Isolation of fully active and stable corn coleoptile lectins.
It appears that the reason for the lack of information and data about corn coleoptile lectin is due to the instability of preparation with a rapid loss of hemagglutinating activity and abundant precipitate. In this paper, we assayed phosphate, borate, tris, and ascorbate-sucrose extraction buffers to compare lectin activity and protein yield. The ascorbate-sucrose buffer (AS-buffer) proved to be the best extracting solution. In a second step, cold acetone was employed to concentrate crude lectin. An increase of specific activity from the first to the third acetone precipitation was obtained. The protective effect on hemagglutinating activity of AS-buffer led us to test cysteine, metabisulfite, borohydride, and dithiothreitol (DTT) as reducing agents. The compounds were ineffective. Dissociating gel electrophoresis of the acetone-purified lectin disclosed a band pattern of components around 60 kD, a second band at 29 kD, and minor bands close to 15 KD. The procedure is useful for the preparation of stable, high activity corn coleoptile lectin. Further purification using affinity chromatography, as in reference (1) could become a major advance to obtain corn lectin of adequate purity for sequential analysis.
Biochemistry and Immunology Unit, Technological Institute of Oaxaca, Mexico.
M
M
Martínez
F
F
Córdoba
D000818:Animals; D001205:Ascorbic Acid / Q000737:chemistry; D002021:Buffers; D011232:Chemical Precipitation; D018548:Cotyledon / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D006384:Hemagglutination; D006801:Humans; D037102:Lectins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D037121:Plant Lectins; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism; D019163:Reducing Agents / Q000494:pharmacology; D013395:Sucrose / Q000737:chemistry; D003313:Zea mays / Q000737:chemistry*
D002021:Buffers; D037102:Lectins; D037121:Plant Lectins; D010940:Plant Proteins; D019163:Reducing Agents; D013395:Sucrose; D001205:Ascorbic Acid