To inve
stigate potential
structure
s ofd(CGG/CCG)
n that might relate to theirbiological functionand a
ssociation with triplet repeat expan
sion di
sea
se
s (TRED
s), the
structure of a
single-
stranded (
ss)oligonucleotide containing d(CCG)
15[
ss(CCG)
15] wa
s examined by
studie
s of the pH andtemperaturedependence of electrophoretic mobility, UV ab
sorbance, circulardichroi
sm, chemical modification, andP1 nuclea
se dige
stion.
ss(CCG)
15 had an unu
suallyhigh p
Ka (7.7 ± 0.2). At pH 8.5,
ss(CCG)
15 formeda relatively un
stable (
Tm = 30
s/entitie
s/deg.gif">C in 1 mMNa
+) hairpin containing CpG ba
se-pair
step
s. At pH7.5, thehairpin contained protonated cyto
sine
s but no detectableC·
+C ba
se pair
s, increa
sed thermal
stability(
Tm= 37
s/entitie
s/deg.gif">C), increa
sed
stacking of the CpG ba
se-pair
step
s, and a
single cyto
sine that wa
s flipped awayfrom the central portion of the helix. Examination of
ss(CCG)
18 and
ss(CCG)
20, which werede
signed toadopt hairpin
s containing alternative GpC ba
se-pair
step
s, revealedhairpin
s containing CpG ba
se-pair
step
s, p
Kas of ~8.2 and ~8.4, re
spectively,and di
storted helice
s. The re
sult
s sugge
st that DNA
sequence
scontaining (CCG)
ns/entities/ge.gif">15adopt hairpin conformation
s that contain CpG rather than GpC ba
se-pair
step
s; themi
smatched cyto
sine
s are protonated at phy
siological pH but are notH-bonded. We propo
se thatprotonation ari
se
s from the
stacking of two cyto
sine
s in the minorgroove of a di
storted helix.