Physicochemical properties and oxidative inactivation of soluble lectin from water buffalo (Bubalus bubalis) brain.
Lectins are carbohydrate-binding proteins present in a wide variety of plants and animals, which serve various important physiological functions. A soluble beta-galactoside binding lectin has been isolated and purified to homogeneity from buffalo brain using ammonium sulphate precipitation (40-70%) and gel permeation chromatography on Sephadex G50-80 column. The molecular weight of buffalo brain lectin (BBL) as determined by SDS-PAGE under reducing and non-reducing conditions was 14.2 kDa, however, with gel filtration it was 28.5 kDa, revealing the dimeric form of protein. The neutral sugar content of the soluble lectin was estimated to be 3.3%. The BBL showed highest affinity for lactose and other sugar moieties in glycosidic form, suggesting it to be a beta-galactoside binding lectin. The association constant for lactose binding as evidenced by Scatchard analysis was 6.6 x 10(3) M(-1) showing two carbohydrate binding sites per lectin molecule. A total inhibition of lectin activity was observed by denaturants like guanidine HCl, thiourea and urea at 6 M concentration. The treatment of BBL with oxidizing agent destroyed its agglutination activity, abolished its fluorescence, and shifted its UV absorption maxima from 282 to 250 nm. The effect of H2O2 was greatly prevented by lactose indicating that BBL is more stable in the presence of its specific ligand. The purified lectin was investigated for its brain cell aggregation properties by testing its ability to agglutinate cells isolated from buffalo and goat brains. Rate of aggregation of buffalo brain cells by purified protein was more than the goat brain cells. The data from above study suggests that the isolated lectin may belong to the galectin-1 family but is glycosylated unlike those purified till date.
Rizvi
Sabika
S
Department of Biochemistry, Faculty of Life Sciences, A.M. University, Aligarh, UP 202002, India.
Banu
Naheed
N
D000645:Ammonium Sulfate; D000818:Animals; D001923:Brain Chemistry / Q000502:physiology*; D002020:Buffaloes / Q000378:metabolism*; D002241:Carbohydrates / Q000737:chemistry; D002449:Cell Aggregation / Q000187:drug effects; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002850:Chromatography, Gel; D003956:Dialysis; D004591:Electrophoresis, Polyacrylamide Gel; D006384:Hemagglutination / Q000187:drug effects; D006861:Hydrogen Peroxide / Q000737:chemistry; D037102:Lectins / Q000737:chemistry* / Q000302:isolation & purification; D008670:Metals / Q000737:chemistry; D008970:Molecular Weight; D010084:Oxidation-Reduction; D011489:Protein Denaturation; D019163:Reducing Agents / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
D002241:Carbohydrates; D037102:Lectins; D008670:Metals; D019163:Reducing Agents; D006861:Hydrogen Peroxide; D000645:Ammonium Sulfate