Transcript Principles of DNA and RNA Structure PHAR 201/Bioinformatics I Philip E. Bourne Department of Pharmacology, UCSD Prerequisite Reading: Structural Bioinformatics Chapters 3 Thanks to Helen Berman.

Principles of DNA and RNA
Structure
PHAR 201/Bioinformatics I
Philip E. Bourne
Department of Pharmacology, UCSD
Prerequisite Reading: Structural Bioinformatics Chapters 3
Thanks to Helen Berman for many slides
PHAR201 Lecture 2 2012
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We start with DNA
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History
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1946 – DNA is the main constituent of genes (Avery)
1950 – First X-ray pictures of DNA (Franklin)
1953 – DNA structure revealed (Watson and Crick)
1970 onwards - Multiple conformations and structures,
initially from fibers
• 1973 - X-ray structure confirms double helix (Rich)
• 1974 - t-RNA structure (Kim)
• 1980 – Structure of first complete turn of B (“normal”)
DNA (Dickerson)
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What Have we Learnt from These
Structures?
• Hydration, ionic strength and sequence all
impact the type of structure
• We see single stranded helices, double,
triple and quadruple
• Alone DNA and RNA does not crystallize
easily, hence strands are short – eg 10-mer
(unless complexed)
• Contrast this to the ribosome (1FFK)
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DNA and
RNA Structure
NOTE:
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• Components
• Sugar
• Base
• Phosphate
5’ to 3’ direction
T->U in RNA
RNA - extra –OH at 2’ of
pentose sugar
DNA - deoxyribose
Numbering
• Single vs double
strands
• DNA more stable
Voet, Donald and Judith G. Biochemistry.
John Wiley & Sons, 1990, p. 792.
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The 5 Bases
of DNA and RNA
Purines
NOTE:
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Pyrimadines and Purines
T->U in RNA
Names
Numbering
Bonding character
Position of hydrogen
Tautomers
Pyrimadines
Neidle, Stephen. Nucleic Acid Structure and Recognition.
Oxford University Press, 2002, p. 18.
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Tautomeric
Structures
• Keto vs enol (OH)
• Different hydrogen
bonding patterns
Saenger, Wolfram. Principles of Nucleic Acid Structure.
Springer-Verlag New York Inc., 1984, p. 113.
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Geometry of
Watson Crick
Base Pairs
• A:T and G:C pairs are
spatially similar
• 3 H-bonds vs 2 (GC rich?)
• Sugar groups are attached
asymmetrically on the same
side of the pair
• Leads to a major and
minor grove
• Bases are flat but the
hydrogen bonding leads to
considerable flexibility
• Base stacking is flexible
Voet, Donald and Judith G. Biochemistry.
John Wiley & Sons, 1990, p.
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Definition of Major
and Minor Groove
Hydrogen
bonding of WC
base pair
Mechanisms of
recognition
The canonical Watson-Crick base pair, shown as the G-C pair. Positions of the
minor and major grooves are indicated. The glycosidic sugar-base bond is shown
by the bold line; hydrogen bonding between the two bases is shown in dashed
lines.
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Base Stacking is a Major Defining
Feature of DNA Morphology
• Dependant on:
– Nature of the bases and base pairs
– Stacking interactions
• Explains sequence dependant features
• Important for understanding molecular
recognition
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Base
Morphology
The base-pair reference frame is
constructed such that the x-axis
points away from the (shaded)
minor groove edge.
Images illustrate positive values
of the designated parameters.
Reprinted with permission from
Adenine Press from (Lu, et al., 1999).
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Backbone
Conformation
Voet, Donald and Judith G. Biochemistry.
John Wiley & Sons, 1990, p. 807.
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A Beta-nucleoside
• Ring is never flat – has 5
internal torsional angles
• The pucker is determined
by what is bound
• A variety of puckers have
been observed
• Pucker has a strong
influence on the overall
conformation
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The Ribose
Ring is
Never Flat
Voet, Donald and Judith G. Biochemistry.
John Wiley & Sons, 1990, p. 808.
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The Glycosidic Bond
Anti
Syn
• Connects ribose sugar to the base
Neidle, Stephen. Nucleic Acid Structure and Recognition.
PHAR201 Lecture 2 2012 Oxford University Press, 2002,
15 p. 27.
Change in sugar
conformation
affects the
backbone
C3’
C2’
C3’-Endo
C3’
Voet, Donald and Judith G. Biochemistry.
John Wiley & Sons, 1990, p. 808.
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C2’
C2’-Endo
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..and the
position of A DNA
the bases
relative to the
helix axis
B DNA
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Canonical B DNA
Neidle, Stephen. Nucleic Acid Structure and Recognition.
Oxford University Press, 2002, p. 34.
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Canonical B DNA
• First determined experimentally by fiber diffraction
(Arnott)
• C2’-endo sugar puckers
• High anti glycosidic angles
• Right handed – 10 base pairs per turn
• Bases perpendicular to the helix axis and stacked over the
axis
• Overall bending as much as 15 degrees (result of base
morphologies – twist and roll) – {machine learning –
sequence vs overall conformation?}
• Over 230 structures 25 with base mis-pairing – only cause
local perturbations
• Strong influence of hydration along spine
http://ndbserver.rutgers.edu/index.html
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Major vs Minor Groove – distinctly
different environments – important
for recognition and binding
• Major
– Richer in base
substituents
• Minor
– Hydrophobic H atoms
of ribose groups
forming its walls
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Spine of Hydration
Neidle, Stephen. Nucleic Acid Structure and Recognition.
Oxford University Press, 2002, p. 97.
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A DNA
Neidle, Stephen. Nucleic Acid Structure and Recognition.
Oxford University Press, 2002, p. 36.
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Canonical A DNA
Voet, Donald and Judith G.
Biochemistry.
John Wiley & Sons, 1990, p. 800.
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Canonical A DNA
• C3’-endo sugar puckers – brings consecutive
phosphates closer together 5.9A rather than 7.0
• Glycosidic angle from high anti to anti
• Base pairs twisted and nearly 5A from helix axis
• Helix rise 2.56A rather than 3.4A
• Helix wider and 11 base pairs per repeat
• Major groove now deep and narrow
• Minor grove wide and very shallow
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Z-DNA
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Helix has left-handed sense
Can be formed in vivo, given proper sequence and superhelical tension, but
function remains obscure.
Narrower, more elongated helix than A or B.
Major "groove" not really groove
Narrow minor groove
Conformation favored by high salt concentrations, some base substitutions,
but requires alternating purine-pyrimidine sequence.
N2-amino of G H-bonds to 5' PO: explains slow exchange of proton, need for
G purine.
Base pairs nearly perpendicular to helix axis
GpC repeat, not single base-pair
– P-P distances: vary for GpC and CpG
– GpC stack: good base overlap
– CpG: less overlap.
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Zigzag backbone due to C sugar conformation compensating for G glycosidic
bond conformation
Conformations:
– G; syn, C2'-endo
– C; anti, C3'-endo
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Z-DNA
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Z-DNA
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Convex major groove
Deep minor groove
Alternate C then G
Spine of hydration
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Drug complexes to DNA
• Bound to the base pair
– double helix can
accommodate this
• Bound in the minor
grove – show base
specificity
• Cis-platinum drugs
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Quadruplex DNA
1NP9
Jmol
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tRNA
Invariant L-shape
1EVV
jmol
Saenger, Wolfram. Principles of Nucleic Acid Structure.
Springer-Verlag New York Inc., 1984, p. 333.
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tRNA H
bonds
between
distant
regions
Neidle, Stephen. Nucleic Acid Structure and Recognition.
p. 148.
PHAR201 Lecture 2 2012Oxford University Press, 2002,31
The Ribosome
• Complex of protein
and RNA
• Small 30S subunit –
controls interactions
between mRNA and
tRNA
• Large 50S subunit –
peptide transfer and
formation of the
peptide bond
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Putting it all Together –
Major Categories of DNA Binding Proteins
Protein residues that make no
contacts with the DNA are colored
blue, those contacting the sugarphosphate backbone are colored red,
and those making base contacts are
colored yellow. (a) Proteins with a
single binding head: T4
endonuclease V (1vas), PU.1 ETS
domain (1pue). (b) Proteins with a
double binding head: lambda
repressor (1lmb), papillomavirus-1
E2 DNA-binding domain (2bop). (c)
Proteins with an enveloping mode of
binding: NF-kB (1nfk),EcoRI
restriction endonuclease (1eri).
Jones et al. 1999 JMB 287(5) 877
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