SURVEY OF BIOCHEMISTRY Nucleic Acids

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SURVEY OF BIOCHEMISTRYNucleic Acids
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PRS Question

• Which statement is INCORRECT?
• B-DNA is double helical
• In DNA, two H-bonds form between adenine and
  thymine
• In DNA, molar amounts of AT molar amounts of
  GC
• DNA strands run antiparallel to each other

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PRS Question

• Which is not found in DNA?
• Thymine
• 2-OH on pentose
• 3-OH on pentose
• Phosphodiester bond

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PRS Question

• The link between the adenine base and the ribose
  sugar occurs
• From N1 of A to the C1 of ribose
• From N3 of A to the C1 of ribose
• From N7 of A to the C1 of ribose
• From N9 of A to the C1 of ribose

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Nucleic Acids

• Basics of Nucleic Acid Structure
• Supercoiling Topoisomerases
• RNA Structure
• Recombinant DNA Technology

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Limitations to DNA Flexibility

• Glycosidic Bond
• Sugar Pucker
• Sugar-Phosphate Backbone

What is a torsion angle and how does it play a
role in biomolecular structure?
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Rotational Degrees of Freedom
Only 2 permissible conformations of basesyn or
anti
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Glycosidic Bond Conformations
Anti conformation is most stablein B-DNA and
A-DNA
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Sugar Pucker

• B-DNA has a C2-endo conformation

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Sugar Pucker

• A-DNA has a C3-endo conformation

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DNA Supercoiling
Increasing supercoiling
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DNA Supercoiling
L T W
Linking Number (L) Number of times one strand
crosses another strand Here the red strand of
DNA crossesthe blue strand of DNA 10 times L 10
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DNA Supercoiling
L T W
Twist (T) Number of times one strand makes a
complete revolution about the helical axis. Here
the red strand of DNA makes 10 revolutions about
the helical axis T 10
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DNA Supercoiling
L T W
Writhe (W) Number of times a duplex makes a
complete revolution about itself. Counterclockwise
turns are negativeClockwise turns are
positive W 0
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Where do we observe supercoiling of DNA?
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Topoisomerases

• Topoisomerases are enzymes that cleave DNA,
  thereby relieving supercoiling tension.
• Type 1A ss breaks
• Type 1B ss breaks
• Type 2 ds breaks

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Types of RNA
Messenger RNA - carries genetic information for
protein synthesis Transfer RNA - covalently
binds amino acids corresponding to its anticodon
sequence Ribosomal RNA - RNA molecules that
play a role in protein synthesis in conjugate
with ribosomal proteins
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Transfer RNA Structures
Carnegie Mellon U.
NIH
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Ribosomal RNA Structures
Frog5S rRNASubunit
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Recombinant DNA Technology

• Goal Introduce foreign gene into a host
  organism
• Reproduction of the gene or gene product
• Function of the new gene product on the host
• Players Gene insert, host, vector used to carry
  gene into host

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Gene Insert

• Genomic DNA
• All of the DNA carrying genetic information for
  normal functioning of an organism
• cDNA
• DNA copied from mRNA
• PCR
• DNA that has been copied from a template in a
  polymerase chain reaction

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What is Polymerase Chain Reaction (PCR)?

• In vitro DNA synthesis
• Components include
• Heat-stable DNA polymerase (Taq polymerase)
• Two Primers (DNA oligonucleotides)
• Deoxynucleotides dATP, dTTP, dCTP, dGTP
• DNA template
• Mg, buffer components, and water

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How does PCR work?
One PCR Cycle