Physicochemical characteristics of the glycosaminoglycan-lysosomal enzyme interaction in vitro. A model of control of leucocytic lysosomal activity

Abstract

The activities of 30 different lysosomal enzymes [from human polymorphonuclear leukocytes] were determined in vitro in the presence of the sulfated glycosaminoglycans, heparin and chondroitin sulfate, all the enzymes being measured on a density-gradient-purified lysosomal fraction. Each enzyme was studied as a function of the pH of the incubation medium. In general the presence of sulfated glycosaminoglycans induced a strong pH-dependent inhibition of lysosomal enzymes at pH values lower than 5.0, with full activity at higher pH values. In the particular case of lysozyme [EC 3.2.1.17] and phospholipase A2 the heparin-induced inhibition was maintained in the pH range 4.0-7.0. For certain enzymes, such as acid .beta.-glycerophosphatase, .alpha.-galactosidase [EC 3.2.1.22] acid lipase, lysozyme and phospholipase A2, the pH-dependent behavior obtained in the presence of heparin was quite different to that obtained with chondroitin sulfate, suggesting the existence of physicochemical characteristic factors playing a role in the intermolecular interaction for each of the sulfated glycosaminoglycans studied. Except in the particular case of peroxidase activity, in all other lysosomal enzymes measured, the glycosaminoglycan-enzyme complex formation was a temperature- and time-independent phenomenon. The effects of the ionic strength and pH on this intermolecular interaction reinforce the concept of an electrostatic reversible interaction between anionic groups of the glycosaminoglycans and cationic groups on the enzyme molecule. As leukocytic primary lysosomes have a very acid intragranular pH and large amounts of chondroitin sulfate, this glycosaminoglycan might act as molecular regulator of leukocytic lysosomal activity, by inhibiting lysosomal enzymes when the intragranular pH is below the pI of lysosomal enzymes. This fact, plus the intravacuolar pH changes described during the phagocytic process, might explain the unresponsiveness of lysosomal enzymes against each other existing in primary lysosomes as well as its full activation at pH values occurring in secondary lysosomes during the phagocytic process.