DNA Gymnastics Flashcards
bDNA twist
10.5 bp/turn
what must happen when nucleosome bends dna?
E bend from E torsion and E stacking must be counterbalanced by E bend
measure of flexibility of DNA
persistence length: length of DNA which can form a complete circle without strain @ temp T
525 bp for bdna
E input vs. bp
picture
straight dna
made of
defined as
poly a poly t
base pair midpoint aligns on axis
DNA is locally flexible
bend over a few bp
find angle by drawing line thru midpoints.
\/ apex of theta
can still bp all
bending and grooves
bending always changed groove size
good to fit things like protein pieces
kink
sharp bend: bp are interrupted
what allows protein to bind nucleics (physically)?
bend open DNA - can intercalate: replace base stacking with aromatic
bendable vs bent
independent of each other
bendable: easily deformed, determined by stacking E
bent: lowest energy conformation isnt straight; determined by base-step roll angle
base pair (A or C, etc) determine bendability
parameters that independently define base-step parameters
roll and slide!
which base steps are the flattest? ie., have the lowest___
lowest roll angle
RpR and YpY
YpR and RpY roll angle relation
are opposites!
bent base steps can be straight if
the overall is straight even if individual base steps have nonzero roll angle
roll of bDNA
slightly negtive-> slight negative writhe
how to determine bendability
distribution of crystal structure has large variance
flexibility of DNA :=
how easy to deform a base step away from characteristic geometry
related to stacking energy
slide vs. roll chart tells us:
picture poly A poly T straight stiff CpA TpG TpA most flexible stiff bents exist CpG most bent naturally towards major (major compresse)
circular permutation assay
finds where the bend is
cleave circle with 1 bend at different places-> linear with bent at different place
fastest has bend on end bc has lower radius of gyration
bend site phasing: what happens if there is more than 1 bend?
phase is an angle aroud the mid axis of helix
bp are in phase if htey face same direction
ie, if angular distance =n360/10.5
if bends are in phase, the whole is more bent/less straight
electrophoretic mobility vs. bend site phase
minima whenever in phase
a-tract bending paradox A5T5
(cT4A4g)n straight but (cA4T4g)n highy bent
A tracts are very straight (AAAAA/TTTTT)
A tracts have very narrow minor grooves; locally bent, but a tracts bent out of phase, so overall straight
DNA looping
short persistance -> bendable -> form loops easily
has less to do with sequence; more with topological constrained
topological contraight if closed or end contrained (chromosomes); linear or knicked circles has no toppo constraints-only persistence length
supercoil density
Writhe/turn: 1 supercoil per 20-25 bp
can store E in supercoil like spring- release 10kcal/mol=E/ATP
plectonemic
paranemic
toroidal
angle between outer strand and loop dna \0/
mmore than 90 (small loop) smaller than 90 (large loop)
Linking number=
DNA twists + write (supercoil number)
how to change twist when lk constant
unwinding
change from B to Z or A( z larger because left handeD)
intercalating agents
topoisomerases that ceave and religate
2D gel to discern +- supercoiling
- run gell 1 D: more supercoiled will run farther
2. add ethidium bromide(-2 T->+2w) and run 2nd D
