Effects of melittin on lipid-protein interactions in sarcoplasmic reticulum membranes.
Effects of melittin on lipid-protein interactions in sarcoplasmic reticulum membranes.
复制标题
蜂毒肽对肌浆网膜脂质-蛋白质相互作用的影响。
DOI：
10.1016/s0006-3495(92)81736-8
复制
发表时间：
1992
影响因子：
3.4
通讯作者：
Thomas,DD
中科院分区：
文献类型：
--
作者：
Mahaney,JE;Kleinschmidt,J;Marsh,D;Thomas,DD
为了研究蜂毒肽抑制肌浆网 (SR) 膜中钙腺苷三磷酸酶 (ATPase) 活性的物理机制，我们使用电子顺磁共振波谱来探讨蜂毒肽对 SR 中脂质-蛋白质相互作用的影响。先前的研究表明，蜂毒肽显着限制了 Ca-ATP 酶的旋转流动性，但仅略微降低了 SR 中的平均脂质烃链流动性。因此，在本研究中，我们询问蜂毒肽是否对 Ca-ATPase 边界脂质（即围绕蛋白质的运动限制脂质的环形壳）具有优先作用。硬脂酸和磷脂酰胆碱的顺磁性衍生物（在 C-14 处进行自旋标记）被掺入 SR 膜中。这些探针的电子顺磁共振光谱包含两个成分，对应于运动受限和运动流体脂质，通过光谱减法对其进行分析。添加越来越多的蜂毒肽，达到 10 mol 蜂毒肽/mol Ca-ATP酶的水平，逐渐增加了限制性脂质的分数，并增加了复合光谱中两种成分的超精细分裂，表明蜂毒肽降低了流体和限制性脂质群体的烃链旋转迁移率。当浓度高于 10 mol 蜂毒肽/mol Ca-ATP酶时，没有观察到进一步的影响。在对照样品和含有蜂毒肽的样品的光谱中，限制性脂质的比例随着温度的升高而显着降低。蜂毒肽的效果与降低温度的效果类似，即，在蜂毒肽 (10:1) 存在下获得的每个光谱与在温度降低约 5°C 时没有蜂毒肽获得的光谱几乎相同。结果表明，蜂毒肽对 SR 膜的主要作用是诱导蛋白质聚集，而蜂毒肽与脂质的直接结合反过来会增强这种聚集，从而导致观察到的脂质流动性降低。蛋白质聚集被认为是蜂毒肽使 Ca-ATP 酶失活的主要原因，也可能通过边界层脂质流动性的降低进行调节。
To investigate the physical mechanism by which melittin inhibits Ca-adenosine triphosphatase (ATPase) activity in sarcoplasmic reticulum (SR) membranes, we have used electron paramagnetic resonance spectroscopy to probe the effect of melittin on lipid-protein interactions in SR. Previous studies have shown that melittin substantially restricts the rotational mobility of the Ca-ATPase but only slightly decreases the average lipid hydrocarbon chain fluidity in SR. Therefore, in the present study, we ask whether melittin has a preferential effect on Ca-ATPase boundary lipids, i.e., the annular shell of motionally restricted lipid that surrounds the protein. Paramagnetic derivatives of stearic acid and phosphatidylcholine, spin-labeled at C-14, were incorporated into SR membranes. The electronic paramagnetic resonance spectra of these probes contained two components, corresponding to motionally restricted and motionally fluid lipids, that were analyzed by spectral subtraction. The addition of increasing amounts of melittin, to the level of 10 mol melittin/mol Ca-ATPase, progressively increased the fraction of restricted lipids and increased the hyperfine splitting of both components in the composite spectra, indicating that melittin decreases the hydrocarbon chain rotational mobility for both the fluid and restricted populations of lipids. No further effects were observed above a level of 10 mol melittin/mol Ca-ATPase. In the spectra from control and melittin-containing samples, the fraction of restricted lipids decreased significantly with increasing temperature. The effect of melittin was similar to that of decreased temperature, i.e., each spectrum obtained in the presence of melittin (10:1) was nearly identical to the spectrum obtained without melittin at a temperature approximately 5 degrees C lower. The results suggest that the principal effect of melittin on SR membranes is to induce protein aggregation and this in turn, augmented by direct binding of melittin to the lipid, is responsible for the observed decreases in lipid mobility. Protein aggregation is concluded to be the main cause of inactivation of the Ca-ATPase by melittin, with possible modulation also by the decrease in mobility of the boundary layer lipids.