STUDIES OF ISOLATION, PURIFICATION AND IODINATION OF ?-BUNGAROTOXIN / 第二军医大学学报

ABSTRACT

The isolation purification, iodination and identification of a-Bungarotoxin(a-BTX)were recorded.Purified a-BTX was prepared from the venom of Hunan Bu-ngarus multicinctus (Blyth) by CM-Sephadex C~50 ionexchange chromatography and Sephadex G~25 chromatography.The homogeneous a-BTX was obtained by rechromatography of purified ct-BTX (fraction 3) on a CM-Cellulose CM-32, and was labelled with 1251 by chloramine-T method.The 1251-?-BTX was then separated from Na125I and purified by Sephadex G-25 column.The results yield a specific activity of 90~100 Ci/mM.40~50% labelling.40~60% recovery of protein and less than 3 % contamination with free iodine in 125I-?-BTX.The purified a-BTX was identified by determining its N-terminal amino acid, amino acid composition, isoelectric and polyacrylamide gel electrophoresis.Biological activity of the ?-BTX was determined by its toxicity to mice, its neuromuscular blocking action and binding to acetylcholine receptors at the motor entplate of the rat diaphragm.