Beavis A D, Brannan R D, Garlid K D
J Biol Chem. 1985 Nov 5;260(25):13424-33.
The amount of light scattered by a mitochondrial suspension depends on matrix volume (Tedeschi, H., and Harris, D.L. (1955) Arch. Biochem. Biophys. 58, 52-67), a correlation which has been extensively exploited for qualitative studies of solute transport across the inner membrane. To obtain reliable, quantitative estimates of solute transport, it is first necessary to characterize the factors determining mitochondrial light scattering. We show that the dependence of absorbance on mitochondrial concentration can be linearized, resulting in an intrinsic light scattering parameter which is independent of the concentration and source of mitochondria. We show that the absorbance osmotic curve is segmentally linear, exhibiting discontinuities which disappear irreversibly following preswelling. In contrast, direct measurements reveal matrix volume to be reversibly and linearly dependent on inverse osmolality. This divergence is a consequence of the fact that the optical technique samples total particle volume, including contributions from folded membranes and trapped medium. These contributions are minimized by structural components, such as intermembrane connections and the outer membrane, which contribute to efficient packaging of the mitochondrion. When these structures are broken, the mitochondrion cannot return to its native state. We observe that the swelling-induced, irreversible transition from efficient packaging to a random packing state begins at a matrix volume of 1.9 microliter/mg and is complete at 3.1 microliter/mg. These findings complicate the interpretation of light scattering results but do not appear to present an insurmountable obstacle to the quantitative application of this technique to transport kinetics.
