are occurred here miss- miss-sense, and protein formed are distinctly sense, and protein formed are distinctly different from the originaldifferent from the original

A TGG GAC GTAA TGG GAC GTACodon: Codon: AAU ACC CUG CAUAAU ACC CUG CAUAmino acid: asn - thr - val – hisAmino acid: asn - thr - val – his

1 Codon Deletion 1 Codon Deletion CodogenCodogen :: TTATTA TAGTAG TATAGG GAGAAA CCACCA CAACAACodon Codon :: AAU AUCAAU AUC AUCAUC CUUCUU GGUGGU GUUGUUAmino acid: asn - ile - ile - leu - gly - valAmino acid: asn - ile - ile - leu - gly - val

Mutation on base sequence GGAMutation on base sequence GGACodogenCodogen :: TTATTA TAGTAG TA TA CodonCodon :: AAU AUCAAU AUCAUAUAmino acid: asn - ile -Amino acid: asn - ile -

AA CCACCA CAACAAUU GGUGGU GUUGUU

ile - gly - valile - gly - valFrame-shift are occurred here, and protein Frame-shift are occurred here, and protein formed are distinctly different from the formed are distinctly different from the originaloriginal

SUBSTITUTION SUBSTITUTION 1. A single-base substitution

Originalcodon

Originalam. acid

Newcodon

Newam. acid

UUAGAU

LeuAsp

UUGGAA

LeuGlu

SILENT MUTATION

MIS-SENSE MUTATION