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Background  Members of the genus  Streptomyces  are Gram-positive bacteria that are used as important cell factories to produce secondary metabolites and secrete heterologous proteins. They possess some of the largest bacterial genomes and thus proteomes. Understanding their complex proteomes and metabolic regulation will improve any genetic engineering approach.    Results  Here, we performed a comprehensive annotation of the subcellular localization of the proteome of  Streptomyces lividans  TK24 and developed the Subcellular Topology of Polypeptides in  Streptomyces  database (SToPSdb) to make this information widely accessible. We first introduced a uniform, improved nomenclature that re-annotated the names of ~ 4000 proteins based on functional and structural information. Then protein localization was assigned de novo using prediction tools and edited by manual curation for 7494 proteins, including information for 183 proteins that resulted from a recent genome re-annotation and are not available in current databases. The  S. lividans  proteome was also linked with those of other model bacterial strains including  Streptomyces coelicolor  A3(2) and  Escherichia coli  K-12, based on protein homology, and can be accessed through an open web interface. Finally, experimental data derived from proteomics experiments have been incorporated and provide validation for protein existence or topology for 579 proteins. Proteomics also reveals proteins released from vesicles that bleb off the membrane. All export systems known in  S. lividans  are also presented and exported proteins assigned export routes, where known.    Conclusions  SToPSdb provides an updated and comprehensive protein localization annotation resource for  S. lividans  and other streptomycetes. It forms the basis for future linking to databases containing experimental data of proteomics, genomics and metabolomics studies for this organism.    Electronic supplementary material  The online version of this article (10.1186/s12934-018-0892-0) contains supplementary material, which is available to authorized users.

Comprehensive subcellular topologies of polypeptides in Streptomyces