Nucleosides, Nucleotides &
Nucleic Acids
•  Genetic (heritable) information of the
cell
•  Also have roles as
–  energy transfer molecules
–  cofactors/coenzymes
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Nucleotides
•  Building block of the
nucleic acids
•  3 characteristic
components
–  Nitrogen containing base
–  Pentose sugar
–  Phosphoryl group
•  If no phosphoryl group,
then nucleoside
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Nitrogenous bases
•  Heterocyclic ring
•  Two parent components
–  Purine
–  Pyrimidine
•  Sugar link at N
–  N9 purines
–  N1 pyrimidines
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Pentose Sugar
•  Pentose sugar in β-furanose
form
•  Numbered by carbohydrate
convention
–  Distinguished from base
numbering by ‘prime’ (´)
•  Ring is puckered
•  Linked to the nitrogenous base
at C1´ position
–  N-β-glycosidic bond
•  Classification based upon
which sugar is present
–  Ribose or deoxyribose
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Phosphate Group
•  Nucleotides have
anywhere from 1 to 3
phosphoryl groups
•  Linked to sugar
•  Named by number of
phosophates
–  Mono-, di-, tri-
•  Position of bonds identified
by sugar position
–  5’
–  2’3’ cyclic monophosphate
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3
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Cellular Roles
•  Nucleic acid constituents
–  Genetic information
–  RNA, DNA
•  Components of enzyme cofactors
•  Energy currency
•  Act as messengers
–  Link cellular responses to extracellular
stimuli (I.e.hormones)
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Nucleic Acids
•  Covalently linked nucleotides by
phosphoryl group bridges
•  The 5’ phosphate linked to 3’ hydroxyl
of next nucleotide
–  Phosphodiester linkages
–  backbone is alternating phosphate and
pentose residues
–  Nitrogenous bases, “side-group”, face out
at regular intervals
•  Phosphate is completely ionized
•  Sugar hydroxyl H-bonded to water
•  Nitrogenous bases are hydrophobic
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Nomenclature
•  Oligonucleotide
–  Normally less
than 50 bases
•  Polynucleotide
–  Termed nucleic
acid
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Directional molecule
•  Phosphodiester links
have same orientation
giving the linear nucleic
acid specific polarity
•  Schematic
representation via line
drawing
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DNA vs RNA
•  Based upon the identity of the sugar
•  Both subject to slow hydrolysis of the
phosphodiester bond
•  Basic conditions will rapidly hydrolyze
RNA
–  Hydroxyl on 2’ position makes 3’
phosphodiester susceptible to hydrolysis
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Structure/Composition
•  DNA stores genetic information
•  1868 first isolation of “nuclein”
–  Fredrich Mieschner suspected that it was
responsible for inheritance
•  1st direct evidence of DNA being
heritable information molecule
–  Avery-MacLeod-McCarty experiment 1944
–  Hershey-Chase experiment 1952
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Chargaff’s Rules
•  Distinctive base composition noted
•  Proposed in 1940’s
–  Base composition of DNA varies by species
–  DNA from different tissues, but same species
same
–  Base composition doesn’t change due to nutrition/
age/environment
•  Regardless of species, for all cellular DNA
–  A=T; C=G
–  A+G=C+T
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•  Non-direct evidence of
DNA inheritance
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Hershey-Chase
•  Identification of which
part of viral particle
infects the cell
•  Direct evidence of
material transferred
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Watson Crick Double Helix
Structure
•  Knew prior information
–  Chargaff’s rules
–  Rosaline Frank/Maurice
Wilkins X-ray diffraction pattern
•  Helices, two perodices along
long axis
•  One 3.4 Å, 2nd 34 Å
•  Proposed a model that corelated all the information
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Model proposed in 1953
•  2 helical chains wound around same axis
–  Right-handed double helix
•  Hydrophilic backbone alternating
deoxyribose and phosphate groups on
exterior
–  C2’ endo conformation of sugar
•  Hydrophobic and nearly planer nucleotide
base close together
–  perpendicular to long axis
•  Pairing yields major and minor grooves
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•  Parallel or antiparallel strands?
–  Antiparallel yields two complementary
structures
•  when A on one strand, T on opposite
–  The complement
•  Double helix held together by:
–  hydrogen bonding between bases
–  base stacking interactions of the ring
structures
•  Hydrophobic interactions of the planer rings
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3 forms of DNA
•  DNA is a relatively flexible
molecule
•  Flexibility due to
–  Possible conformations of the
deoxyribose ring
–  Rotation about the contiguous
bonds that make up the
phosphodiester backbone
–  Free rotation about the C1´-N
glycosyl bond
•  3 structural forms
predominate
–  A,B and Z
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B-form DNA
•  Watson-Crick proposed
model
•  Most stable under
physiological conditions
•  Standard reference molecule
•  Physical parameters
–  10.5 bases per turn
–  3.4 Å rise per base
–  34 Å per turn
–  Right-handed helix
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A-form DNA
•  Favored in solutions devoid
of water
•  Physical parameters
–  11 bases/turn
–  Base plane tilted
•  Deepened major groove
•  Shallower minor groove
•  Uncertain if found
physiologically
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Z-form DNA
•  Radical departure from prior
structures
•  Physical parameters
–  left-handed helix
–  12 bp/turn
–  2.7 Å rise per base
–  Elongated (zig-zag) appearance
–  Purine residues flip to the syn
conformation
•  Found, role is uncertain
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Palindromic sequences
•  Word or phrase that reads the same
forwards or backwards
–  “Level, madam, level!"
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RNA structure
•  Complex structures exist for RNA
molecules
•  RNA molecule has more functions
within the cell
–  mRNA - messenger
–  rRNA - ribosomal
–  tRNA - transfer
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mRNA
•  Messenger RNA carries genetic message
from the nucleus to the cytoplasm
•  mRNA from different genes vary in length,
mRNA from a single gene has defined size
–  Monocistronic vs polycistronic
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•  Stem-loop structures
–  Both double and single
stranded areas
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rRNA
•  Ribosomal RNA
•  Jacob-Monod proposal
–  ribosomes were not manufactured anew each time a
protein was made
–  the ribosomes did not contain the template necessary
for the manufacture
–  Were structures that when supplied with the
necessary building blocks and instruction (mRNA)
•  Conserved within organisms
–  Most conserved gene
–  16S/18S RNA allows taxonomic identification
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tRNA
•  transfer RNA
•  Adaptor molecule allowing interaction
between codon (nucleotide sequence) and
amino acid
•  Has specific 3-dimensional structure
–  Acceptor stem
–  Anticodon stem-loop
–  TΨC loop
–  D (dihydrouridine) loop
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RNA and DNA can be denatured
•  Denaturation - removal of Hbonding
–  Temperature, pH
–  No covalent bonds broken
•  Renaturation called annealing
•  One step
–  Rapid if molecules still associated
at some point
–  “Zipping-up” of molecule
•  Two step
–  Slower, molecules not associated
–  First must associate, then have
regions of complementarity before
the zip can occur
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Tm
•  Temperatures are
content specific
•  G-C higher melting
temp than A-T
•  Greater number of Hbonds
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Transformation
•  Spontaneous loss of groups
–  Deamination
•  exocyclic amino group
–  Depurination
•  Loss of nitrogenous bases
•  Hydrolysis of N-b-glycosyl bond higher for purines than
pyrimidines
•  Pyrimidine dimers induced by UV irradiation
–  Formation of a cyclobutyl ring between two adjacent thymines
•  Methylation
–  Addition of a methyl (-CH3) group
–  Adenine and cytodine more often methylated
–  S-adenosylmethionine methyl group donor
•  Mutation
–  If in DNA, daughter chains will have permanent change45to
the sequence
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Sanger dideoxy sequencing
•  Requires
–  Template
–  Primer
–  All four dNTPs +
ddNTPs radiolabeled
–  Enzyme (polymerase)
•  Label incorporated is
complementary to base
present on template
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Energy Currency
•  Phosphoanhydride bond
between phosphoryl groups in
nucleotidetriphosphates is a
high energy bond
–  Have α, β, γ phosphoryl groups
–  The α-β, β-γ are
phosphoanhydride
–  Sugar to α is a ester
•  Most common carrier of
energy is the adenine base
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Enzyme cofactors
•  Have nucleotide as
part of the structure
•  Nucleotide doesn’t
participate in the
function, but acts as
a handle, allowing
binding energy to
participate in
enzyme-substrate
interaction
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Regulatory molecules/messengers
•  Serve to modulate the
activity of enzymes/
pathways
•  Second messengers as
they are produced in
the cell in response to
message from outside
of the cell
•  Commonly cyclic AMP
(cAMP, 3’5’cAMP) and
ppGpp
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