Montejo J M, Naqvi K R, Lillo M P, González-Rodríguez J, Acuña A U
Instituto de Química Física, CSIC, Serrano, Madrid, Spain.
Biochemistry. 1992 Aug 25;31(33):7580-6. doi: 10.1021/bi00148a020.
The rotational motions of human fibrinogen in solution at 20 degrees C have been examined, in the 0.2-12-microseconds time range, by measuring the laser-induced dichroism of the triplet state of an erythrosin probe covalently bonded to the protein. The decay of the anisotropy was multiexponential, and up to three correlation times (phi 1 = 380 +/- 50 ns, phi 2 = 1.1 +/- 0.1 microseconds, and phi 3 = 3.3 +/- 0.6 microseconds) were needed to obtain a satisfactory analysis. The experimental data are consistent with the brownian motions of an elongated, rigid particle. If the correlation times are combined with previous data on the intrinsic viscosity of fibrinogen, the rotational and translational diffusive properties of the protein can be reproduced with high accuracy by idealizing it as an elongated ellipsoid of revolution with dimensions (2a x 2b) of (54 +/- 6) x (7.2 +/- 0.5) nm, having rotational diffusion constants of D parallel = (6.2 +/- 0.7) x 10(5) s-1 and D perpendicular = (5 +/- 1) x 10(4) s-1. The possibility of Ca(2+)-dependent changes in the rigidity or conformation of fibrinogen was excluded by examining the submicrosecond time-resolved fluorescence depolarization of 1-methylpyrene conjugates of the protein in the presence of different calcium concentrations. Although there are inherent difficulties to extrapolate the data on isolated fibrinogen molecules to the polymerizing species, this relatively stiff conformation meets the requirements of the classical half-staggered double-stranded model of fibrin polymerization rather better than those of the recently proposed interlocked single-stranded mechanism.
通过测量与蛋白质共价结合的赤藓红探针三重态的激光诱导二色性，在0.2 - 12微秒的时间范围内，研究了20℃下溶液中人类纤维蛋白原的旋转运动。各向异性的衰减是多指数的，需要多达三个相关时间（φ1 = 380 ± 50纳秒，φ2 = 1.1 ± 0.1微秒，φ3 = 3.3 ± 0.6微秒）才能获得令人满意的分析结果。实验数据与细长刚性颗粒的布朗运动一致。如果将相关时间与先前关于纤维蛋白原特性粘度的数据相结合，将该蛋白质理想化为尺寸为（54 ± 6）×（7.2 ± 0.5）纳米的细长旋转椭球体，其平行旋转扩散常数为D平行 =（6.2 ± 0.7）× 10⁵ 秒⁻¹，垂直旋转扩散常数为D垂直 =（5 ± 1）× 10⁴ 秒⁻¹，就可以高精度地再现该蛋白质的旋转和平移扩散特性。通过在不同钙浓度存在下检查该蛋白质的1 - 甲基芘结合物的亚微秒时间分辨荧光去极化，排除了纤维蛋白原刚性或构象中钙依赖性变化的可能性。尽管将关于分离的纤维蛋白原分子的数据外推到聚合物种存在固有困难，但这种相对僵硬的构象比最近提出的互锁单链机制更符合纤维蛋白聚合经典半交错双链模型的要求。
