The Journal of Medical Microbiology, Vol 41, Issue 3 173-178, Copyright © 1994 by Society for General Microbiology
Purification and partial characterisation of a novel 66-kDa seroreactive protein of Mycobacterium tuberculosis H37Rv
R. G. Deshpande, M. B. Khan, D. A. Bhat and R. G. Navalkar
Department of Microbiology and Immunology, Morehouse School of Medicine, Atlanta, Georgia 30310.
A seroreactive protein (TB66) was purified from culture filtrate (CF) and
cell sonicate (CS) of Mycobacterium tuberculosis H37Rv by immobilised metal
affinity chromatography (IMAC) on a Ni-nitrilotriacetic acid (NTA) column.
The TB66 preparations obtained by IMAC contained predominantly a 66-kDa
protein with a pI of c. 5.5 as determined by two-dimensional
electrophoresis. TB66 was detected in the CF as early as 1 week of growth
of H37Rv. The NH2-terminal amino-acid sequence showed 85% homology with the
N-terminal sequence of bovine serum albumin (BSA) and 80% homology with
human serum albumin. Amino-acid analysis indicated a difference in the
amino-acid content of TB66 when compared to BSA, with an abundance of
acidic amino acids. A monoclonal antibody (MAb) OD4AG3, raised in this
laboratory against an M. avium complex (MAC 101) sonicate cross-reacting
with H37Rv sonicate, recognised a heat-stable and trypsin-sensitive epitope
of this protein. TB66 was also recognised by MAbs IT1 and IT20 which also
react with the 14-kDa antigen of the M. tuberculosis complex. Antibodies
against TB66 were present in the sera of 62 of 64 patients with
tuberculosis; sera from normal healthy individuals showed no significant
reactivity. TB66 appears to be a predominant secretory protein of M.
tuberculosis and could play an important role in the pathogenesis of this
organism.
