The base pair lifetimes and apparent dissociation constants of a 21 ba
se DNA hairpin and an analog possessing a disulfide cross-link bridgin
g the 3'- and 5'-terminal bases were determined by measuring imino pro
ton exchange rates as a function of exchange catalyst concentration an
d temperature, A comparison of the lifetimes and apparent dissociation
constants for corresponding base pairs of the two hairpins indicates
that the cross-link neither increases the number of base pairs involve
d in fraying nor alters the lifetime, dissociation constant, or the op
ened structure from which exchange occurs for the base pairs that are
not frayed. The cross-link does, however, stabilize the frayed penulti
mate base pair of the stem duplex. Significantly, it appears that the
disulfide cross-link is more effective at preventing fraying of the pe
nultimate base pair than is the 5 base hairpin loop, Because this disu
lfide cross-link can be incorporated site specifically, and does not a
dversely affect static or dynamic properties of DNA, it should prove v
ery useful in studies of nucleic acid structure and function.