Crystalline Kidney-Bean Leucoagglutinin

1. Kidney-bean (Phaseolus vulgaris) leucoagglutinin was purified on a preparative scale by fractional precipitation with ethanol followed by ion-exchange chromatography on DEAE-cellulose and SP-Sephadex and exclusion chromatography on Sephadex G-150. The final purification step consisted of crystallization in aqueous solution. 2. The crystallized leucoagglutinin was homogeneous by analytical ultracentrifugation, disc electrophoresis, isoelectric focusing and immuno-diffusion. 3. Leucoagglutination was demonstrable at 1–3 μg/ml and optimum lymphocyte stimulation occurred at 1–5 μg/ml. No erythroagglutinating activity could be detected. 4. The leucoagglutinin lacks sulphur-containing amino acids completely, but contains the other common amino acids, aspartic acid and serine being predominant. It contains glucosamine and mannose as the only carbohydrates. In addition it contains considerable amounts of calcium and manganese, which are essential for the lymphocyte-stimulating and leucoagglutinating activities. 5. The leucoagglutinin has the following physico-chemical properties: Mr= 126000, s020,W= 6.87 S, f/f0= 1.45, partial specific volume = 0.728 ml/g, pI= 5.1, A1%280 nm/cm= 11.4. The leucoagglutinin is composed of 4 identical subunits, Mr= 31000.