Abstract Filamentous bacteria of the genus Streptomyces possess linear chromosomes and linear plasmids. Theoretically, linear replicons may not need a decatenase for post-replicational separation of daughter molecules. Yet, Streptomyces contain parC and parE that encode the subunits for the decatenase topoisomerase IV. The linear replicons of Streptomyces adopt a circular configuration in vivo through telomere–telomere interaction, which would require decatenation, if the circular configuration persists through replication. We investigated whether topoisomerase IV is required for separation of the linear replicons in Streptomyces. Deletion of parE from the Streptomyces coelicolor chromosome was achieved, when parE was provided on a plasmid. Subsequently, the plasmid was eliminated at high temperature, and ΔparE mutants were obtained. These results indicated that topoisomerase IV was not essential for Streptomyces. Presumably, the telomere–telomere association may be resolved during or after replication to separate the daughter chromosomes. Nevertheless, the mutants exhibited retarded growth, defective sporulation and temperature sensitivity. In the mutants, circular plasmids could not replicate, and spontaneous circularization of the chromosome was not observed, indicating that topoisomerase IV was required for decatenation of circular replicons. Moreover, site-specific integration of a plasmid is impaired in the mutants, suggesting the formation of DNA knots during integration, which must be resolved by topoisomerase IV.