Cyanobacteria are complex prokaryotes, incorporating a Gram-negative cell wall and internal thylakoid membranes (TMs). However, localisation of proteins within cyanobacterial cells is poorly understood. Using subcellular fractionation and quantitative proteomics we produced an extensive subcellular proteome map of an entire cyanobacterial cell, identifying ~67% of proteins in Synechocystis sp. PCC 6803, ~1000 more than previous studies. 1,712 proteins were assigned to six specific subcellular regions. Proteins involved in energy conversion localised to TMs. The majority of transporters, with the exception of a TM-localised copper importer, resided in the plasma membrane (PM). Most metabolic enzymes were soluble although numerous pathways terminated in the TM (notably those involved in peptidoglycan monomer, NADP+, heme, lipid and carotenoid biosynthesis), or PM (specifically, those catalysing lipopolysaccharide, molybdopterin, FAD and phylloquinol biosynthesis). We also identified the proteins involved in the TM and PM electron transport chains. The majority of ribosomal proteins and enzymes synthesising the storage compound polyhydroxybuyrate formed distinct clusters within the data, suggesting similar subcellular distributions to one another, as expected for proteins operating within multi-component structures. Moreover, heterogeneity within membrane regions was observed, indicating further cellular complexity. Cyanobacterial TM protein localisation was conserved in Arabidopsis thaliana chloroplasts, suggesting similar proteome organisation in more developed photosynthetic organisms. Successful application of this technique in Synechocystis suggests it could be applied to mapping the proteomes of other cyanobacteria and single-celled organisms. The organisation of the cyanobacterial cell revealed here substantially aids our understanding of these environmentally and biotechnologically important organisms.