Differential T-cell recognition of native and recombinant Mycobacterium tuberculosis GroES

Mycobacterium tuberculosis GroES was purified from culture filtrate, and it s identity was confirmed by immunoblot analysis and N-terminal sequencing. Comparing the immunological recognition of native and recombinant GroES, we found that,whereas native GroES elicited a strong proliferative response a nd release of gamma interferon-gamma by peripheral blood mononuclear cells from healthy tuberculin reactors, the recombinant protein failed to do so. The same difference in immunological recognition was observed in a mouse mo del of TB infection. Both the native and recombinant preparations mere reco gnized by mice immunized with the recombinant protein. Biochemical characte rization including sodium dodecyl sulfate-polyacrylamide gel electrophoresi s, two-dimensional electrophoresis, and mass spectrometry analysis of both proteins demonstrated no differences between the native and recombinant for ms of GroES except for the eight additional N-terminal amino acids derived from the fusion partner in recombinant GroES, The recombinant fusion protei n, still tagged with the maltose binding protein,was recognized by T cells isolated from TB-infected mice if mixed with culture filtrate before affini ty purification on an amylose column. The maltose binding protein treated i n the same manner as a control preparation was not recognized. Based on the data presented, we suggest that the association of biologically active mol ecules from culture filtrate with the chaperone GroES may be responsible fo r the observed T-cell recognition of the native preparation.