Characterization of native and denatured ricin using MALDI-TOF/MS


P. Sehgal, M. K. Rao, O. Kumar, R. Vijayaraghavan

Abstract


Ricin is a toxic protein present in the seeds of castor bean plant. It can be inactivated by heat; therefore characterization of denatured ricin is essential to differentiate it from native ricin and to avoid any ambiguity in its identification. In this study, potential of mass spectrometry using MALDI-TOF/MS has been exploited to investigate the effects of heat treatment on ricin and spiked food matrices. The molecular weights of ricin, ricin A (A1 and A2) and B chain were found to be 62.8 kDa, 31.2 kDa, 32.5 kDa and 32 kDa respectively. The mass spectrum revealed a polypeptide chain of 11.1 kDa for denatured ricin. The peptide mass fingerprinting showed 24 peptides, six were common both in native and denatured ricin. The differentiating peptide at position 294-318 (m/z 934.533) was observed only in denatured ricin. The three selected marker peptides m/z 1013.6, 1310.7, 1728.9 are chosen for identification of ricin inactivated by heat in spiked apple juice and milk samples by immunocapture analysis. There is always a probability of denatured non- toxic ricin being confused with native (toxic) ricin to create unnecessary panic. Keeping this probability in mind, our study will be of immense value in minimising such risk.

Keywords


Ricin, denatured, ambiguity, MALDI-TOF/MS, peptide mass fingerprinting, marker, immunocapture.

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