[Purification of recombinant proteins with an example of tumor necrosis factor thymosin-alpha1].

Prikladnaia biokhimiia i mikrobiologiia2010PMID: 20391771

Plain Language Summary

Russian researchers developed an efficient method to purify a hybrid protein combining tumor necrosis factor with thymosin alpha-1 from bacterial cultures. The purification process achieved 80% recovery of the protein from bacterial biomass while maintaining its cancer cell-killing activity.

Abstract

Hybrid protein, cancer necrosis factor thymosin-alpha1 (CNF-T), when synthesizing in strain-producer of Escherichia coli SG200-50 with plasmid pThy315, was a part of "inclusion bodies" mostly in the form of a high-molecular complex with other proteins due to the S-S bonds formation. An approach of purification of CNF-T has been proposed, which is based on the destruction of the complex in the presence of sodium dodecylsulfate (DDS-NA) and dithiotreitol (DDT) followed by gel-filtration on Sephadex G-100 and renaturation by ultrafiltration on hollow fibers. The method allows the isolation of electrophoretically homogeneous CNF-T containing no DDS-Na and having high cytotoxic activity against cancer cells of mouse adenocarcinome L-929. The yield of CNF-T achieved 80% relative its content in biomass and 30% relative the total protein.

Authors

Fedorov, T V; Korobov, V I; Nazarov, V G; Smolkina, A E; Shmelev, V A