Isolation, characterization and production of a new recombinant lectin protein from leguminous plants

Lectins are present in microorganisms, plants and animals and have attracted great interest due to their varied physiological roles in cell agglutination, anti-tumour, immunomodulatory, antifungal and antiviral effects. Legume lectins are important to the pharmaceuticals but they are produced in low amounts in the plant seeds. Moreover, the genes controlling these proteins are conditionally active, i.e., they work under specific circumstances and not in regular manner. Looking into these limitations, we aimed this work to produce a recombinant lectin for pharmaceutical use especially for the treatment of cancer. Three different legume plant species were collected from Aseer region of Kingdom of Saudi Arabia viz., Acacia seyal, Pisum sativum (wild type) and Pisum sativum (Pea). The plant tissues were subjected to RNA extraction, the extracted RNA was used for lectin gene amplification using specific primers. Cloning, subcloning of the Acacia 400bp gene was carried out and in vitro transcription, combined with protein purification was undertaken. The cytotoxicity of the recombinant lectin was performed on two cell lines such as breast cancer MCF-7 and liver cancer HepG-2 cells. Our study resulted in the observation of two amplicones with all the three examined species, the amplicone molecular sizes were 800 and 400bp. The 400bp amplicone was excised from the agarose gel, purified and sequenced. The sequence analysis revealed that the nucleotide sequence belongs to lectin gene. The sequence analysis revealed that the lectin gene isolated from Pisum sativum (wild plant) is similar to the Pisum sativum lect1 with identity 90%, whereas, lectin isolated from Pisum (pea) showed similarity of 91% with the other lectins. On the other hand, Acacia lectin showed similarity with Lotus japonicus nod factor binding lectin gene with identity of 95%. Thus we conclude that new lectin protein of 17 and 15 kDa was produced that can be used by pharmaceutical industries.