Molecular crowding and viscosity as determinants of translational diffusion of metabolites in subcellular organelles

The role of molecular crowding and viscosity on the apparent translational diffusion coefficient (ADC) of small metabolites was investigated in differ ent subcellular organelles using the pulse-field gradient spin-echo H-1 NMR technique. ADCs of metabolites with increasing radius of gyration (0.7 Ang strom < R-G < 4.5 Angstrom) were measured in the cytoplasm of rat or chicke n erythrocytes, in the nucleus of chicken erythrocytes, and in isolated rat liver mitochondria. Metabolite ADCs in these systems were compared with th e corresponding ADCs determined in model solutions of increasing bulk visco sity but different molecular crowding. For solutions having the same viscos ity, metabolite ADCs decreased with increasing concentration of cosolutes, This effect is adequately described by the modified Stokes-Einstein relatio nship, ADC = k/R-G (1 + 2.5 Phi), where k is a constant for a given tempera ture and Phi is an obstruction factor reporting the fractional volume of so lution occupied by cosolutes, a measure of the molecular crowding in the so lution. Cytoplasmic values of Phi for metabolites of different sizes did no t depend exclusively on metabolite RG but On additional factors including t he chemical nature of the metabolite, the presence of diffusional barriers, and metabolite-specific binding sites, In the case of water, nuclear Phi v alues approached those of the extracellular space while mitochondrial Phi v alues were significantly higher than those of the cytoplasm, Taken together , these results reveal important differences in molecular crowding within t he different subcellular compartments, suggesting considerable diffusional heterogeneity for small metabolites within the different intracellular orga nelles, (C) 1999 Academic Press.