(164446829) Molecular Genetic

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    MOLECULARGENETICPART IGene Regulation in Protein

    Synthesis

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    1 GeneRegulationSynthesisinProteinA.

    B.

    C.

    D.

    E.

    Nucleic Acid Structure

    Replication of DNA

    Transcription of Bacterial DNA

    Transcription of Eukaryote DNA

    Translation

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    A.Nucleic AcidStructure

    Two types of nucleic acids- DNA & RNA

    DNA (DeoxyriboNucleic Acid) & RNA

    (RiboNucleic Acid)

    Covalent link between atoms IN nucleic

    acids building blocksprotect the

    biological polymers

    Weak bond BETWEEN different parts

    allow functions

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    DNA

    Molecule in which the genetic materialstored

    Nucleoside: purine base (Adenine,Guanine) pyrimidine base (Thymine,Cytosine)

    1. is

    2.

    3.

    4.

    5.

    F(x) as a

    F(x) as a

    DNA canA form,

    template for its self-replication

    template for RNA synthesis

    adopt different conformationsB form, Z form

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    DNA

    DNA can exist in two conditionssupercoiled & relaxed

    Naturally occurred in negatively supercoiledcondition

    Enzyme alter DNA structure

    6.

    7.

    8.Topoisomerase I (A & B) for single strand break

    Topoisomerase II for double strand break,multimeric enzyme, require ATP hydrolysis to

    complete two DNA strands cleavageProkaryotes has a special topoisomerase IIknown as DNA gyrase which introduce negativesupercoil

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    DNA

    Stabilize by hydrogen bonding, stackinginteractions and ionic interactions

    9.

    10.Undergo denaturation when heated and

    anneal when cooled

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    bond

    PhosphodiesterHydrogen

    bond

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    RNA

    1.

    2.

    3.

    Chemically very similar to

    Three differences:

    DNA

    deoxyribose vs ribose

    thymine vs uracil

    Typically found in

    cell as singlepolynucleotide chain

    backbone

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    RNA

    1.

    Five functions:

    Intermediate between the geneprotein synthesizing machinery

    Adaptor between the codons inamino acids (tRNA)

    and the(mRNA)mRNA and2.

    3. Structural role as a components of theribosomes (rRNA)

    Regulatory molecule which complement and

    interferes with the translation of certainmRNAsEnzymes that catalyze essential reactions inthe cell (ribozymes)

    4.

    5.

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    RNA

    F(x) as a template for protein

    Comes in various shapes,

    sizes and some with catalytic

    synthesis

    property

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    B. Replication of DNATHE CHEMISTRY OF DNA SYNTHESIS

    Requires TWO key substratesdNTP(dATP, dGTP, dCTP, dTTP) and

    primer:template junction

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    B. Replication of DNATHE CHEMISTRY OF DNA SYNTHESIS

    DNA is synthesized by extending theend of the primer in antiparallelorientation

    Matching dNTP form a base pair byhydrogen bonding

    3

    -

    - hydroxyl group from 3end of primer attack

    the P (-phosphoryl) from dNTP to formphosphodiester bond and release thepyrophosphate (- and -phosphoryl)

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    B. Replication of DNATHE CHEMISTRY OF DNA SYNTHESIS

    Hydrolysis of pyrophosphate as adriving force for DNA synthesis

    - Polymerization of nucleotides is promoted

    by additional free energy provided by the

    rapid hydrolysis of pyrophosphate

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    B. Replication of DNAINITIATION OF DNA REPLICATION

    Involves a replicator and initiatorcomponents

    Replicator is a set of cis-acting DNA

    sequence sufficient to direct an initiationreplication

    Binding site for the initiator protein which

    nucleates the assembly of replicationinitiation machinery

    Consist of AT-rich region which unwindreadily

    of

    -

    -

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    B. Replication of DNA

    Initiator is a sequence-specific DNAbinding protein

    - Binds to a specific sequencereplicator

    Unwind the DNA region

    Attract other factors required

    of replication

    within the

    -

    -for initiation

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    B. Replication of DNA

    Unwinding of duplex DNA strandscatalyzed by DNA helicase

    To stabilize the unwound single-strand

    DNA , a single-strand DNA bindingprotein (SSBs) bind to ssDNA

    Replication fork formation

    Primase synthesis RNA primers onssDNA to initiate DNA synthesis by DNApolymerase

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    B. Replication of DNAREPLICATION FORK

    DNA is synthesized semidiscontinuously

    Both DNA strands are synthesized

    simultaneously

    The junction between newly separated

    strands and the unreplicated duplex DNA

    is known as replication fork

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    B. Replication of DNAREPLICATION FORK

    Synthesis of DNA occur only from 5

    direction. to 3

    In lagging strand, DNA is synthesized

    discontinuously and form a short new

    fragments, known as Okazaki fragments

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    B. Replication of DNADNA Polymerase

    Key enzyme for DNA replication

    Have single active site which catalyze

    addition of any four dNTPs

    the

    Bound to primer:template junction on DNAstrand

    Monitor the accuracy of base-pairing bycatalysis rate. High rate=matched, slow

    rate=mismatched

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    B. Replication of DNADNA Polymerase

    When mismatched occur, primer:template

    junction will slide out from catalytic site of

    DNA polymerase into proofreading

    exonuclease site

    Exonuclease will remove the unmatched

    base from 3-end

    Primer:template junction will slide in into

    the catalytic site of DNA polymerase

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    B. Replication of DNA

    RNA primers are removed by RNAse Hthrough degradation

    The final ribonucleotide bond to DNA end is

    removed by exonuclease which result in gaps

    DNA ligase use ATP to create

    phosphodiester bond between 5phosphate

    and 3OH in a newly synthesized DNA

    strands to close this gap

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    REPLICATION TRANSCRIPTION

    Synthesis of new Synthesis of new

    deoxyribonucleotide strand ribonucleotide strand

    Catalyze by DNA polymerase Catalyze by RNA polymerase

    DNA polymerase require a RNA polymerase doesntprimer to initiate synthesis require any primer to initiate

    synthesis The newly synthesized DNA The newly synthesized RNAform a base-pair with DNA doesnt remain base-pairedtemplate to the DNA template

    Involve extensive Lack of extensive

    proofreading proofreading Copy the entire genome Copy certain parts of the

    once in every cell division genome

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    C. Transcription of DNA

    1.2.

    3.

    Key enzyme is RNA polymerase

    Involve three phases:

    InitiationElongation

    Termination

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    C. Transcription of Bacterial DNAINITIATION

    Binding of polymerase to a promoter

    sequence, form a closed complex

    Upon binding, DNA strands separate

    around the start site and form a

    transcription bubble (open complex)

    First two ribonucleotides are bought into

    active site, aligned and joined

    Polymerase move ahead opening the DNA

    duplex

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    C. Transcription of Bacterial DNAINITIATION

    Formation of first 10 bp ribonucleotides is

    a rather inefficient process, usually being

    released and transcription start again-

    abortive initiation

    When transcribed sequence reach morethan 10 bp, a stable ternary complex is

    established

    Followed by elongation phase

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    C. Transcription of Bacterial DNAELONGATION

    DNA duplex enter polymerase

    separated at the catalytic cleft

    Ribonucleotides enter the site

    and

    and joinedthe growing RNA chain guided by the DNAtemplate

    Only 8-9 growing RNA chain remain on theDNA template and the previously

    generated are peeled off and directed out

    from polymerase

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    C. Transcription of Bacterial DNAELONGATION

    During elongation, polymerase carries out twoproofreading functions:

    Pyrophosphorolytic editing

    1.

    Polymerase uses its active site to catalyzeremoval of incorrect ribonucleotide by

    incorporating the pyrophosphate (PPi)

    2. Hydrolytic editingPolymerase is backtrack