Interaction of Ruthenium II Polypyridyl Complexes with DNA

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Interaction of Ruthenium (II) Polypyridyl
Complexes with DNA

• Japeck Tang
• Ch221A
• March 15, 2007

Mihailovic A, et al. Exploring the interaction of
ruthenium(II) polypyridyl complexes with DNA
using single-molecule techniques. LANGMUIR 22
(10) 4699-4709 MAY 9 2006.
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Outline

• Introduction
• Theory
• Overstretching B-DNA
• Ruthenium Complexes
• Experimental Setup
• Results
• Conclusion

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Introduction

• Binding of Ruthenium(II) Polypyridyl Complexes
• Used to study DNA structure, small molecule
  binding
• Noncovalent (electrostatic/van der Waals)
• Intercalation
• vs electrostatic/groove binding
• Lengthens DNA (observe binding rate)
• Stabilizes DNA (increased melting temperature)
• Study binding via AFM optical tweezers

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Theory (B-DNA)

• Overstretching
• Normal 3.4Å/bp
• Overstretched 5.8Å/bp
• Transition force 65pN
• Max force 100pN

Smith SB, Cui YJ, Bustamante C. Overstretching
B-DNA The elastic response of individual
double-stranded and single-stranded DNA
molecules. SCIENCE 271 (5250) 795-799 FEB 9 1996.
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Theory (Ruthenium)

• Various Complexes
• Photophysical/redox properties
• Absorb visible light (changes when bound)
• Dipyridophenazine (dppz) light switches
• Oxidize guanine bases

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Theory (Ruthenium)
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Experimental Setup

• Methods of Characterization
• AFM Measurements
• Immobile DNA on mica surface
• Manually trace and measure lengths
• Optical Tweezers
• DNA stretching
• Shift in force-extension curves

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Experimental Fitting (AFM)

• Binding equilibrium
• Binding constant

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Experimental Fitting (AFM)

• Fit data using AFM measured DNA lengths
• Gaussian fit ? average length
• Assume 0.34nm lengthening/intercalation, 172pM
  DNA strands

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Experimental Fitting (AFM)

• Given initial Ru concentration Ru0
• 750nM base pairs, neighbor exclusion (n)
• Solve for Kb and n

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Experimental Fitting (Optical)

• McGhee-von Hippel model for binding
• T fractional occupancy of Ru to DNA
• b extension/bp n binding site size in bp c
  Ru
• Solve for Kb, n given measured T

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Results (AFM)
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Results (AFM)
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Results (Optical)
Ru-(phen)2dppz2
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Results (Optical)
Ru-(phen)32
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Results (Optical)
Ru-(bpy)32
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Summary of Results

• Kb lower for AFM due to Mg2
• DNA stretching is better for weakly binding
  complexesAFM limited to lower concentrations due
  to curling
• Ru(phen)32 binds 3 orders weaker than
  Ru(phen)2dppz2

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Conclusion

• Ruthenium(II) polypyridyl complexes can be used
  to study DNA (photophysical/redox)
• Intercalation lengthens and stabilizes DNA
• Intercalation can be observed, and binding
  constants determined via either AFM or optical
  tweezers measurements
• Optical tweezers are better than AFM for weakly
  binding complexes