脂肪族类两亲物对肌浆网膜蛋白结构的影响

Effects of Fatty Amphiphiles on Protein Structure of Sarcoplasmic Reticulum

本文观察和比较了六种C_(18)脂肪族类两亲物(fatty amphiphile,FA),包括硬脂酸(stearic acid)、硬脂胺(stearyl amine)、硬脂醇(stearyl alcohol)、油酸(oleic acid)、油胺(oleylamine)和油醇(oleyl alcohol),对肌浆网(sarcoplasmic reticulum,SR)钙泵蛋白结构的影响。当FA∶SR(μmol∶mg)的比例为2.67∶1—21.33∶1时,除油醇(oleyl alcohol)外,其余五种FA引起天然兔骨骼肌肌浆网蛋白内源荧光强度降低。随FA∶SR比例升高,降低幅度加大。五种FA的最大降低幅度在10—32％之间。其中带电荷FA比不带电荷FA的作用强,但均未见峰位位移。当FA∶SR的比例为2.67∶1时,除硬脂醇(stearyl alcohol)外,其余五种FA使N-(3-芘)-马来酰胺(N-(3-pyrene)maleimide,N-(3-p)-M)修饰的SR蛋白巯基荧光强度分别上升9％,40％,150％,193％和5％,但也未见峰位位移。ATP可减弱胺类FA引起的SR蛋白内源荧光降低和巯基荧光升高的幅度。油酸、油胺和硬脂胺既抑制SRCa^(2+),Mg^(2+)-ATPase活力和SR钙蓄积,又使SR钙泵蛋白的巯基修饰荧光显著上升。提示C_(18)脂肪族类两亲物使SR功能受损与其引起钙泵蛋白构象的显著变化有关。

Effects of six kinds of C_(18) saturated and cis-monounsaturated fatty amphiphiles (stearic acid, stearylamine, stearyl alcohol, oleic acid, oleylamine and oleylalcohol) on the structure of calcium pump protein in rabbit skeletal muscle sarcoplasmic reticulum (SR) were studied. When the amphiphile:SR ratios (μmol:mg) was from 2.67:1 to 21.33:1, with an increase in the ratios, the intensities of intrinsic protein fluorescence of SR were reduced by the amphiphiles except oleyl alcohol, and the maximal decreases in emission intensity induced, by the five amphiphiles were measured by 10—30%, but no shift of emission maximum was monitored, Following addition of the above five kinds of fatty amphiphiles except stearyl alcohol at an amphiphile:SR ratio of 2.07:1,the SH-modified protein fluorescences of SR labelled with N-3-pyrene-maleimide were enhanced by 9% ,40%, 150%, 193% and 5%, respectively, and no shift of emission maximum was also monitored. Adding of MgATP was found to decrease the effects of saturated and unsaturated fatty amines on both the emission spectra. Oleic acid, oleylamine and stearylamine among the amphiphiles inhibited both calcium sequestration and Ca^(2+),Mg^(2+)-ATPase activity as well as strongly increased the SH-modified fluorescence intensity of SR calcium pump protein. These results indicated that inhibition of the function of calcium pump protein by amphiphiles may be involved in the remarkable conformational changes.