Protein & Peptide Letters, 2012, 19, 000-000 1 0929-8665/12 $58.00+.00 © 2012 Bentham Science Publishers Purification, Characterizations of a Snake Guard Seed Lectin with Anti- tumor Activity Against Ehrlich Ascites Carcinoma Cells In Vivo in Mice Syed Rashel Kabir 1 *, Farhadul Islam 1 , Jahangir Alom 1 , Abu Zubair 2 and Nurul Absar 1 1 Department of Biochemistry and Molecular Biology, Rajshahi University, Rajshahi-6205, Bangladesh; 2 Department of Food Technology and Nutritional Science, Mawlana Bhashani Science and Technology University, San- tosh, Tangail-1902, Bangladesh Abstract: A lectin was purified (designated as TCSL) from the Snake guard seeds with molecular mass of 56±2 kDa con- taining two subunits (34±1 and 22±1 kDa.). TCSL exhibited high agglutination activity at the temperature range 30 to 70ºC and did not lose its activity between pH 3.0 to 12.0. The lectin was stable in the presence of denaturants and aggluti- nated mouse, goat, cow, chicken and human erythrocytes. TCSL did not show antifungal activity whereas it agglutinated six pathogenic bacteria and the lectin showed less toxicity against brine shrimp nauplii with the LC 50 of 261±29 μg/ml. TCSL showed 28% and 72% inhibition against Ehrlich ascites carcinoma (EAC) cells in vivo in mice when administered 1 mg/kg/day and 2 mg/kg/day (i.p.) respectively for five days. TCSL enhanced the number of macrophages remarkably in the normal mice. The lectin reduced the tumor burden to 62% of EAC cells and significantly increased the hemoglobin and RBC. Treating the EAC bearing mice with TCSL at 2 mg/Kg/day for ten days with a monitoring of 20 days decreased the total WBC towards the normal level and it increased the life span by 39%. Keywords: Antitumor, bacterial agglutination, blood group, life span, lectin, toxicity. INTRODUCTION Lectins are the heterogeneous group of proteins of non- immune origins that bind reversibly to mono and oligosac- charides with high specificity but without any catalytic activ- ity [1]. According to the molecular weight and evolutionary relationships, plant lectins can be subdivided into seven dif- ferent families which include the legume lectins, type-2 ribo- some-inactivating proteins, monocot mannose binding lect- ins, jacalin-like lectins and chitin-binding lectins [2]. In the recent years, lectins have been proved to be attractive for research works due to their role in cell agglutination, toxic- ity, antifungal, antibacterial, antiviral, anti-proliferative and antitumor effects [3-5]. Legume seed lectin has been studied most widely due to its higher presence in the seed. Trichosanthes cucumerina (Snake guard) belongs to the Leguminosae group that grows both in Bangladesh and India, and the seed is used as a popu- lar food in both the countries. A lectin with Mr. 62 kDa was purified from Trichosanthes cucumerina seed (designated as TCSL) cultivated in India. The lectin contained 3% neutral sugar and two subunits with Mr. of 41 and 22 kDa linked by disulfide bridge(s) and the involvement of histidine residues to its carbohydrate-binding activities were identified by chemical modification [6, 7]. Steady state and time-resolved fluorescence studies show that the tryptophan residues of the lectin were buried in the hydrophobic interior of the protein matrix [8]. Galactose, specifically p-nitrophenyl-- *Address correspondence to this author at the Department of Biochemistry and Molecular Biology, Faculty of Science, Rajshahi University, Rajshahi- 6205, Bangladesh; Tel: +880-721-750041/4109 (Office); E-mail: rashelkabir@ru.ac.bd; rashelkabir@gmail.com galactopyranoside was the best inhibitor among a variety of sugars [7]. TCSL exhibited immunological cross-reactivity with Cucurbitaceae seed lectin SGSL [6]. It bound to a vari- ety of porphyrins with considerable avidity and the thermo- dynamics and kinetics of porphyrin binding by this lectin had been investigated [7]. The pH and temperature optima, isola- tion profile and secondary structure content were also stud- ied [9]. But till now no detailed information about the chemical stability and biological properties such as toxicity, blood group specificity, antibacterial, antifungal and antitu- mor activities of the lectin were known. In order to check these properties, we have purified the lectin from Trichosan- thes cucumerina seed cultivated in Bangladesh and the mo- lecular mass of the newly purified lectin did not match with the reported one. So, it was also necessary to reinvestigate some characteristic features (sugar specificity, heat and pH stability) of the purified lectin. In this study we have focused to the differences of the characteristic attributes and report- ing first time the blood group specificity, bacterial agglutina- tion activity, antifungal activity, toxicity and antitumor activ- ity of snake guard seed lectin against EAC cells in vivo in mice. MATERIALS AND METHODS Materials Diethyl aminoethyl (DEAE)-cellulose and Sepharose-4B was procured from Wako (Japan) and Fluka (Sweden) re- spectively. All other chemicals/reagents were of the highest grades available commercially. The seeds of Trichosanthes cucumerina were collected from the local market.