genetics TEST III (1).txt

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genetics TEST III (1).txt
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genetics replication dnap rnap
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study cards for genetics test III
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  1. REPLICATION
    how DNA is copied & maintained from cell division to the next.
  2. Watson/Crick structure of DNA gave what insight into the mechanism/process of DNA replication?
    • -Complimentary nature of the 2 strands of the double helix
    • -suggested each strand of helix could serve as template to direct the synthesis of a new complimentary strand
  3. SEMI-CONSERVATIVE MODEL (Watson/Crick)
    • SEMI-CONSERVATIVE MODEL OF DNA REPLICATION
    • EACH PARENTAL STRAND OF THE DOUBLE HELIX ACTS AS A TEMPLATE TO DIRECT THE ASSEMBLY OF A NEW DAUGHTER STRAND THAT IS COMPLEMENTARY TO ITSELF
    • RESULT: EACH NEW MOLECULE OF DNA WOULD CONSIST OF A PARENTAL & A DAUGHTER STRAND
    • THUS: THE PARENTAL STRAND IS SEMI-CONSERVED IN THE REPLICATED STRUCTURE
    • 
  4. CONSERVATIVE MODEL
    • PARENTAL STRANDS ARE FULLY CONSERVED IN THAT THE ENTIRE DNA MOLECULE SERVES AS A TEMPLATE FOR A NEW MOLECULE COMPRISED OF ONLY NEWLY SYNTHESIZED DNA
  5. DISPERSIVE MODEL
    • LARGE AMOUNTS OF DNA
    • THE ORIGINAL DOUBLE HELIX GETS BROKEN DOWN INTO MANAGEABLE SIZED FRAGMENTS
    • EACH OF THESE FRAGMENTS, THEN SERVES AS TEMPLATE TO MAKE NEW DNA
    • REASSEMBLE THE FRAGMENTS AFTER REPLICATION IS COMLETED
    • LEAST LIKELY MODEL B/C THERE CAN BE ERRORS IN ASSMEBLY PROCESS
  6. MESELSON & STAHL - '58
    • -5 YEARS AFTER STRUCTURE OF DNA
    • CARRIED OUT A SET OF EXPERIMENTS IN WHICH THEY COULD LOOK FOR CORRECT MODEL OF DNA REPLICATION
    • NEEDED A WAY TO DISTINGUISH B/W PARENT & DAUGHTER DNA
    • USED HEAVY 15N & LIGHT 14N ISOTOPES OF NITROGEN TO LABEL THE NITROGENOUS BASES OF DNA
  7. WHAT WOULD YOU EXPECT TO FIND IF DNA REPLICATES BY THE SEMI-CONSERVATIVE MECHANISM??
  8. WHAT WOULD YOU EXPECT TO FIND IF DNA REPLICATES BY THE CONSERVATIVE MECHANISM??
  9. WHAT WOULD YOU EXPECT TO FIND IF DNA REPLICATES BY THE DISPERSIVE MECHANISM??
  10. WHICH MODEL DID MESELSON & STOHLS EXPERIMENT SUPPORT?
    SEMI-CONSERVATIVE
  11. HOW DOES SEMI-CONSERVATIVE REPLICATION OCCUR?
    • DIFFER LARGELY BASED ON TEMPLATE
    • *TYPE OF TEMPLATE
    • *# OF REPLICATION ORIGINS
    • 1.THETA REPLICATION
    • 2. ROLLING CIRCLE REPLICATION
    • 3. EUKARYOTIC DNA REPLICATION
  12. REPLICON
    INDIVIDUAL UNIT OF REPLICATION
  13. REPLICATION ORIGIN
    • POINT ON CHROMOSOME WHERE REPLICATION ORIGINATES
    • *EACH REPLICON CONTAINS A SINGLE ORIGIN OF REPLICATION
  14. REPLICATION BUBBLE
    • REGION OF UNWINDING OF DOUBLE HELIX
    • *EXPOSES ssDNA THAT CAN BE COPIED
    • *CENTERED OVER THE ORIGIN
  15. REPLICATION FORK
    SITE OF ACTIVE REPLICATION AS IT MOVES DOWN REPLICATION TEMPLATE DURING COPYING STEPS
  16. THETA REPLICATION
    • NAMED AFTER THE GREEK LETTER
    • COMMON TO CIRCULAR GENOMES
    • EXAMPLE: BACTERIAL CHROMOSOME
    • GENERATES A REPLICATION INTERMEDIATE THAT RESEMBLES THETA
    • 1ST DESCRIBED BY JOHN CAIRNS
    • EXP:
    • *GREW E.COLI IN PRESENCE OF TRITILATED THYMIDINE (RADIOACTIVE)
    • *SAMPLED THE CELLS AT VARIOUS TIMES OF REPLICATION PROCESS
    • EACH SAMPLE WAS "LYSED" OPEN ONTO A GLASS SLIDE - RELEASE THE DNA
    • *EXPOSED SLIDE TO X-RAY FILM OR PHOTOGRAPHIC EMMALISM - THE "HOT" DNA WILL BE CAPTURED IN A PHOTOGRAPHIC IMAGE
    • *LOOK AT IMAGE OF DNA

    • CONCLUSION:
    • E.COLI CHROMOSOME HAS SINGLE ORIGIN OF REPLICATION=SINGLE REPLICON
    • PROPOSED:
    • DNA UNWINDS @ THE ORIGIN OriC (CHROMOSOMAL) - ALWAYS MEANS REPLICATION
    • --GENERATES LOCALIZED REGION OF ssDNA THAT CAN NOW SERVE AS TEMPLATES TO DIRECT THE SYNTHESIS OF A NEW, COMPLEMENTARY DAUGHTER STRAND

    • -PRESCOTT SHOWED E.COLI REPLICATION IS BIDIRECTIONAL
  17. ROLLING CIRCLE REPLICATION
    • -COMMON TO CIRCULAR GENOMES
    • -UNIDIRECTIONAL REPLICATION
    • EX:
    • *CONJUGATION & REPLICATION OF F FACTOR
    • * BACTERIOPHAGE THETA REPLICATION
  18. EUKARYOTIC DNA REPLICATION
    • Linear DNA template
    • A lot larger genome
    • Utilizes multiple origins of replication

    • **the human genome has
    • 20k-30k origins
    • If there was use of only a single origin

    • Take 5-7 days
    • Reality 3-4 hours

    • Proceeds bidirectional
    • Replication rate ~500-5k
    • bp/min

    • E.coli replicates
    • ~1000bp/second

    A lot slower in eukaryotes



    • In large part due to all of the DNA packaging into nucleosomes
    • 1st have to disassemble the nucleosomes to expose the ssDNA template & then reassemble after copying

    Components of DNA replication

    • ssDNA template
    • dNTPs
    • Primer
    • MgH
    • DNA polymerase
  19. WHAT ARE THE COMPONENTS OF DNA REPLICATION
    • -ssDNA TEMPLATE
    • -dNTPs
    • -PRIMER
    • -MGH
    • -DNA POLYMERASE
  20. BASIC CELLULAR COMPONENTS OF DNA REPLICATION
    • A short chain of nucleotides
    • that are complementary to the template DNA (base pairs w/the ssDNA template)
    • *purpose of the primer is to
    • provide a 3'OH group to which DNA polymerase can add a base to via a phosphodiester bond
    • ALL DNA polymerases require
    • a pre-existing 3'OH group




    • MgH ->metal co-factor that stabilizes the DNA
    • DNA polymerase ->enzyme that carries out DNA synthesis
  21. DNA SYNTHESIS
    • SIMPLY THE JOINING OF NUCLEOTIDES IN A POLYNUCLEOTIDE CHAIN BY LINKING THE NUCLEOTIDES W/A PHOSPHODIESTER BOND
    • *THE PRODUCT IS COMPLEMENTARY & ANTI-PARALLEL TO THE TEMPLATE STRAND
    • *DIRECTION OF SYNTHESIS IS ALWAYS 5'-3'
  22. DNA REPLICATION IN E. COLI
    • DNA replication in E.coli
    • begins @ the Ori-C
    • Ori-C
    • -245bp region of the chromosome

    • Contains 2 sets of tandem
    • repeats

  23. STEPS OF BACTERIAL REPLICATION
    • -INITIATION
    • -UNWINDING
    • - PRIMING
    • -ELONGATION
    • -FINISHING TOUCHES
  24. INITATION
  25. Initiation - binding of the HU proteins & initiator protein to
    the series of 9-mer repeats
  26. Results in coiling of the DNA around these proteins (requires ATP for energy)
    • This coiling puts a torsional strain of the double helix
    • This tension needs to be released
  27. UNWINDING
    • Unwinding ->to release the tension caused by coiling w/HU
    • & Initiator protein

    • Get unwinding @ the weak A/T
    • pairing of 13-mers repeats
    • Generates your ssDNA
    • template
    • This unwound state is stabilized by single stranded DNA binding proteins (ssb proteins)

    • Keeps the replication origin
    • open for DNA synthesis
    • Prevents the helix from
    • reforming
    • Further unwinding of the double helix by the enzyme-->

    • Helicase->enzymatically breaks the H-bonds of the double helix
    • as replication proceeds

    *helicase binds to ssDNA @ the replication fork

    • -breaks the H-bonds


    • -removes
    • these supercoiled knots that result from helicase unwinding

    Removes 2 supercoils @ a time

    • As the DNA unwinds
    • -supercoiled knots are formed in front of the replication fork
  28. PRIMING
    • carried out by the enzyme primase (a special
    • type of RNA polymerase)
  29. Primase binds to the helicase @ the replication fork & sets down a short oligonucleotide primer of RNA
    • bases that are complementary to the template DNA
    • Promer provides the 3'OH
    • group for DNAP to add DNA bases to
    • **all DNA synthesis begins
    • w/a short stretch of RNA (to be removed later & replaced w/DNA)
  30. ****ALL DNAPS REQUIRE A PRE-EXISTING 3'OH GROUP BUT RNAPS DON’T
    HAVE THIS REQUIREMENT
  31. ELONGATION
  32. addition of DNA nucleotides complementary to the
    template
  33. **once DNA synthesis is
    • primed, replication proceeds in a 5'-3' direction until a termination signal is encountered or 2
    • replication forks meet
    • ***elongation is carried out
    • by DNAPs
  34. CHARACTERISTICS OF ALL DNAPs
    • Synthesize new strands of DNA that are
    • complementary & anti-parallel to the template
    • By
    • definition they have 5'-3' polymerase activity
    • Use dNTPs
    • as substrates for synthesis
    • Require a
    • primer to initiate synthesis


    (unlike the RNAP primase)


    • Catalyze the formation of a phosphodiester
    • linkage b/w the 3'OH of 1 nucleotide & the 5' PO4 of the incoming, joining nucleotides
  35. WHAT ARE THE 3 MAIN DNAPs INVOLVED IN E.COLI?
    • -DNAPI, DNAPII, DNAPIII
    • -IN ADDITION TO THE (1) 5'-3' POLYMERASE ACTIVITY, SOME DNAPs MAY HAVE ADDITIONAL ACTIVITIES (2) 3'-5' EXONUCLEASE ACTIVITY (INVOLVED IN PROOFREADING/CORRECTING MISTAKES IN SYNTHESIS) (3)5'-3' EXONUCLEASE ACTIVITY (INVOLVED IN REMOVING THE RNA PRIMERS & REPLACING W/DNA
  36. EXONUCLEASE
    ENZYME THAT BINDS TO THE ENDS OF DNA MOLECULES & REMOVES/DEGRADES NUCLEOTIDES (NUCLEASE)
  37. 3'-5' EXONUCLEASE
    • BINDS TO THE 3' END & REMOVES NUCLEOTIDES BY MOVING IN A 3'-5' DIRECTION = PROOFREADING ACTIVITY
    • -BACKTRACKS & REMOVES THE BASES 3'-5' INCLUDING THE MISINCORPORATION
  38. DNAP III
    • -PRIMARY FXN
    • *MAIN WORKHORSE OF DNA REPLICATION.
    • *ACTIVITY @ THE REPLICATION FORK ->SYNTHESIZED THE DNA
    • -5'-3' POLYMERASE - YES
    • -3'-5' EXONUCLEASE - YES (PROOFREADING)
    • - 5'-3' EXONUCLEASE - NO
  39. DNAP I
    • -PRIMARY FXN
    • *REMOVES THE RNA PRIMERS (W/5'-3'EXONUCLEASE) & REPLACES W/DNA (5'-3'POLYMERASE)
    • -5'-3' POLYMERASE - YES
    • -3'-5' EXONUCLEASE - YES
    • -5'-3' EXONUCLEASE - YES
  40. DNAP II
    • -PRIMARY FXN
    • *POST-REPLICATION REPAIRS/PROOFREADING ->CATCHES THOSE MISTAKES NOT CORRECTED BY DNAP III
    • -5'-3' POLYMERASE - YES
    • -3'-5' EXONUCLEASE - YES
    • -5'-3' EXONUCLEASE - NO
  41. WHY IS DNA REPLICATION ALSO KNOWN AS SEMI-DISCONTINUOUS??
    • The strands of the double helix areanti-parallelBut, DNAsynthesis occurs only 5'3'Therefore,DNA synthesis proceeds in opposite directions (physically not biologically5'-3') on the parental strandsOne strandis said to be synthesized - continuously = leading strandThe otherstrand is synthesized - discontinuously = lagging strandThe lagging strand is madein a series of fragments = Okazaki fragment
  42. FINISHING TOUCHES
    • Removal of the RNA primers
    • & replacing them w/DNA =DNAP I

    • Chews away the RNA 5'-3'
    • while replacing w/DNA 5'-3'

    • Uses the 3'OH group of the
    • proceeding Okazaki fragment to initiate synthesis




    • Joining of the Okazaki fragments on
    • the lagging strand

    • DNA ligase=molecular glue
    • ->joins Okazaki fragments

    • Joins 2 DNA fragments w/a
    • phosphodiester linkage
    • *but it cannot add bases
    • (not a polymerase)

    DNA ligase seals the nick
  43. FIDELITY OF DNA REPLICATION
    • ACCURACY
    • ~OVERALL ERROR RATE OF 1MISTAKE/BILLION BASES
    • ~DNAPIII MAKES MISTAKES ~1/1000000BASES
    • ~PROOFREADING ACTIVITY CORRECTS SOME OF THESE MISTAKES = 1/1000000 CHANCES
    • ~POST-REPLICATIVE REPAIR (DNAPII) - 1/BILLION MISTAKES
  44. HAYFLICK LIMIT
    • (ASSOCIATED W/CELLULAR AGING)
    • -THE CHROMOSOME GETS SHORTER W/EACH REPLICATION/DIVISION CYCLE
  45. TELOMERASE
    • THE BODIES WAY OF FIGHTING BACK AGAINST CHROMOSOME SHRINKAGE
    • -AN ENZYME THAT BINDS TO THE ENDS OF LINEAR CHROMOSOMES (TELOMERES) & ADDS ON TANDEM COPIES OF THE TELOMERIC REPEAT SEQUENCE
    • ~IT FIGHTS BACK AGAINST SHRINKAGE BY ELONGATING & ADDING MORE REPEATS
    • -RIBONUCLEIC PROTEIN (RNA COMPONENT & PROTEIN COMPONENT)
    • ~SPECIFICALLY RNA DEPENDENT DNAP
    • ~USES AN RNA TEMPLATE TO DIRECT THE SYNTHESIS OF COMPLEMENTARY DNA STRAND
    • ~IT USES ITS OWN RNA AS THE TEMPLATE
  46. DNA RECOMBINATION
    • ~HOMOLOGOUS
    • ~NONHOMOLOGOUS
  47. HOMOLOGOUS DNA RECOMBINATION
    • COMMON IN GERMLINE CELLS THAT ARE UNDERGOING MEISOSIS (MEIOSIS I)
    • ~THE MECHANISM IS NOT COMPLETELY UNDERSTOOD-BUT THERE IS A MODEL FOR THIS RECOMBINATION BASED ON OBSERVABLE INTERMEDIATES - HOLLIDAY MODEL
  48. NON-HOMOLOGOUS DNA RECOMBINATION
    NO HOMOLOGY INVOLVED IN THIS GENETIC EXCHANGE -> COMMON IN SOMATIC CELLS THAT DO NOT GO THROUGH MEIOSIS
  49. HOLLIDAY MODEL
    • MODEL OF HOMOLOGOUS DNA RECOMBINATION BASED ON OBSERVABLE INTERMEDIATES
    • 1.RECOGNITION & ALIGNMENT
    • 2.SINGLE-STRAND BREAKAGE
    • 3.CROSSING OVER & STRAND INVASION
    • 4.JOINING OF THE END (DNA LIGASE)
    • 5.BRANCH MIGRATION & HETERODUPLEX FORMATION
    • 6.CLEAVAGE & RESOLUTION
  50. HOLLIDAY MODEL
    1.RECOGNITION & ALIGNMENT
  51. HOLLIDAY MODEL
    2.SINGLE-STRAND BREAKAGE
  52. HOLLIDAY MODEL
    3.CROSSING OVER & STRAND INVASION
  53. HOLLIDAY MODEL
    4.JOINING OF THE END (DNA LIGASE)
    *FORMS HOLLIDAY JUNCTION
  54. HOLLIDAY MODEL
    5.BRANCH MIGRATION & HETERODUPLEX FORMATION
  55. HOLLIDAY MODEL
    6. CLEAVAGE & RESOLUTION
    • -HAVE VERTICAL AND HORIZONTAL CLEAVAGE PRODUCTS
  56. VERTICAL CLEAVAGE PRODUCTS IN THE HOLLIDAY MODEL
  57. HORIZONTAL CLEAVAGE PRODUCTS IN THE HOLLIDAY MODEL
  58. WHAT IS DNA'S PRIMARY FUNCTION?
    • to store the cellular info in a stable form ->such that it isn't
    • lost when a cell divides & replicates
    • The DNA sequence is a code that when deciphered directs
    • the assembly of amino acids intoproteins
    • It is the proteins that carry out all the
    • cellular/biochemical processes.


    • -->
    • proteins control the phenotype


    • Although DNA stores all this
    • info -->it is NOT the primary template used for protein synthesis.
  59. GENE EXPRESSION
    • -the term used to describe the process of taking the info in DNA, & using it to direct the synthesis of proteins.
    • -there are 2 main processes in gene expression
    • *TRANSCRIPTION
    • *TRANSLATION
  60. TRANSCRIPTION
    the synthesis of ssRNA (mRNA) from a DNA template -> produces an mRNA molecule that is complementary & anti-parallel to the DNA strand

    • 5'-3' synthesis of ssRNA from a dsDNA template
    • *Although the mRNA is transcribed from dsDNA template,

    --->only 1 strand of the double helix actually serves as the template

    • mRNA has the same polarity as the non-template strand
    • mRNA has the same sequence as
    • the non-template strand except T are replaced by U

    • b/c of this = the nontemplate strand is also referred
    • to as the "sense" strand

    Template strand = antisense

    Process of transcription is similar to replication

    ------>5' - 3' synthesis of a polynucleotide chain from dsDNA
  61. TRANSLATION
    • synthesis of a polypeptide (protein) from an
    • mRNA template
  62. CENTRAL DOGMA OF MOLECULAR
    BIOLOGY
    • --->FLOW OF GENETIC INFORMATION
  63. REPLICATION VS TRANSCRIPTION
  64. 4 TYPES OF RNA PRODUCED BY TRANSCRIPTION
    • Messenger RNA
    • (mRNA) - encodes the amino acid sequence that
    • determines a protein

    • mRNA is copy of the gene coding region
    • Define protein sequence

    • Transfer RNA
    • (tRNA) - a functional RNA molecule that serves as an
    • adapter that links the amino acids with the correct coding in Mrna
    • Ribosomal RNA (rRNA) - structural & functional compiment of ribosomes

    Small nulcear RNAs (smRNA) - involved in splicing eukaryotic genes
  65. 3 BASIC STEPS OF TRANSCRIPTION
  66. Initiation
    • Elongation
    • Termination
  67. 3 REGIONS OF A TRANSCRIPTION UNIT
  68. Promoter - initiation point for transcription =RNA binding site
    • RNA coding sequence - gene coding region
    • Terminator -where transcription terminates
  69. PROKARYOTIC TRANSCRIPTION UNIT
  70. CISTRON
    -gene coding region
  71. transcription initiation
    • begins with the
    • RNAP binding to the promotor

    • The core enzyme of the RNAP is a tetrameric protein
    • comprised of 4 polypeptide subunits

    • 2 molecules of alpha subunit
    • 1 molecule of beta subunit
    • 1 moluecule of beta prime
    • subunit
    • Shorthand Α2ββ'

    • Core RNAP - responsible for catalyzing elongation by
    • joining rNTPs with a
    • phosphodister bond

    Enzymatic component of the RNAP

    • Inititation
    • is a 2-step process

    Loose binding of the holoenzyme to the -35 site

    This binding facilitates localized unwinding at the A/T rich pribnow box (-10 site)

    Creates a "bubble" of ssDNA template

    • Results in tight binding to the now available ssDNA
    • Centered over the -10 site
  72. holoenzyme
    σ- factor associates w/the core RNAP to form the HOLOENZYME (σ2ββ'α)
  73. PROMOTOR OF TRANSCRIPTION
    • THE
    • SPECIFICITY OF BINDING IS DETERMINED BY THE SEQUENCE OF DNA W/I THE PROMOTOR

    • *IF YOU MUTATE OR
    • CHANGE THESE CONSERVED SEQUENCES -> REDUCE OR ABOLISH TRANSCRIPTION

    • *NOTE: PRIBNOW BOX =
    • A/T RICH

    • WEAK
    • h-BONDS

    • FACILIATATES
    • UNWINDING OF HELIX TO EXPOSE ssDNA template for transcription
  74. TRANSCRIPTION ELONGATION
    • Transcription proceeds in 5'-3' direction until a
    • termination signal is met
    • After 8-10 nt have been
    • synthesized the sigma factor falls off, & the core RNAP (catalytic
    • component) continues on w/transcription until the end


    • ***the
    • sigma factor is recycled to form another holoenzyme that can begin a second
    • round of transcription from the same template

    • ***the association of the mRNA w/the DNA template is temporary (unlike replication)
    • As the RNA synthesized, it dissociates from the template -> DNA helix reforms behind it
    • -> RESULTS: the promotor becomes available again for a sequential round of transcription

    -> Multiple mRNAs can be transcribed simultaneously from the same template

    • (drawing in book) - looks like a christmas tree
    • -> each of these
    • mRNAs can then be simultaneously translated into proteins by the ribosomes

    • Can
    • have multiple ribosomes along same mRNA

    • Very
    • efficient system for making lots of protein

    • ->Polyribosome/polysome
    • => simultaneous translation of a single mRNA by multiple ribosomes
  75. WHY IS THE RATE OF TRANSCRIPTION SLOWER THAN THAT OF REPLICATION?
    • Rate of
    • transcription is approximately 20X slower than replication

    • Transcription
    • is 50nt/sec

    • Replication
    • is 1000nt/sec

    • Slower
    • rate due in part due to single stranded mRNA forming secondary structures due
    • to complementary base pairing - slows down to forward synthesis
  76. *Why do RNAPs NOT have proofreading activity (no 3'-5' exonuclease activity)
    • mRNAs have a short half-life and they get degraded
    • quickly

    • So even if you make a mistake it's not going to be
    • around that long for a bad protein to be made
    • And then right behind that
    • bad mRNA is another copy being synthesized which would probably be a good
    • copy



    • *The lack of RNA
    • proofreading is an advantage to RNA viruses b/c it allows them to evolve
    • quickly
  77. TRANSCRIPTION TERMINATION
    • The termination signal is found downstream of the Gene
    • Coding Region

    • In bacteria there are 2 mechanisms of transcription
    • termination:
    • Self-termination/Rho-independent
    • Rho-dependent
  78. self-termination/Rho-independent transcription termination
    • Based solely on the characteristics of the sequenceat the termination site
    • A non-enzymatic dissociation of mRNA from the DNA template
    • Template characteristicsG/C-rich region that is followed by a poly-A tail (6or more Adenine in a row)
    • G/C -rich region ispalindromic (inverted repeat)-facilitatescomplementary pairing/hairpin formation in the transcribed mRNA.
  79. Rho-dependent transcription termination
    • Enzymatic dissociation of mRNA from the DNA template
    • Rho- enzyme= HELICASE
  80. DISSOCIATES THE mRNA from the template by
    enzymatically breaking H- bonds
  81. G/C rich region - but not followed by a poly A tail
    • Upstream of this region is a
    • rut-site (rho-utilization)
  82. Site where rho binds to the mRNA
    • Once bound, rho moves down
    • the transcript in a 5'-3' direction following the RNAP
  83. When the RNAP reaches the G/C rich region ->slows
    • down (harder to transcribe through
    • the triple bonds of G/C pairs)
    • This allows rho to catch-up
    • & it breaks the H-bonds holding the transcript template - termination
  84. eukaryotic transcription
    • More proteins involved = these proteins are called
    • transcription factors (tfs)

    • A transcription factor is any protein that influences
    • the rate of transcription in either an increase or decrease in the amt of mRNA made




    More RNAPs

    RNAP II that transcribes mRNA




    • The mRNA must go through several processing steps
    • (post-transcriptionally) beofre it is ready for translation pre-mRNA----->mature mRNA
    • Transcription takes place in
    • the nucleus, while translation occurs separately in the cytoplasm =UNCOUPLED

    • For prokaryotes there is no nucleus and transcription
    • and translation occur simultaneously or
    • COUPLED




    Euk. Transcription units =monocistronic


    • Prok.
    • Transcription units = may be polycistronic
  85. eukaryotic transcription unit
  86. EXON
    expressed DNA
  87. INTRON
    interventing DNA
  88. Eukaryotic Promotor binding site
  89. enhancer
  90. regulatory
    • elements that when bound by transcription factors called ACTIVATORS serve
    • to increase the rate of
    • transcription
    • They don't have to be partof the promoter In fact they can be located upstream, downstream oreven w/I the gene itselfAnd they can act over greatdistance
  91. activator
    positive transcription factor - they act bystabilizing RNAP binding to the promotor
  92. silencers
  93. regulatory
    elements similar to enhancers w/regards to position
  94. Except when bound by tfs called REPRESSORS they
    decrease or inhibit transcription by the RNAP
  95. Destabilize RNAP binding to the promoter
  96. EUKARYOTIC 3 PRIMARY PROCESSING STEPS
    • -5' CAPPING
    • -3'POLYADENTATION
    • -SPLICING
  97. 5' CAPPING
    • Shortly after transcription
    • begins, a 7-methyl guanosine cap is added to 5' end of mRNA (McG)
    • Serves 2 functions
  98. Protects the 5' end from
    • degradation
    • Serves as the Ribosome
    • Binding Site (RBS) for translation
  99. (*no cap->no
    translation->no protein made)
  100. 3' POLYADENTATION
    • the addition of 300-500 adenines to the 3' end
    • of transcript
  101. Serves 2 fxns
  102. Protects the 3'end from
    • degredation
    • Involved in transcriptional
    • termination
  103. Ends the transcript lenth
  104. The enzyme responsible =
    poly A polymerase=PAP
  105. A template INDEPENDENT
    polymerase
  106. **adds adenines w/out a
    template of thymines to direct it
  107. At the 3' end of pre-mRNA
    -->there is a conserved consensus sequence that is recognized by a complex of cleavage proteins
  108. SPLICING
    the removal of the non-coding intron & the joining of the coding exons
  109. End result = mature fxnal mRNA that is ready for transport & translation
  110. Splicing is very important
    ->must be done accurately ->to the exact base level
  111. If you improperly join the
    • exons->non-fxnal mRNA ->bad, non-fxnal protein
    • Precise slicing is directed
    • by specific DNA sequences in the mRNA that are located @ the exon/intron boundaries
  112. WHY HAVE INTRONS (HYPOTHESIS)
    • Introns may protect the important coding regions of genes from errors in homologous meitoic recombination
    • Exon shuffling -rearrangement of coding exons @ the DNA level
  113. Rearrange the exons to create new genes
  114. Exons are typically functional domains of a protein
  115. Phosphorylation activity
    • DNA binding domain
    • Nuclease domain
  116. Inherited changes - evolutionarily important
  117. Alternative
    splicing/deferential splicing
  118. The rearrangement of exons @ the RNA level->occurs during splicing of the mRNA
    Result: create more than one mature mRNA from a single pre-mRNA
  119. Single transcript can actually end up coding for more than one protein, depending on how the exons are joined

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