Nucleosome Positioning Histones and DNA Bending

DNA packaging 3 X 10 9 base pairs in human genome ~1 m if unraveled Compacted into nucleus –100  m in diamter

Histones

PDBID = 1KX3

Nucleosome Arrays Dorigo B, et al. Science (5701):

Current Opinion in Structural Biology 2006, 16:336–343

Chromatin Represses Transcription Clark & Wolffe 1991

Histones vs. DNA Histones fold DNA DNA sequence determines histone binding –Does histone bind sequence? Absence of direct sequence specific recognition –Bending flexibility of DNA determines affinity for complementary protein surface

DNA as a Rod

Twist Bend

3 pN, we observe a similar coexistence of B-DNA and a new, highly twisted structure. Experimental data and molecular modeling suggest that this structure has approximately 2.62 bases per turn and an extension 75% larger than B-DNA. This structure has tightly interwound phosphate backbones and exposed bases in common with Pauling's early DNA structure

Major & Minor Groove

Flexibility Torsional Flexibility –Angle between base pairs Bending Flexibility –Properties of individual base steps Flexibility of DNA is a Sequence- dependent property

Flexibility determinants Hydrogen bonds Electronic configuration of base pairs Base Pair Stacking energy Exocyclic groups in major and minor grooves

Base Pairing

Base Pairing

Base Stacking

Base Stacking The structural basis of DNA flexibility.Philos Transact A Math Phys Eng Sci Jul 15;362(1820):

Base Stacking Richmond & Davey 2003

Electrostatics Philos. Transact. A Math Phys. Eng. Sci Jul 15;362(1820):

Major & Minor Groove 12A 22A

DNA wrapping alters helical periodicity Chromatin: Structure and Function Alan P. Wolffe

Sequence-directed positioning Widlund et al Mouse Genome Micrococcal nuclease 146 bp DNA fragments DNA seq added to end Amplify using PCR 10 rounds cycles –competitive nucleosome assembly Chromatin: Structure and Function Alan P. Wolffe

Intrinsically curved DNA sequences Oligo(dA).oligo(dT) tracts (4-6bp) –Phased with a periodicity similar to that of DNA helix TATA boxes repeated every 10 bp CA and CTG di/tri-nucleotide repeats

Energy Landscape

Matrix Method Nucleotide phasing of 10bp repeats influences bending of DNA

Periodic occurrence of A/T & GC A/T & G/C out of phase A/T narrow minor groove G/C wide minor groove Satchwell et al. J. Mol. Biol :659-75

Minor Groove bending facilitated by Arg side chain into minor groove when it faces histone octamer Major groove narrows to ~9Å Minor groove narrows to ~3Å

Richmond & Davey 2003 Overtwisted DNA –Outside Dyad B-DNA conformation with local conformational variation DNA groove –Minor 3A protein-DNA –Major 9A protein-DNA –Twist -> narrow minor groove –Roll 3x vs tilt to cause curvature Smooth bending into the major groove –Uniform positive roll angle and undertwisting at adjacent base steps Smooth bending into minor groove –Constant negative roll angle with alternating twist-shift coupling Relieves steric hindrance between the base edges Discontinuos bending -> Kinking of CA/TG base steps with roll angles and coupled overtwisting –Roll angles reducing the stacking energy –Unstacking energy is less for TA than for TA/TG TA bends the greatesd

Structure of DNA in Nucleosome Discontinuous curvature Richmond & Davey 2003

Phase of Roll & Tilt Positive Roll in Major Groove Positive Tilt in Minor Groove 2:1 Roll/Tilt

Smooth Major Positive roll Under winding –Reduced twist –Negative slide

Minor Groove Smooth Bending –Large shift flux –Constant – Roll –Twist-shift Relieves steric bp edge clash Kinking –Large slide Avoid purine clash –Overtwisted –Always at CA/TG bp steps

Consistent with Studies of DNA in database Hassan and Calladine 1997 CA=TG most flexible as judged from SD of Roll angle Hassan and Calladine 1997

Stacking Energy Kink and Overtwist –CA/TG has low stacking energy –Can adopt high or low roll angles by reducing stacking energy

Summary Ong, Richmond & Davey 2007 Major Gray Minor White

Unusual DNA conformation parameters induced by histone binding Implicates sequence-dependent –protein recognition –nucleosome positioning –nucleosome mobility Can we predict DNA:Nucleosome binding based on sequence?

Consistant with Studies of DNA in database Hassan and Calladine 1997 Rigid and loose (continuously flexible) –Roll slide and twist correlated linearly –RIGID: AA/TT, AT and GA/TC –LOOSE: GG/CC, GC, CG