Association of prolyl hydroxylase activity with membranes

Abstract

Addition of ionic and nonionic detergents to whole homogenates of liver, kidney and lung prepared by a mild homogenization technique resulted in a two- to three-fold increase of prolyl hydroxylase activity. After subcellular fractionation of whole homogenates of liver, particulate and supernatant fractions were incubated in the presence and absence of triton X-100 and assayed for prolyl hydroxylase activity. All particulate fractions tested were able to release significant amounts of prolyl hydroxylase activity in the presence of triton. The release of enzyme activity by triton was observed with the 1000 × g and 17,000 × g supernatants but not with the 105,000 × g supernatant; thus indicating that detergent does not activate soluble enzyme nor make the substrate more accessible to hydroxylation by the enzyme during incubation. Rigorous homogenization of the 17,000 × g particulate fraction with the Polytron ST system resulted in a substantial loss of the amount of prolyl hydroxylase activity released by treatment with triton. These data suggest that a significant amount of prolyl hydroxylase activity is associated with membranes under physiological conditions.