Stenotrophomonas maltophilia is increasingly emerging as a multiresistant pathogen in the hospital environment. In immunosuppressed patients, these bacteria may cause severe infections associated with tissue lesions such as pulmonary hemorrhage. This suggests proteolysis as a possible pathogenic mechanism in these infections. This study describes a protease with broad specificity secreted by S. maltophilia. The gene, termed StmPr1, codes for a 63-kDa precursor that is processed to the mature protein of 47 kDa. The enzyme is an alkaline serine protease that, by sequence homology and enzymic properties, can be further classified as a new member of the family of subtilases. It differs from the classic subtilisins in molecular size, in substrate specificity, and probably in the architecture of the active site. The StmPr1 protease is able to degrade several human proteins from serum and connective tissue. Furthermore, pan-protease inhibitors such as ␣ 1 -antitrypsin and ␣ 2 -macroglobulin were unable to abolish the activity of the bacterial protease. The data support the interpretation that the extracellular protease of S. maltophilia functions as a pathogenic factor and thus could serve as a target for the development of therapeutic agents.Stenotrophomonas maltophilia, formerly referred to as Xanthomonas maltophilia or Pseudomonas maltophilia (1, 2), is an aerobic nonfermentative Gram-negative bacterium of widespread occurrence. For healthy humans, it is regarded as an opportunistic germ; it has been implicated in a variety of infections without distinctive clinical features (for a review, see Ref.3). However, in immune-compromised patients, particularly those with bone marrow aplasia or receiving intensive chemotherapy, cases of fulminant hemorrhagic pneumonia have been reported, even with fatal outcome (4 -6). In patients not surviving infections with S. maltophilia, histological inspection of the lung tissue revealed massive bleeding caused by damage to the lung epithelium (4). There are further reports demonstrating involvement of this bacterium in massive hemorrhagic processes of the small intestine and the subclavian artery accompanied by severe lesions of the tissue (5, 6). These observations strongly suggest the participation of proteolytic activity, produced by the bacteria, which may damage the infected tissue. Indeed, it is known that members of the Pseudomonaceae express and secrete a variety of proteases (cf. Ref. 7). Whereas the primary function of these enzymes is to provide a source of free amino acids for bacterial survival and growth, there is accumulating evidence that bacterial proteases may play a pathogenic role in the infected host by involvement in tissue invasion and destruction, evasion of host defenses, and modulation of the host immune system (8).The broad administration of antibiotics currently applied in cases of intensive care patients leads to selection of multiresistant S. maltophilia strains. Consequently, these bacteria are found with increasing frequency in the hospital environmen...