ESR studies of stearic acid binding to bovine serum albumin.
ESR studies of stearic acid binding to bovine serum albumin.
复制标题
硬脂酸与牛血清白蛋白结合的 ESR 研究。
DOI：
10.1016/0304-4165(90)90039-y
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发表时间：
1990
期刊：
影响因子：
--
通讯作者：
Freed,JH
中科院分区：
文献类型：
--
作者：
Ge,MT;Rananavare,SB;Freed,JH
用慢速ESR模拟的非线性最小二乘法分析了一系列链标记硬脂酸(5-、7-、12-和16-DSA)和硬脂酸甲酯(5-、7-、12-和16-DMS)与牛血清白蛋白(BSA)高亲和力结合部位的ESR波谱。运动分析表明，这些硬脂酸盐围绕垂直于长碳氢链的轴的旋转扩散受到极大的阻碍，这表明它们被紧密地固定在蛋白质的通道中。各系列的各向同性超精细分裂A0的比较表明，5-、16-DSA和16-DMS的A0值比其他自旋标记物大。此外，DSA和DMS的16-C位置上的标记相对于其他位置表现出显著增加的运动。这些观察表明，通道开始于链的5-C，结束于13-C和15-C之间的某个地方，导致通道长度的估计为11±1Å。与类似的硬脂酸标记相比，硬脂酸甲酯标记绕链轴旋转的速度明显更快，这表明脂肪酸与BSA结合是双氢键机制。酸形成两个氢键的能力显然更严格地将其固定在蛋白质中，阻止围绕其中一个氢键旋转。双氢键机制与酸的带负电的羧酸盐和带正电的精氨酸基团或两个赖氨酸残基的带正电的ω-氨基之间形成盐桥最一致。对DSA结合的pH依赖性的ESR研究表明，与赖氨酸形成的盐桥至少是BSA长链脂肪酸结合部位的一部分。
The ESR spectra of a series of chain-labelled doxyl stearic acids (5-, 7-, 12-and 16-DSA) and doxyl methyl stearates (5-, 7-, 12-and 16-DMS) bound to the high-affinity binding sites of bovine serum albumin (BSA) have been analyzed using nonlinear least-squares fitting of slow-motional ESR simulation. The motional analysis reveals that the rotational diffusion of these stearates around the axis perpendicular to the long hydrocarbon chain is greatly hindered, suggesting that they are held tightly in a channel of the protein. Comparison of the isotropic hyperfine splitting, A 0, among each series shows that 5-and 16-DSA and 16-DMS have larger A 0 values than the other spin labels. In addition, labels at the 16-C position of both DSA and DMS exhibit significantly increased motion relative to the other positions. These observations suggest that the channel starts at 5-C of the chain and ends somewhere between 13-C and 15-C, leading to an estimate of 11±1 A ̊ for the lenght of the channel. The methyl stearate lables exhibit significantly faster rotation around the chain axis than the analogous stearic acid labels, suggesting a double hydrogen-bonding mechanism for fatty acid binding to BSA. The ability of the acid to form two hydrogen bonds apparently fixes it more rigidly in the protein preventing rotation about either single hydrogen bond. A double-hydrogen bonding mechanism is most consistent with the formation of a salt bridge between the negatively charged carboxylate of the acid and either a positively charged guanidino group of arginine, or the positively charged ω-amino groups of two lysine residues. An ESR study of the pH dependence of DSA binding indicates that salt bridge formation with lysine is responsible for at least some of the long chain fatty acid binding sites of BSA.