Significance Genome inheritance requires the complete resolution of all intertwines within parental DNA. This is facilitated by fork rotation and precatenation of the newly replicated DNA. However, the general importance and frequency of fork rotation in vivo are poorly understood. We find that the evolutionarily conserved Timeless and Tipin proteins actively inhibit fork rotation in budding yeast. In their presence, fork rotation appears restricted to hard-to-replicate fragile sites. In their absence, excessive fork rotation leads to damage accumulating in the replicated sister chromatids, especially at known yeast fragile sites. Therefore, fork rotation appears to be restricted to contexts where it is absolutely required for unwinding, and this restriction is required to prevent precatenation inducing excessive chromosomal fragility.