Abstract: The study was aimed to produce recombinant ricin A chain (r-RTA) with high biological activity and compare its toxicity with native ricin (n-RT). We synthesized the published RTA gene sequence on NCBI and cloned it into the pET-28a vector, and then prokaryoticly expressed by the E. coli. After purification with Ni²⁺-NTA resin column, the r-RTA was eluted by 500 mmol/L of imidazole solutions. The purified protein was identified by SDS-PAGE and Western blotting, and the immunogenicity was determined by ELISA. Animal experiments and cell toxicity analysis were conducted to compare the toxicity between the r-RTA and n-RT. The rate of recombinant expression was 31.2%. About 20 mg fusion proteins were obtained from 1 000 mL cultures with the protein purity of ≥90%. The results of ELISA showed that the immunogenicity of r-RTA were about 1.27 times of that of n-RT. The half inhibitory concentration (IC₅₀) of n-RT to RAW264.7 cells was 0.01 μg/mL and the median lethal dose (LD₅₀) to mouse was (3.27±0.44) μg/kg. We used the same dose of toxin to challenge the Raw264.7 cells or mouse, and found that the virulence of n-RT was 2 700 times of that of the r-RTA. The mortality rate of n-RT was 40%, while r-RTA was all survive. Our results suggested that single RTA, without the assistance of RTB, had a significantly reduced toxicity, which provided vital data and theoretical supports for the development of RTA-based vaccine.

Key words: ricin toxin A chain (RTA); prokaryotic expression; natural ricin toxins (n-RT); comparison of toxicity