兔骨骼肌肌球蛋白的纯化及溶液浊度和溶解度研究

Extraction and Purification of Myosin from Rabbit Skeletal Muscles and Its Solution Turbidity and Solublility

从雄兔的腰大肌(Pasoasmajor,简称PM)和半膜肌(Semimembranosusproprius,简称SMp)中提取纯化了肌球蛋白,并用双缩脲法测定其浓度,采用Laemli的不连续SDS-PAGE系统检验其纯度。结果表明,提取纯化的肌球蛋白浓度为20～30mg/ml,提取率为1%左右,在SDS-PAGE图谱上出现四条带,一条为肌球蛋白重链,其分子量为205kDa和三条轻链,分子量分别为25.9、17.8和13.8kDa。研究了肌球蛋白质量浓度、pH值、离子强度、温度对PM和SMp肌球蛋白溶液的浊度和溶解度的影响。结果表明,肌球蛋白浓度与浊度呈正线性相关(R2=0.97)。PM和SMp肌球蛋白在离子强度为0.6的KCl溶液中pH值为5.5时浊度最大,溶解度在pH值大于5.0时显著上升。离子强度下降,肌球蛋白溶液的浊度会上升,而溶解度下降。相同浓度下温度升高肌球蛋白溶液浊度上升,PM肌球蛋白在55℃,SMp肌球蛋白在60℃其浊度达到最大值。而温度升高溶解度下降。

The myosin was extracted and purified from Pasoas major (PM) and Semimembranosus proprius (SMp) of male rabbit, and effects of myosin concentration, pH, ionic strength, and heating temperature on the turbidity and solubility of PM and SMp myosin were studied. The results showed that concentration of the purified myosin was about 20～30mg/ml, the rate of extraction was about 1%. Four bands were observed in the purified myosin SDS-PAGE pattern, which were assigned to one myosin heavy chains (MHC) and three myosin light chains,with molecular weight of 205, 25.9, 17.8 and 13.8kDa, respectively. The results showed that the effect of myosin concentration on turbidity was linear. The turbidity reached its maximum at pH5. 5 and in 0.6mol/L KCl. The solubility increased significantly when pH value was greater than pH5.0. The turbidity increased and the solubility decreased with the decrease of the ionic strength. The turbidity increased with the increase of temperature under the same myosin concentration. PM and SMp myosin turbidity reached the maximum at 55℃ and 60℃ respectively. The solubility decreased when the temperature increased.