Welcome to My Molecular Biology Lecture

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Welcome to My Molecular Biology Lecture
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Molecular Biology of the Gene, 5/E --- Watson et
al. (2004)
Part I Chemistry and Genetics Part II
Maintenance of the Genome Part III Expression
of the Genome Part IV Regulation Part V Methods
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Part II Maintenance of the Genome
Dedicated to the structure of DNA and the
processes that propagate (??), maintain (??) and
alter (??) it from one cell generation to the next
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Maintenance of the Genome
Ch 6 The structures of DNA and RNA Ch 7
Chromosomes, chromatins and the nucleosome Ch 8
The replication of DNA Ch 9 The mutability and
repair of DNA Ch 10 Homologous recombination at
the molecular level Ch 11 Site-specific
recombination and transposition of DNA
PROPAGATE MAINTAIN
ALTER
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CHAPTER 6

• The Structures of DNA and RNA

How do the structures of DNA and RNA account for
their functions?
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OUTLINE
1.DNA Structure
2.DNA Topology
3.RNA Structure
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DNA STRUCTURE

• The building blocks and base pairing.
• The structure two polynucleotide chains are
  twisting around each other in the form of a
  double helix.

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DNA building blocks

• DNA STRUCTURE (1)

Base (??) Nucleoside (??) Nucleotide (???) is
the fundamental building block of DNA.
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Bases in DNA
Adenine (A)
Purines
N9
Guanine (G)
Cytosine (C)
pyrimidines
Thymine (T)
N1
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Each bases has its preferred tautomeric form
(Related to Ch 9)
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Waston-Crick pairing
The strictness of the rules for Waston-Crick
pairing derives from the complementarity both of
shape and of hydrogen bonding properties between
adenine and thymine and between guanine and
cytosine.
Maximal hydrogen bonding
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AC incompatibility
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Nucleosides Nucleotides
Nucleoside
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Asymmetric
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3
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A DNA molecule is composed of two antiparallel
polynucleotide chains
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Phosphodiester linkages repeating,
sugar-phosphate backbone of the polynucleotide
chain
DNA polarity is defined by the asymmetry of the
nucleotides and the way they are joined.
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The two strands are held together by base pairing
in an antiparallel orientation a stereochemical
(?????) consequence of the way that A-T and G-C
pair with each other. (Related to replication and
transcription)
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DNA structure

• DNA STRUCTURE (2)

two antiparallel polynucleotide chains are
twisting around each other in the form of a
double helix.
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1. The Two Chains of the Double Helix Have
Complementary Sequences
Watson-Crick Base Pairing
Example If sequence 5-ATGTC-3 on one chain,
the opposite chain MUST have the complementary
sequence 3-TACAG-5
(Related to replication and transcription)
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2. Hydrogen Bonding determines the Specificity of
Base Pairing, while stacking interaction
determines the stability a helix.
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• Hydrogen bonding also contribute to the
  thermodynamic stability of the helix (?)
• Stacking interactions (p-p) between bases
  significantly contribute to the stability of DNA
  double helix

H2O molecules lined up on the bases are displaced
by base-base interactions, which creates
disorder/hydrophobicity.
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3. Two different models illustrate structure a
DNA double helix.
Schematic model
Space-filling model
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4. DNA is usually a right-handed double helix.
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5. The double helix has Minor and Major grooves
(What Why)
It is a simple consequence of the geometry of the
base pair.
(See the Structural Tutorial of this chapter for
details)
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The Major groove is rich in chemical information
(What are the biological relevance?)
The edges of each base pair are exposed in the
major and minor grooves, creating a pattern of
hydrogen bond donors and acceptors and of van der
Waals surfaces that identifies the base pair.
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A H-bond acceptors
D H-bond donors
H non-polar hydrogens
M methyl groups
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6. The double helix exists in multiple
conformations.

• The B form (10 bp/turn), which is observed at
  high humidity, most closely corresponds to the
  average structure of DNA under physiological
  conditions
• A form (11 bp/turn), which is observed under the
  condition of low humidity, presents in certain
  DNA/protein complexes. RNA double helix adopts a
  similar conformation.

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DNA strands can separate and reassociate
DNA STRUCTURE (3)

• Key terms to understand
• Denaturation (??)
• Hybridization (??)
• Annealing/renature (??)
• Absorbance (???)
• Hyperchromicity (???)
• Tm (melting point) (??)

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DNA TOPOLOGY
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Structure (1) Linking number is an invariant
topological property of covalently closed,
circular DNA (cccDNA)
DNA TOPOLOGY (1)
Linking number is the number of times one strand
have to be passed through the other strand in
order for the two strands to be entirely
separated from each other.
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• Species of cccDNA
• Plasmid and circular bacterial chromosomes
• Linear DNA molecules of eukaryotic chromosomes
  due to their extreme length, entrainment (??) in
  chromatin and interaction with other cellular
  components (Ch 7)

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Structure (2) Linking number is composed of
Twist and Writhe
DNA TOPOLOGY (2)
The linking number is the sum of the twist and
the writhe. Twist is the number of times one
strand completely wraps around the other
strand. Writhe is the number of times that the
long axis of the double helical DNA crosses over
itself in 3-D space.
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Local disruption of base pairs
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Function (1) DNA in cells is negatively
supercoiled nucleosomes introduces negative
supercoiling in eukaryotes
DNA TOPOLOGY (3)
Negative supercoils serve as a store of free
energy that aids in processes requiring strand
separation, such as DNA replication and
transcription. Strand separation can be
accomplished more easily in negatively
supercoiled DNA than in relaxed DNA.
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Function (2) Topoisomerases (P115-119)
DNA TOPOLOGY (4)

• The biological importance of topoisomerase?
• The functional difference of the two types of
  topoisomerases?
• The working mechanism of topoisomerase (See the
  animation for detail)

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RNA STRUCTURE
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Biological roles of RNA
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• RNA is the genetic material of some viruses
• RNA functions as the intermediate (mRNA) between
  the gene and the protein-synthesizing machinery.
• RNA functions as an adaptor (tRNA) between the
  codons in the mRNA and amino acids.
• Through sequence complementarity, RNA serves as a
  regulatory molecule to bind to and interfere with
  the translation of certain mRNAs or as a
  recognition molecule to guide many
  post-transcriptional processing steps.
• Through the tertiary structures, some RNAs
  function as enzymes to catalyze essential
  reactions in the cell (RNase P ribozyme, large
  rRNA in ribosomes, self-splicing introns, etc).

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Structures of RNA

• Primary structure
• 2.Sequence complementarity base pairing as DNA
• 3.Secondary structure
• 4. Tertiary structure

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• Primary structure

RNA STRUCTURE
RNA contains ribose and uracil and is usually
single-stranded
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2.Sequence complementarity inter- and
intra-molecular base pairing
RNA STRUCTURE (1)
Watson-Crick base pairing
G-C

A-U
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3.Secondary structures and interactions
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RNA chains fold back on themselves to form local
regions of double helix similar to A-form DNA
RNA STRUCTURE (2)
2nd structure elements
hairpin
RNA helix are the base-paired segments between
short stretches of complementary sequences, which
adopt one of the various stem-loop structures
bulge
loop
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Some tetraloop sequence can enhance the stability
of the RNA helical structures
For example, UUCG loop is unexpectedly stable due
to the special base-stacking in the loop
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Special interactions
3
4
1
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Pseudoknots are complex secondary structure
resulted from base pairing of discontiguous RNA
segments
Figure 6-32 Pseudoknot.
Structurally special base-pairing
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Non-Watson-Crick GU base pairs represent
additional regular base pairing in RNA, which
enriched the capacity for self-complementarity.
Figure 6-33 GU base pair
Chemically special base-pairing
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The double helical structure of RNA resembles the
A-form structure of DNA.
The minor groove is wide and shallow, but offers
little sequence-specific information. The major
groove is so narrow and deep that it is not very
accessible to amino acid side chains from
interacting proteins. Thus RNA structure is less
well suited for sequence-specific interactions
with proteins.
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4. RNA can fold up into complex tertiary
structures
RNA STRUCTURE
Why?

• RNA has enormous rotational freedom in the
  backbone of its non-base-paired regions.

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The structure of the hammerhead ribozyme
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Interactions in the tertiary structure

• Unconventional base pairing, such as base
  triples, base-backbone interactions
• Proteins can assist the formation of tertiary
  structures by large RNA molecule

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The crystal structure of a 23S ribosme
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Some RNAs with tertiary structures can catalyze
RNA STRUCTURE (4)
Ribozymes are RNA molecules that adopt complex
tertiary structure and serve as biological
catalysts. RNase P and self-splicing introns are
ribozymes
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Structure Function The hammerhead ribozyme
cleaves RNA by formation of a 2,3 cyclic
phosphate
RNA STRUCTURE (5)
C17
See animation for detail
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Key points for Chapter 6

• DNA structure
• Building blocks and base pairing
• Double helical structure
• Application of the property of strand separation
  and association in DNA techniques
• Critical thinking how DNA structure influence
  the processes of genome maintenance and
  expression? You are encouraged to take this
  question and find out the answers when we discuss
  the related contents

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• 2. DNA topology
• The biological relevance of cccDNA
• Linking number, twist and writhe how these
  topological features are changed during DNA
  replication answer the question after the
  related lecture.
• Topoisomerases
• 3. RNA structure
• Composition, structure (2nd and tertiary) and
  functions (differences from DNA)