Microbial Pathogenesis 1996; 20: 41–55 Molecular characterization of a surface-exposed superoxide dismutase of Mycobacterium avium Vincent Escuyer*, Nadia Haddad, Claude Frehel and Patrick Berche Unite ´ INSERM U411, Faculte ´ de Me ´ decine Necker-Enfants Malades, 156 rue de Vaugirard, 75015 Paris, France (Received July 24, 1995; accepted in revised form October 16, 1995) Escuyer, V. (Unite ´ INSERM U411, Faculte ´ de Me ´decine Necker-Enfants Malades, 156 rue de Vaugirard, 75015 Paris, France), N. Haddad, C. Frehel and P. Berche. Molecular char- acterization of a surface exposed superoxide dismutase of Mycobacterium avium . Microbial Pathogenesis 1996; 20: 41–55. Mycobacterium avium is an intracellular pathogen capable of growing inside the phago- somal compartment of macrophages. In this work, we characterized the superoxide dis- mutase of M. avium , as a putative candidate to resist the oxidative stress. The gene sodA encoding superoxide dismutase (SOD:EC1.15.1.1) from Mycobacterium avium TMC724 was cloned and sequenced. It encodes a 23 kDa protein (207 aminoacids) showing identity with the Mycobacterium leprae SOD (91%) and the M. tuberculosis SOD (83%). This enzyme was functionally expressed in both Escherichia coli and Mycobacterium smegmatis, and identified as a manganese (Mn) SOD on the basis of sequence comparison with other MnSODs from different organisms, and by activity inhibition studies. By indirect im- munogold labeling of M. avium with a mAb directed against M. leprae SOD, the enzyme was found to be exposed at the cell surface of M. avium . It was also shown that SOD was released in supernates of M. avium TMV724 during exponential growth, suggesting a role of this enzyme during interactions with the environment. When SOD was expressed in the non-pathogenic M. smegmatis, it was also exposed at the surface of bacteria and released in supernates, but this was not sufficient to protect this recombinant mycobacterium from the killing mechanisms of macrophages. 1996 Academic Press Limited Key words: Mycobacterium avium , superoxide dismutase. Introduction Members of Mycobacterium avium complex have recently emerged as major opportunistic pathogens in humans, with the advent of AIDS epidemic. 1,2 The frequency of M. avium infections has reached as high as 15–24% of AIDS patients. 3,4 This microorganism is widely spread throughout nature, and can be isolated from surface water, soil and even house dust, 5,6 thus explaining why humans are frequently exposed to this pathogen. 7 Like the other two human pathogens, Mycobacterium tuberculosis and Mycobacterium leprae, M. avium is an intracellular pathogen capable of surviving and growing within the phagosomal compartment of phagocytic cells. 8–11 It is established that M. avium partially prevents the fusion between the phagosome and the lysosomal vesicles. 12,13 However, nothing is known about the molecular mechanisms of the virulence of * Author to whom correspondence should be addressed. 0882–4010/96/010041+15 $12.00/0  1996 Academic Press Limited