<span style="font-size:12.0pt;line-height:115%; font-family:"Times New Roman";mso-fareast-font-family:"Times New Roman"; mso-ansi-language:EN-IN;mso-fareast-language:EN-IN;mso-bidi-language:HI">Affinity purification and characterization of a seed lectin from <i>Crotalaria medicaginea</i></span>

Abstract

Purification of lectin from the seeds of <i>Crotalaria medicaginea </i>Lamk by affinity chromatography on asialofetuinlinked amino activated silica, yielded a single band on non-denatured PAGE at <i>p</i>H 4.5 and 8.3 and, a single peak on HPLC size exclusion and cation exchange columns. The molecular mass of the native C. <i>medicaginea </i>lectin was determined to be 125 kDa by gel filtration. In SDS-PAGE, the lectin migrated as a single band of M_r 31.6 kDa under reducing and nonreducing conditions, indicating that it is a tetramer of apparently identical subunits. It agglutinated red blood cells (RBCs) from rabbit and human ABO blood groups. It also reacted with RBCs from rat, sheep, goat and guinea pig but after desialylation with neuraminidase. The hemagglutination activity of the lectin was inhibited by D-galactose and its derivatives. Amino acid analysis showed that lectin was rich in aspartic and glutamic acid and, did not contain sulphur containing amino acids. The lectin is a glycoprotein having 1.41% of neutral sugars. It is labile at temperature above 60°C. It needs divalent cations for its activity, as a loss of activity was observed on removal of Ca²⁺ and Mn²⁺. Denaturing agents like urea, thiourea and guanidine-HCl have no effect on its activity.