Molony L, McCaslin D, Abernethy J, Paschal B, Burridge K
J Biol Chem. 1987 Jun 5;262(16):7790-5.
This paper describes the structural and biochemical characterization of talin, a protein localized to various cellular sites where bundles of actin filaments attach to the plasma membrane. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the protein has a molecular mass of 225,000 +/- 5,000 daltons. Hydrodynamic measurements at protein concentrations less than 0.72 mg/ml indicate a monomeric protein with a native molecular mass of 213,000 +/- 15,000 daltons. Sedimentation equilibrium experiments indicate self-association at protein concentrations of 0.72 mg/ml and higher. The data suggest that this self-association is a simple monomer:dimer equilibrium over the range of concentrations observed. At low protein concentrations where talin is a monomer, the Stokes radius and sedimentation coefficient vary with ionic strength. Under low ionic strength conditions (5-20 mM NaCl), talin has a Stokes radius of 6.5 nm and a sedimentation value of 9.4, suggesting an asymmetric globular molecule; whereas under high ionic strength conditions (200 mM NaCl), the Stokes radius increases to 7.7 nm and the sedimentation coefficient decreases to 8.8, suggesting a more elongated protein. This conformation change is confirmed by electron microscopy which reveals a more globular protein at low ionic strength which unfolds to become an elongated flexible molecule as the ionic strength is increased to physiological and higher levels. The amino acid composition of talin indicates a low level of aromatic residues, consistent with its relatively low extinction coefficient, talin has an isoelectric point between pH 6.7 and 6.8 based on isoelectric focusing. The detailed purification of talin is described.
本文描述了踝蛋白的结构和生化特性，踝蛋白是一种定位于肌动蛋白丝束附着于质膜的各种细胞位点的蛋白质。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳，该蛋白质的分子量为225,000±5,000道尔顿。在蛋白质浓度低于0.72 mg/ml时进行的流体动力学测量表明，该蛋白质为单体，天然分子量为213,000±15,000道尔顿。沉降平衡实验表明，在蛋白质浓度为0.72 mg/ml及更高时会发生自缔合。数据表明，在观察到的浓度范围内，这种自缔合是一种简单的单体：二聚体平衡。在踝蛋白为单体的低蛋白质浓度下，斯托克斯半径和沉降系数随离子强度而变化。在低离子强度条件下（5-20 mM NaCl），踝蛋白的斯托克斯半径为6.5 nm，沉降值为9.4，表明是一个不对称的球状分子；而在高离子强度条件下（200 mM NaCl），斯托克斯半径增加到7.7 nm，沉降系数降低到8.8，表明是一个更细长的蛋白质。电子显微镜证实了这种构象变化，其显示在低离子强度下是一个更球状的蛋白质，随着离子强度增加到生理水平及更高水平，该蛋白质展开成为一个细长的柔性分子。踝蛋白的氨基酸组成表明芳香族残基含量较低，这与其相对较低的消光系数一致，基于等电聚焦，踝蛋白的等电点在pH 6.7至6.8之间。本文还描述了踝蛋白的详细纯化过程。
