PHAR201 Lecture 2 20121 Principles of DNA and RNA Structure PHAR 201/Bioinformatics I Philip E....

PHAR201 Lecture 2 2012 1

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


We start with DNA


History

• 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)


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)


NOTE:

• 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.

DNA and RNA Structure


NOTE:

• Pyrimadines and Purines• T->U in RNA• Names• Numbering• Bonding character• Position of hydrogen• Tautomers

Neidle, Stephen. Nucleic Acid Structure and Recognition.Oxford University Press, 2002, p. 18.

The 5 Basesof DNA and RNA

Purines

Pyrimadines


• Keto vs enol (OH)• Different hydrogen

bonding patterns

Saenger, Wolfram. Principles of Nucleic Acid Structure.Springer-Verlag New York Inc., 1984, p. 113.

Tautomeric Structures

• 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

Geometry of Watson Crick

Base Pairs

Voet, Donald and Judith G. Biochemistry.John Wiley & Sons, 1990, p. 797.8PHAR201 Lecture 2 2012


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.

Definition of Major and Minor Groove


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


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).


Backbone Conformation



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



The Ribose Ring is

Never Flat

PHAR201 Lecture 2 2012 15Neidle, Stephen. Nucleic Acid Structure and Recognition.

Oxford University Press, 2002, p. 27.

The Glycosidic Bond

• Connects ribose sugar to the base

AntiSyn



Change in sugar conformation

affects the backbone

C2’-Endo

C3’-Endo

C3’

C3’

C2’

C2’


A DNA

B DNA

..and the position of the bases

relative to the helix axis



Canonical B DNA


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


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



Spine of Hydration



A DNA



Canonical A DNA


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


Z-DNA• 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.

• Zigzag backbone due to C sugar conformation compensating for G glycosidic bond conformation

• Conformations: – G; syn, C2'-endo – C; anti, C3'-endo


Z-DNA


Z-DNA

• Convex major groove

• Deep minor groove

• Alternate C then G

• Spine of hydration


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


Quadruplex DNA

1NP9Jmol


Saenger, Wolfram. Principles of Nucleic Acid Structure.Springer-Verlag New York Inc., 1984, p. 333.

tRNA

1EVVjmol

Invariant L-shape

PHAR201 Lecture 2 2012 31Neidle, Stephen. Nucleic Acid Structure and Recognition.

Oxford University Press, 2002, p. 148.

tRNA H bonds

between distant regions


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


Putting it all Together –Major Categories of DNA Binding Proteins

Jones et al. 1999 JMB 287(5) 877

Protein residues that make no contacts with the DNA are colored blue, those contacting the sugar-phosphate 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).

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