Single-Step Purification and Characterization of A Recombinant Serine Proteinase Inhibitor from Transgenic Plants

Jha, Shweta; Agarwal, Saurabh; Sanyal, Indraneel; Amla, D.
May 2016
Applied Biochemistry & Biotechnology;May2016, Vol. 179 Issue 2, p220
Academic Journal
Expression of recombinant therapeutic proteins in transgenic plants has a tremendous impact on safe and economical production of biomolecules for biopharmaceutical industry. The major limitation in their production is downstream processing of recombinant protein to obtain higher yield and purity of the final product. In this study, a simple and rapid process has been developed for purification of therapeutic recombinant α-proteinase inhibitor (rα-PI) from transgenic tomato plants, which is an abundant serine protease inhibitor in human serum and chiefly inhibits the activity of neutrophil elastase in lungs. We have expressed rα-PI with modified synthetic gene in transgenic tomato plants at a very high level (≃3.2 % of total soluble protein). The heterologous protein was extracted with (NH)SO precipitation, followed by chromatographic separation on different matrices. However, only immunoaffinity chromatography resulted into homogenous preparation of rα-PI with 54 % recovery. The plant-purified rα-PI showed molecular mass and structural conformation comparable to native serum α-PI, as shown by mass spectrometry and optical spectroscopy. The results of elastase inhibition assay revealed biological activity of the purified rα-PI protein. This work demonstrates a simple and efficient one-step purification of rα-PI from transgenic plants, which is an essential prerequisite for further therapeutic development.


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