Subcellular localisation and sedimentation behaviour of antigen 60 from Mycobacterium bovis BCG

Abstract

Preparation, composition and immunological properties of A60 of Mycobacterium bovis BCG were previously described (Cocito and Vanlinden 1986). The present study focused on the intracellular distribution of this antigen. Fractionation of mycobacterial homogenates by ultracentrifugation indicated that most of A60 was present within the cytoplasm. Some of the antigen was located within the cell wall, from which it was released by extraction with alkali. Submission of cytoplasm to high speed centrifugation caused A60 to cosediment with ribosomes; however, dissociation of ribosomes in low-Mg buffer did not alter the sedimentation pattern of A60. Labelled A60, after ultracentrifugation in sucrose density gradients without Mg2+, was distributed throughout the entire gradient: treatment of (125I)A60 with urea or detergents produced a peak of radioactivity located in the upper part of the gradient. It is concluded that A60 is represented by a heterogeneous family of molecules of increasing sizes: polymerization being enhanced by Mg2+ and reversibly prevented by urea. Some or all of the biological properties hitherto attributed to ribosomal particles may, in fact, be due to their contamination with cosedimented A60.