Mushroom lectins have important biological and biomedical applications. Most lectins purified from these organisms exhibit high toxicity in animal cells and toward microbial agents. They are able to induce cell growth inhibition and metabolism by their ability to interact with glyconjugate components (glycoproteins receptors, glycolipids) present in their membrane. After lectins bind to these membrane receptors, they induce cellular signalization chains in which gene expression is regulated and cell death programming (apoptosis) is activated. In this work, a new multimeric lectin was characterized from the rare saprobic edible mushroom, Laetiporus sulphureus strain TMES43, grown in the Algerian forest. Lectin was isolated with ammonium sulfate precipitation followed by affinity chromatography on a Sepharose 4B column, with specific activity of 1204.7 units of hemagglutination activity/mg and 35.55% yield. The protein has a tetrameric structure with a molecular weight of 36 kDa for each subunit, with a total molecular weight of approximately 140 kDa. In addition, a Mascot peptide fingerprint study on a matrix-assisted laser desorption ionization-time of flight tandem fragment showed identity with autophagy-related protein 16 from Meyerozyma guilliermondii (strain ATCC 6260/CBS 566/DSM 6381/JCM 1539/NBRC 10279/NRRL Y-324; Expasy ID: ATG16_PICGU) and no sequence similarity to known mushroom lectins. L. sulphureus hemagglutination activity was reduced by 5 mM of lactose and 10 mM of EDTA incubation and was recovered by metallic cations such as CaCl2, MgCl2, and ZnCl2. L. sulphureus purified lectin had no human ABO group specificity and showed low temperature and alkaline pH stabilities. The MTT preliminary assay showed that L. sulphureus purified lectin induced high cytotoxicity for tumor cells and normal cells.