MYOPLASMIC BINDING OF FURA-2 INVESTIGATED BY STEADY-STATE FLUORESCENCE AND ABSORBANCE MEASUREMENTS
MYOPLASMIC BINDING OF FURA-2 INVESTIGATED BY STEADY-STATE FLUORESCENCE AND ABSORBANCE MEASUREMENTS
复制标题
DOI：
10.1016/s0006-3495(88)83045-5
复制
发表时间：
1988-12-01
影响因子：
3.4
通讯作者：
BAYLOR, SM
中科院分区：
文献类型：
--
作者：
KONISHI, M;OLSON, A;BAYLOR, SM
结合的荧光Ca 2+指示剂染料Fura-2细胞内成分已被研究的稳态光学测量。在含有纯化的肌浆蛋白溶液的玻璃毛细管中测量Fura-2的（a）荧光强度、（B）荧光发射各向异性、（c）荧光发射光谱和（d）吸收光谱;还在显微注射Fura-2的青蛙骨骼肌纤维中测量特性B和c。结果表明，超过一半，可能高达85%，在肌浆中的fura-2分子是在蛋白质结合的形式，并结合改变染料的许多特性。例如，Ca 2 +-染料反应的体外表征表明，当fura-2与醛缩酶（一种大而丰富的肌浆蛋白）结合时，染料对Ca 2+的解离常数比在不存在蛋白质的情况下测量的解离常数大三到四倍。由fura-2与细胞质蛋白的细胞内结合引起的问题可能很好地适用于骨骼肌纤维以外的细胞。
Binding of the fluorescent Ca2+ indicator dye fura-2 by intracellular constituents has been investigated by steady-state optical measurements. Fura-2''s (a) fluorescence intensity, (b) fluorescence emission anisotropy, (c) fluorescence emission spectrum, and (d) absorbance spectra were measured in glass capillary tubes containing solutions of purified myoplasmic proteins; properties b and c were also measured in frog skeletal muscle fibers microinjected with fura-2. The results indicate that more than half, and possibly as much as 85%, of fura-2 molecules in myoplasm are in a protein-bound form, and that the binding changes many properties of the dye. For example, in vitro characterization of the Ca2+-dye reaction indicates that when fura-2 is bound to aldolase (a large and abundant myoplasmic protein), the dissociation constant of the dye for Ca2+ is three- to fourfold larger than that measured in the absence of protein. The problems raised by intracellular binding of fura-2 to cytoplasmic proteins may well apply to cells other than skeletal muscle fibers.