Gene to Protein(KFogler)

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    AP Biology 2007-2008

    From Geneto Protein

    How GenesWork

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    AP Biology

    What do genes code for?

    proteins cells bodies

    How does DNA code for cells & bodies? how are cells and bodies made from the

    instructions in DNA

    DNA

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    AP Biology

    The Central Dogma

    Flow of genetic information in a cell How do we move information from DNA to proteins?

    replication

    proteinRNADNA trait

    DNA getsall the glory,

    but proteins doall the work!

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    AP Biology

    Inheritance of metabolic diseases suggested that genes coded for enzymes

    each disease (phenotype) is caused by

    non-functional gene product lack of an enzyme

    Tay sachs

    PKU (phenylketonuria)

    albinism

    Am I just thesum of my proteins?

    Metabolism taught us about genes

    A B C D E

    disease disease disease disease

    enzyme 1 enzyme 2 enzyme 3 enzyme 4

    metabolic pathway

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    AP Biology

    Beadle & Tatum1941 | 1958

    George Beadle

    Edward Tatum

    "for their discovery that genes act byregulating definite chemical events"

    one gene : one enzyme hypothesis

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    AP Biology

    Wild-typeNeurospora

    Minimalmedium

    Select one ofthe spores

    Grow oncomplete medium

    Minimalcontrol

    Nucleicacid

    CholinePyridoxine RiboflavinArginine

    Minimal media supplemented only with

    ThiamineFolicacid NiacinInositolp-Aminobenzoic acid

    Test on minimalmedium to confirmpresence of mutation

    Growth oncompletemedium

    X rays or ultraviolet light

    asexualspores

    spores

    Beadle & Tatum

    create mutations

    positive control

    negative control

    experimentals

    amino acidsupplements

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    AP Biology

    mRNA

    From gene to protein

    DNAtranscription

    nucleus cytoplasm

    aa

    aa

    aa

    a

    a

    aa

    aa

    a

    a

    aa

    aa

    aa

    aa

    proteintranslation

    ribosome

    trait

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    AP Biology 2007-2008

    Transcription

    fromDNA nucleic acid language

    to

    RNA nucleic acid language

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    AP Biology

    RNA

    ribose sugar N-bases

    uracil instead of thymine

    U : A C : G

    single stranded

    lots of RNAs mRNA, tRNA, rRNA, siRNA

    RNADNA

    transcription

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    AP Biology

    Transcription

    Making mRNA transcribed DNA strand = template strand

    untranscribed DNA strand = coding strand same sequence as RNA

    synthesis of complementary RNA strand transcription bubble

    enzyme RNA polymerase

    template strand

    rewinding

    mRNA RNA polymerase

    unwinding

    coding strand

    DNAC C

    C

    C

    C

    C

    C

    C

    C CC

    G

    G GG

    G G

    G G

    G

    G

    GA

    A

    AA A

    A

    A

    A

    A

    A A

    A

    AT

    T T

    T

    T

    T

    T

    T

    T T

    T

    T

    U U

    5

    353

    3

    5build RNA 53

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    AP Biology

    RNA polymerases

    3 RNA polymerase enzymes RNA polymerase 1

    only transcribes rRNA genes

    makes ribosomes RNA polymerase 2

    transcribes genes into mRNA

    RNA polymerase 3

    only transcribes tRNA genes

    each has a specific promoter sequenceit recognizes

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    AP Biology

    Which gene is read?

    Promoter region binding site before beginning of gene

    TATA box binding site

    binding site for RNA polymerase& transcriptionfactors

    Enhancer region

    binding site farupstream of gene

    turns transcription

    on HIGH

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    AP Biology

    Transcription Factors

    Initiation complex transcription factors bind to promoter region

    suite of proteins which bind to DNA

    hormones?

    turn on or off transcription

    trigger the binding of RNA polymerase to DNA

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    AP Biology

    Matching bases of DNA & RNA

    Match RNA bases to DNAbases on one of the DNAstrands

    U

    AG GGGGGT T A C A C T T T T TC C C CA A

    U

    UU

    U

    U

    G

    G

    A

    A

    A C CRNA

    polymerase

    C

    C

    C

    C

    C

    G

    G

    G

    G

    A

    A

    A

    A

    A

    5' 3'

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    AP Biology

    Eukaryotic genes have junk!

    Eukaryotic genes are not continuous

    exons = the real gene

    expressed / coding DNA

    introns = the junk inbetween sequence

    eukaryotic DNA

    exon = coding (expressed) sequence

    intron = noncoding (inbetween) sequence

    intronscome out!

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    AP Biology

    mRNA splicing

    eukaryotic DNA

    exon = coding (expressed) sequence

    intron = noncoding (inbetween) sequence

    primary mRNAtranscript

    mature mRNAtranscript

    pre-mRNA

    spliced mRNA

    Post-transcriptional processing eukaryotic mRNA needs work after transcription

    primary transcript = pre-mRNA

    mRNA splicing

    edit out introns

    make mature mRNA transcript

    ~10,000 bas

    ~1,000 bas

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    AP Biology

    1977 | 1993

    RichardRoberts Philip

    SharpCSHL

    MIT

    adenovirus

    common cold

    Discovery of exons/introns

    beta-thalassemia

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    AP Biology

    Splicing must be accurate

    No room for mistakes! a single base added or lost throws off the

    reading frame

    AUG|CGG|UCC|GAU|AAG|GGC|CAU

    AUGCGGCTATGGGUCCGAUAAGGGCCAUAUGCGGUCCGAUAAGGGCCAU

    AUG|CGG|GUC|CGA|UAA|GGG|CCA|U

    AUGCGGCTATGGGUCCGAUAAGGGCCAUAUGCGGGUCCGAUAAGGGCCAU

    Met|Arg|Ser|Asp|Lys|Gly|His

    Met|Arg|Val|Arg|STOP|

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    AP Biology

    RNA splicing enzymes

    snRNPs

    exonexon intron

    snRNA

    5' 3'

    spliceosome

    exonexcisedintron

    5'

    5'

    3'

    3'

    3'

    lariat

    exonmature mRNA

    5'

    No,not smurfs!snurps

    snRNPs small nuclear RNA

    proteins

    Spliceosome

    several snRNPs

    recognize splicesite sequence

    cut & paste gene

    Whoa! I thinkwe just broke

    a biological rule!

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    AP Biology

    Alternative splicing

    Alternative mRNAs produced from same gene when is an intron not an intron

    different segments treated as exons

    Starting to gethard to

    define a gene!

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    AP Biology

    AmRNA

    5'

    3'

    G PPP

    More post-transcriptional processing

    Need to protect mRNA on its trip fromnucleus to cytoplasm

    enzymes in cytoplasm attack mRNA protect the ends of the molecule

    add 5 GTP cap add poly-A tail longer tail, mRNA lasts longer: produces more protein

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    AP Biology

    mRNA

    From gene to protein

    DNAtranscription

    nucleus cytoplasm

    aa

    aa

    aa

    a

    a

    aa

    aa

    a

    a

    aa

    aa

    aa

    aa

    ribosome

    trait

    proteintranslation

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    AP Biology 2007-2008

    Translation

    fromnucleic acid language

    to

    amino acid language

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    AP Biology

    How does mRNA code for proteins?

    TACGCACATTTACGTACGCGGDNA

    AUGCGUGUAAAUGCAUGCGCCmRNA

    MetArgValAsnAlaCysAlaprotein

    ?

    How can you code for 20 amino acids

    with only 4 nucleotide bases (A,U,G,C)?

    4

    4

    20

    ATCG

    AUCG

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    AP Biology

    AUGCGUGUAAAUGCAUGCGCCmRNA

    mRNA codes for proteins in triplets

    TACGCACATTTACGTACGCGGDNA

    AUGCGUGUAAAUGCAUGCGCCmRNA

    MetArgValAsnAlaCysAlaprotein

    ?

    codon

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    AP Biology

    Cracking the code1960 | 1968

    Crick determined 3-letter (triplet) codon system

    Nirenberg & Khorana

    WHYDIDTHEREDBATEATTHEFATRAWHYDIDTHEREDBATEATTHEFATRA

    Nirenberg (47) & Khorana (17)

    determined mRNAamino acid match

    added fabricated mRNA to test tube ofribosomes, tRNA & amino acids

    created artificial UUUUU mRNA

    found that UUU coded for phenylalanine

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    AP Biology

    1960 | 1968Marshall Nirenberg

    Har Khorana

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    AP Biology

    The code

    Code for ALL life! strongest support for

    a common origin forall life

    Code is redundant

    several codons foreach amino acid

    3rd base wobble

    Start codon

    AUG

    methionine

    Stop codons

    UGA, UAA, UAG

    Why is the

    wobble good?

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    AP Biology

    How are the codons matched to

    amino acids?

    TACGCACATTTACGTACGCGGDNA

    AUGCGUGUAAAUGCAUGCGCCmRNA

    aminoacid

    tRNA anti-codon

    codon

    5 3

    3 5

    3 5UAC

    Met GCAArg

    CAU

    Val

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    AP Biology

    mRNA

    From gene to protein

    DNA transcription

    nucleus cytoplasm

    aa

    aa

    aa

    a

    a

    aa

    aa

    aa

    aa

    a

    a

    aa

    aa

    ribosome

    traitaa

    proteintranslation

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    AP Biology

    Transfer RNA structure

    Clover leaf structure anticodonon clover leaf end

    amino acid attached on 3 end

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    AP Biology

    Loading tRNA

    Aminoacyl tRNA synthetase enzyme which bonds amino acid to tRNA

    bond requires energy

    ATP AMP bond is unstable so it can release amino acid at ribosome easily

    activatingenzyme

    anticodon

    tRNATrp binds to UGGcondon of mRNA

    Trp Trp Trp

    mRNAACCUGG

    C=O

    OHOH

    H2

    OO

    tRNATrp

    tryptophan attached

    to tRNATrp

    C=O

    O

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    AP Biology

    Ribosomes

    Facilitate coupling oftRNA anticodon tomRNA codon

    organelle or enzyme? Structure

    ribosomal RNA (rRNA) & proteins

    2 subunits

    large

    smallE P A

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    AP Biology

    Ribosomes

    Met

    5'

    3'

    UUA C

    A G

    APE

    A site (aminoacyl-tRNA site) holds tRNA carrying next amino acid to

    be added to chain

    P site (peptidyl-tRNA site) holds tRNA carrying growing

    polypeptide chain

    E site (exit site)

    empty tRNAleaves ribosomefrom exit site

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    AP Biology

    Building a polypeptide

    Initiation brings together mRNA, ribosome

    subunits, initiator tRNA

    Elongation

    adding amino acids based oncodon sequence

    Termination end codon 123

    Leu

    Leu Leu Leu

    tRNA

    Met MetMet Met

    PE A

    mRNA5' 5' 5' 5'

    3' 3' 3'3'

    U UA AAAC

    CC

    AU UG GGU

    UA

    AAAC

    CC

    AU UG GGU

    UA

    AAAC

    CC

    AU UG GGU U

    AAA

    CCAU UG G

    Val Ser

    AlaTrp

    releasefactor

    AAA

    CCU UGG 3'

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    AP Biology

    Protein targeting

    Signal peptide address label

    Destinations: secretion nucleus

    mitochondria chloroplasts cell membrane cytoplasm etcstart of a secretory pathway

    RNA l

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    AP Biology

    Can you tellthe story?

    DNA

    pre-mRNA

    ribosome

    tRNA

    aminoacids

    polypeptide

    mature mRNA

    5' GTP cap

    poly-A tail

    large ribosomal subunit

    small ribosomal subunit

    aminoacyl tRNAsynthetase

    E P A

    5'

    3'

    RNA polymerase

    exon intron

    tRNA

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    AP Biology AAAAAAAAGTP

    20-30b

    3'

    promoter transcriptionstop

    transcriptionstart

    introns

    The Transcriptional unit (gene?)

    transcriptional unit (gene)TAC ACT

    DNA

    DNATATA5'RNA

    polymerase

    pre-mRNA

    5' 3'

    translationstart

    translationstop

    mature mRNA

    5' 3'

    UTR UTR

    exonsenhancer1000+b

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    AP Biology 2007-2008

    ProteinSynthesis in

    Prokaryotes

    Bacterial chromosome

    mRNA

    Cell wall

    Cellmembrane

    Transcription

    Psssstno nucleus!

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    AP Biology

    Prokaryote vs. Eukaryote genes

    Prokaryotes DNA in cytoplasm

    circularchromosome

    naked DNA

    no introns

    Eukaryotes DNA in nucleus

    linearchromosomes

    DNA wound onhistone proteins

    introns vs. exons

    eukaryoticDNA

    exon = coding (expressed) sequence

    intron = noncoding (inbetween) sequence

    introns

    come out!

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    AP Biology

    Transcription & translation are simultaneousin bacteria

    DNA is incytoplasm

    no mRNAediting

    ribosomes

    read mRNAas it is beingtranscribed

    Translation in Prokaryotes

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    AP Biology

    Translation: prokaryotes vs. eukaryotes

    Differences between prokaryotes &eukaryotes

    time & physical separation betweenprocesses takes eukaryote ~1 hour

    from DNA to protein

    no RNA processing

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    AP Biology 2007-2008

    Any Questions??What color would a smurf turn

    if he held his breath?

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    AP Biology 2007-2008

    Substitute Slidesfor Student Print version

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    AP Biology

    Can you tellthe story?

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    20-30b

    3'

    introns

    The Transcriptional unit

    transcriptional unitTAC ACT

    DNATATA5'RNA

    polymerase

    5' 3'

    5' 3'

    exonsenhancer

    1000+b