Immunoproteomic analysis of mycobacterium leprae derived cell membrane antigen

Abstract

The aim of the study was serological characterization of antigens of cell membrane proteins (MLMA) of armadillo derived Mycobacterium leprae, a causative organism of leprosy. Cell membrane proteins of Mycobacterium leprae were separated using two dimensional gel electrophoresis (2-DGE) and then immunoblotted with sera from leprosy patients, tuberculosis patients, healthy individuals and anti-human IgG/IgA/IgM antibodies. The immunoblots thus obtained were compared and proteins corresponding to spots detected only in leprosy immunblots were finally analyzed for their characterization by MALDI-TOF/TOF. Employing this approach, 14 (6 with anti-IgA, 5 with anti-IgM and 3 with IgG) immuno-reactive proteins with anti-human IgA/IgM/IgG were recorded. Out of these 14 proteins only 8 proteins (encoded as: MAL 1, MAL 4, MAL 5, MAL 6, MML 5, MML 6, MML 7 and MML 8) were identified to be M. leprae specific by MALDI-TOF/TOF and MASCOT search. Of these 8 proteins, 1 protein was identified as bacterioferritin (MMP-II) and remaining 7 proteins appeared to be as 5 isoforms of major membrane protein-1 (MMP-1), a 35kDa protein. To our knowledge this is the first report regarding existence of isoforms of MMP-1/35kDa protein. On preliminary examination for serological potential of these antigens, MAL5 was found to be the best (sensitivity= 82.6 specificity = 54.6%, efficiency= 68.9%) among IgA reactive proteins. On the other hand, antigen encoded as MML7 was best (sensitivity 90.9% with a specificity of 33.3%, efficiency of 70.6) among IgM reactive proteins. Though seroreactive, as such both of these antigens do not seem to be very promising serodiagnostic reagents due to their low specificities, information obtained out of this study has opened paths towards dissection of these proteins at the peptide level with the view to look for their efficient serological potential.