Lon is an AAA+ serine-lysine protease that plays a crucial rolein protein quality control in both eukaryotic and prokaryoticorganisms. Dysregulation of Lon is associated with variousdiseases. Despite its ubiquity, the mechanisms governingregulation of Lon remain poorly understood. While Lon isfunctionally active as a hexameric ring, evidence suggests that it can also form larger complexes, which may represent differentregulatory states. Here we investigated the formation andstructural basis of higher-order oligomers of E. coli Lon. UsingcryoEM and mass photometry, we demonstrated the existence ofdistinct oligomeric states, including the dodecamer (12mer),undecamer (11mer), and decamer (10mer). High-resolutionstructural analysis reveals that the 12-, 11- and 10mer states areformed by the arrangement of hexameric and pentameric rings ofthe protomers as (6+6), (6+5), and (5+5) assemblies,respectively. The interfaces between these rings are mediated inoligomers by intricate interactions involving the N-terminaldomain (NTD) of Lon. Notably, the NTD interfaces differsignificantly among the three configurations. The 12mer adopts asymmetrical arrangement with a contiguous channel connectingthe two hexamers, whereas the 11mer and 10mer exhibit anangular arrangement in which the NTD blocks the formation ofa continuous channel. Additionally, we have determined thestructures of these oligomers in complex with nanobodies thatstabilize the flexible interface regions, leading to improvedresolution and identification of low-populated states. We alsodemonstrated that Lon proteases from various organisms formsimilar oligomeric assemblies. Based on our experimentalevidence we hypothesize that formation of higher orderoligomers is driven by NTDs, and these oligomers representtransition states of Lons at various stages of their functioning,leading to regulation of Lon activity in cells.