Bio exam 3

  1. DNA characteristics
    • 1.      Information: necessary for construction of entire organism
    • 2.      Replication: genetic material must be accurately copied
    • 3.      Transmission: Successfully passed from cell to cell and parent to offspring
    • 4.      Variation: must be able to identify variations in species/ and w/in species
  2. Levels of DNA structure
    • 1. Nucleotides are the building blocks of DNA (and RNA).
    • 2. A strand of DNA (or RNA)
    • 3. Two strands form a double helix.
    • 4. In living cells, DNA is associated with an array of different proteins to form chromosomes.
    • 5. A genome is the complete complement of an organism’s genetic material
  3. Chromosome
    cellular structures that contain geneticmaterial
  4. Nucleotides
    • 1.Directionalityà 5’ to 3’ in which C is linked to the phosphate
    • 2. phosphate group
    • 3. pentose sugar (deoxyribose in DNA, ribose in RNA)
    •       a. C are #’d starting w/ C to the right of O, cont. in clockwise direction to count
    •       b. Sugar carbons 1’ to 5’
    •       c. Base attached to 1’
    •       d. Phosphate attached to 5
    •       e. Deoxyribose lacks an O at 2’, therefore why called DE-OXY
    •       f. Phosphodiester bond
    •       g. Backbone= sugars+ phosphates
  5. Phosphodiester bond
    2 phosphoester bonds (covalent bonds betw. P and O) in which phosphate groups attach 2 sugars
  6. Nitrogenous base
    • 1.      Base, projects INWARDLY from backbonea.    
    • Purines: double- ring structure                                                           
    •   i. Adenine(A)                                                             
    • ii. Guanine(G)
    • b. Pyrimidine’s: have single ring structure                                                               i. thymine(T)                                                         
    •     ii. cytosine (C )                                                             iii.      uracil (U)
  7. Complementary
    can predict sequence of 1 strand when given itspair
  8. Major Groove
    • the larger length of DNA rotation
    • · Provides location where proteins can bind and effect gene expression
  9. 1953, James Watson and Francis Crick, with Maurice Wilkins, proposed the structure of the DNA double helix
    used pauling's ball and stick model (purine-pyrimidine pairing)

    Rosalind Franklin’s X-ray diffraction results suggested a helical structure with uniform diameter
  10. Erwin Chargoff
    • Results consistently showed that amount of
    • adenine similar to the amount of thymine and amount of cytosine similar to
    • amount of guanine
  11. DNA Replication
    :a mechanism in which DNA can be copied

    • 1.Origin of replication provides an opening
    • called a replication bubble that forms two replication forks

    2.DNA replication proceeds outward from forks

    3.Bacteria have single origin of replication

    4.Eukaryotes have multiple origins of replication

    5.3 different models:


    2. Conservative

  12. origin of replication
    • Site of start point
    • for replication

    1.w/in chromosome
  13. DNA Polymerase
    responsible for covalently linking nucleotides together to form DNA strands

    1. Free roaming Deoxynucleoside triphosphates H bond to exposed bases

    • 2. DNA polymerase breaks bond between 2nd and 3rd phosphates (EXERGONIC), so provides energy for deoxynucleoside monophosphate to attach to previous
    • nucleotide

    3. Then DNA polymerase recognizes phosphate groups of deox. Triphoshpates, so attaches to remaining phosphate to the 3’ end of growing strand
  14. 2 other enzymatic features
    • DNA polymerase unable to begin DNA
    • synthesis on a bare template strand

    o   DNA primase must make a short RNA primer

    o   RNA primer will be removed and replaced with DNA later

    • • DNA polymerase can only work 5’
    • to 3
  15. DNA Primase
    produces primer that begins the DNA replication process
  16. Leading Strand
    produced in same direction that fork is moving

    1. DNA synthesized as one long cont.  molecule

    2. DNA primase makes one RNA primer

    3. DNA polymerase attaches nucleotides in a 5’ to 3’ direction as it slides forward
  17. Lagging Strand
    DNA synthesized 5’ to 3’ but as Okazaki fragments

    1. Synthesis of fragments occurs away from fork
  18. In both strands
    RNA primers will be removed by DNA polymerase and filled in with DNA

    • DNA ligase will join adjacent DNA fragments
  19. DNA Ligase
    joins adjacent lagging strand DNA fragments together
  20. DNA Helicase
    Binds to DNA and travels 5’ to 3’ using ATP to separate strand and move fork forward
  21. DNA topoisomerase
    Relieves additional coiling ahead of replication fork

    • Functions by nicking one strand
  22. Single -strand binding proteins
    keep parental strands open to act as templates
  23. DNA replication = very accurate
    • 1. Hydrogen bonding between A and T or G and C more stable than mismatches
    •  2. Active site of DNA polymerase unlikely to form bonds if pairs mismatched 
    • 3. DNA polymerase removes mismatched pairs 
    • • Proofreading results in DNA polymerase backing up and digesting linkages 
    • • Other DNA repair enzymes
  24. Transcription
    1st step, produces RNA copy of gene (RNA transcript)

    3 stages of transcription

    1.     initiation

    2.     elongation

    3.     termination
  25. Initiation
    Recognition step

    1. In bacteria, sigma factor attaches to RNA polymerase

    2. causes RNA polymerase to recognize promoter region bc sigma factor recognizes promoter region and attaches there!

    3.Stage completed when DNA strands separated near promoter to form open  complex
  26. Elongation Stage
    1. RNA polymerase synthesizes RNA

    • 2. Template strand used for RNA synthesis
    •         – read 3’ to 5’

    • 3.Coding strand is not used
    •          same sequence as the RNA except uracil substituted for thymine

    4. Synthesized 5’ to 3
  27. Template strand
    used for RNA synthesis
  28. Termination
    1. RNA polymerase reaches termination sequence

    2. Causes it and newly made RNA transcript to dissociate from DNA
  29. Which strand is used?
    · Doesn’t matter! template strand Varied (as whether top or bottom) per gene

    · The direction is modified so that the template is always 3’ to 5’ direction, and RNA is 5’ to 3’
  30. Transcription of Eukaryotic Genes
    1. Basic features identical to prokaryotes

    2.However, each step has more proteins

    3. 3 forms of RNA polymerase

    a. RNA polymerase II: transcribes mRNA

    b. RNA polymerase I and III: transcribes nonstructural genes for rRNA and tRNA
  31. Introns
    (only in eukaryotes) intervening DNA sequences that are not translated
  32. Exons
    coding DNA sequences that are contained in mature mRNA
  33. Splicing
    : removal of introns and connection of exons

    1.  Most structural genes will have 1 or more introns
  34. Spliceosome
    removes introns precisely

    1.Composed of snRNPs – small nuclear RNA and proteins

    2. snRNPs attach to 5’ end, branch site, and eventually 3’end to loop and snip the intron so that neighboring exons connect

    3. Alternative Splicing: splicing can occur more than one way to produce different products
  35. rRNA and tRNA are self-splicing
    • (thanks to RNA that does
    • the job)
  36. Ribozyme
    :RNA molecule that catalyzes a chemical reaction

    ·tRNA and rRNA (self-splicing)
  37. Capping
    1.Modified guanosine attached to 5’ end

    Needed for proper exit of mRNA from nucleus and binding to ribosome
  38. poly A tail
    1. 100-200 adenine nucleotides added to 3’ end

    2. Increases stability and lifespan in cytosol

    3. Not encoded in gene sequence
  39. Bacterial mRNA
    1. 5’ ribosomal-binding site

    2. Start codon is usually AUG

    3. Typical polypeptide is a few hundred amino acids in length

    4. 1 of 3 stop codons

    5. Termination or nonsense codons

    6. UAA, UAG or UGA
  40. Translation
    process of producing polypeptides (proteins) in ribosome

    1. Requires more components than transcription

    2. mRNA, tRNA, ribosomes, translation factors

    3. Most cells use a substantial amount of energy on translation

    4. 3 stages: initiation, elongation, and termination
  41. Initiation of Translation
    mRNA, first tRNA and ribosomal subunits assemble

    1. 3 components:

    a. mRNA,

    b. first tRNA 

    c. ribosomal subunits assemble

    2. initiation factors: proteins that facilitate formation of ribosomal components  

    3. Also requires input of energy (GTP hydrolysis)
  42. initiation factors
    proteins that facilitate formation of ribosomal components (in translation)
  43. Elongation
    Synthesis from start codon to stop codon

    1. Aminoacyl tRNA brings a new amino acid to the A site

    a. Binding of tRNA and mRNA occurs due to codon / anticodon recognition

    • b.Fueled by Elongation factors (hydrolyze GTP to provide energy to bind tRNA to A
    • site)

    c. Peptidyl tRNA is in the P site

    d. Aminoacyl tRNA is in the A site

    2.  peptide bond is formed between the amino acid at the A site and the growing polypeptide chain

    • a.    Peptidyl transfer reaction: polypeptide is removed
    • from the tRNA in the P site and transferred to the amino acid at the A site

    b. rRNA catalyzes peptide bond formation

    (ribosome is a ribozyme)

    3. Movement or translocation of the ribosome toward the 3’ end of the mRNA by one codon

    a. Shifts tRNAs at the P and A sites to the E and P sites

    b. The next codon is now at the A spot

    c. Uncharged tRNA exits from E spot
  44. Termination
    : Complex disassembles at stop codon (found in A site) releasing completed polypeptide

    · 3 stop codons: UAA, UAG, UGA

    1. Release factor (protein shaped like tRNA) binds to stop codon at the A site

    2. Bond between polypeptide and tRNA (in P site) hydrolyzed to release polypeptide

    3. Ribosomal subunits and release factors disassociate
  45. Detailed assembly of bacterial ribosome
    1. mRNA binds to small ribosomal subunit facilitated by ribosomal binding sequence

    2. Start codon a few nucleotides downstream

    3. Initiator tRNA recognizes start codon in mRNA

    4. Large ribosomal subunit associates

    5. At the end, the initiator tRNA is in the P site
  46. eukaryotic differences in ribosome assembly
    1. mRNA have Guanosine cap at 5’ end instead of ribosomal binding sequence

    a. recogn. By cap binding sequences

    2. Variable start codon locations

    Usually AUG is start codon
  47. codon
    code of 3 nucleotides that designate an amino acid
  48. start codon
    defines reading frame
  49. Genetic code
  50. mRNA
    1. Codon – set of 3 RNA nucleotides

    2. T of DNA substituted for U of RNA
  51. tRNA
    1.  Allows binding of tRNA to mRNA codon

    2. Different tRNA molecules encoded by different genes

    3. tRNAser carries serine

    4. Common features

    a.  Cloverleaf structure

    b. Anticodon: 3 RNA nucleotides part of tRNA molecule

    c. Acceptor stem for amino acid binding
  52. aminoacyl- tRNA synthetase
    1. Catalyzes the attachment of amino acids to tRNA

    2. One for each of 20 different amino acids

    3. Reactions result in tRNA with amino acid attached or charged tRNA or aminoacyl tRNA

    4. Ability of aminoacyl-tRNA synthetase to recognize appropriate tRNA has been called the second genetic code
  53. second genetic code
    ability of EACH aminoacyl –tRNA synthetase to recognize an appropriate tRNA
  54. Ribosomes
    1.  Prokaryotes have one kind

    2. Eukaryotes have distinct ribosomes in different cellular compartments

    3. Focus on cytosolic ribosomes (most abundant )

    4. Composed of large and small subunits

    • 5. Structural differences between prokaryotes and eukaryotes exploited by antibiotics to inhibit
    • bacterial ribosomes only

    6. Overall ribosome shape determined by rRNA

    7. Discrete sites for tRNA binding and polypeptide synthesis

    8. P site- peptidyl site

    9. A site- aminoacyl site

    10. E site- exit site
  55. Telomeres
    Series of short nucleotide sequences repeated at the ends of eukaryotic chromosomes

    1. Specialized form of DNA replication only in eukaryotes in the telomeres

    2.  3’ overhang: Telomere at 3’ that does not have a complementary strand

    3. DNA polymerase cannot copy the tip of the DNA strand with a 3’end

    4. No place for upstream primer to be made

    5. If this replication problem were not solved, linear chromosomes would become progressively shorter

    6. Telomerase – enzyme attaches many copies of DNA repeat sequence to the ends of chromosomes

    7. Progressive shortening of telomeres correlated with cellular senescence

    8. Telomerase function reduced as organism ages

    9. 99% of all types of human cancers have high levels of telomerase

    10. Telomerase contains RNA (which is complimentary to DNA) so can attach itself to DNA end
  56. Senescence
    point when cells can no longer double
  57. DNA telomerase
    enzyme that prevents DNA shortening by attaching many copies of DNA repeat sequence to ends of chromosomes
  58. 3' overhang
    Telomere at 3’ that does not have a complementary strand
  59. Elizabeth H. Blackburn, Carol W. Greider, Jack W. Szostak:
    The Nobel Prize in Physiology or Medicine 2009 winners

    · "for the discovery of how chromosomes are protected by telomeres and the enzyme telomerase"
  60. cell division
    When cells prepare to divide, chromosomes become even more compact

    1. Euchromatin not as compact

    2. Heterochromatin much more compact

    3.  Metaphase chromosomes highly compact
  61. Extracellular matrix (ECM)
    • Major macromolecules of are proteins and polysaccharides !
    • Proteins form large fibers !
    • Polysaccharides give a gel-like character
  62. anchoring junctions
    hold adjacent cells together 

  63. cell junctions
    • Hold Cells Together Regulate passage of solutes Plants #
    • Middle Lamella #
    • PlasmodesmataAnimals #
    •       Anchoring Junctions #
    •       Tight Junctions #
    •       Gap Junctions
  64. Middle Lamella
    • ECM layer to adhere cell walls of adjacent cells together !
    • First layer to form when cells divide !
    • Rich in pectins
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Bio exam 3