The genus Borrelia includes the causative agents of Lyme disease and relaps
ing fever. An unusual feature of these bacteria is a genome that includes l
inear DNA molecules with covalently closed hairpin ends referred to as telo
meres, We have investigated the mechanism by which the hairpin telomeres ar
e processed during replication. A synthetic 140 bp sequence having the pred
icted structure of a replicated telomere was shown to function as a viable
substrate for telomere resolution in vivo, and was sufficient to convert a
circular replicon to a linear form. Our results suggest that the final step
in the replication of linear Borrelia replicons is a site-specific DNA bre
akage and reunion event to regenerate covalently closed hairpin ends. The t
elomere substrate described here will be valuable both for in vivo manipula
tion of linear DNA in Borrelia and for in vitro studies to identify and cha
racterize the telomere resolvase.