Isolation And Purification Of BmK AS-like Peptides From The Venom Of Scorpion Buthus Martensi Karsch And Effect Of BmK AS'on[Ca～(2+)]_i In Cultured Rat Skeletal-muscle Cell

Part one :Isolation and purification of BmK AS-like peptides from the venom of scorpion Buthus martensi Karsch In the first section of this work, a new isolation and purification procedure of BmK as-like peptides from the venom of scorpion Buthus rnartensi Karsch was described. In this improved procedure, three different methods of routine column chromatography (Sephadex G-25 Sephadex G-50 DEAE Sephadex A-25) and repeated high performance liquid chromatography were used together. The results showed: Two BmK AS?like peptides(fraction 1-2-4-1 and fraction 1-2-4-3) were found. Their molecular weights were determined to be all about 7500 Da by SDS-polyacrylamide gel slab electrophoresis. Part two: Effect of Bmk AS? an active polypeptide from scorpion Buthus martensi Karsch, on [Ca2+]+ in cultured rat skeletal-muscle cell In the second section, we studied the effect of BmK AS on intracellular free calcium concentration([Ca2+]1) in cultured rat skeletal-muscle cell. The cells were incubated with fluo- 3/AM.Fluoresence was measured under the laser scanning confocal microscope. The change of [Ca2+]+ was represented by fluorescent intensity. The results showed: 1) In the absence of extracellular Ca>, BmK AS 1 .30x109+ 1.3Ox10 +. 1 .30x10 +. 0.65x10 enhanced fluorescence obviously. 2) In the presence of extracellular Ca>, BmK AS. 1.30x10 +. 1.30xl0 + 0.65x10 enhanced fluorescence similarly. 3) In the presence of extracellular Ca>, following the concentration of BmK AS increased, fluorescence increased too. 4) when the concentration ofBmK AS were 1.30x1O + 1.30x10 0.65x10 here were no obvious difference of fluorescence in the presence and absence of extracellular Ca>. 5) BmK AS 1.30xl09M enhanced fluorescence more obviously in the presence of extracellular Ca> than that in the absence of extracellular Ca>. 6) The time of calcium-peak under BmK AS (950s)was longer than that under kcl(30s).From results we can draw a conclusion :BmK AS 1.30x10 ) increased [Ca>]1 through the intracellular mechanism . BmK AS 1.3x109) increased [Ca2+]1 through not only intracellular mechanism but also extracellular mechanism.