Background: The genus Leishmania includes protozoan parasites that are able to infect an array of phlebotomine and vertebrate species. Proteases are related to the capacity of these parasites to infect and survive in their hosts and are therefore classified as virulence factors. Findings: By analyzing protease genes annotated in the genomes of four Leishmania spp [Leishmania (Leishmania) infantum, L. (L.) major, L. (L.) mexicana and L. (Viannia) braziliensis], these genes were found on every chromosome of these protozoa. Four protease classes were studied: metallo-, serine, cysteine and aspartic proteases. Metalloprotease genes predominate in the L. (V.) braziliensis genome, while in the other three species studied, cysteine protease genes prevail. Notably, cysteine and serine protease genes were found to be very abundant, as they were found on all chromosomes of the four studied species. In contrast, only three aspartic protease genes could be detected in these four species. Regarding gene conservation, a higher number of conserved alleles was observed for cysteine proteases (42 alleles), followed by metalloproteases (35 alleles) and serine proteases (15 alleles). Conclusions: The present study highlights substantial differences in the organization of protease genes among L. (L.) infantum, L. (L.) major, L. (L.) mexicana and L. (V.) braziliensis. We observed significant distinctions in many protease features, such as occurrence, quantity and conservation. These data indicate a great diversity of protease genes among Leishmania species, an aspect that may be related to their adaptations to the peculiarities of each microenvironment they inhabit, such as the gut of phlebotomines and the immune cells of vertebrate hosts.