The solution structures of a 21 base long DNA hairpin derived from the
ColE1 cruciform, and an analog possessing a disulfide cross-link brid
ging the terminal bases, have been determined by NMR spectroscopy. The
8 bp long stem of these sequences adopts a B-form helix whereas the f
ive base long single-stranded loop appears to be flexible and cannot b
e represented by a unique static conformation. NOESY cross-peak volume
s, proton and phosphorus chemical shifts, and both homo- and heteronuc
lear coupling constants for the cross-linked hairpin are virtually ide
ntical to those measured for the unmodified sequence, even for the res
idues that are proximal to the cross-link. These results indicate that
both hairpins are structurally isomorphous. Because this cross-link c
an be incorporated site specifically in a sequence independent manner,
and does not appear to alter native conformation, it should prove bro
adly applicable in studies of DNA structure and function.