Two Migration Inhibitory Factors Differ in Density and Susceptibility to Neuraminidase and Proteinases

Abstract

Abstract It has been shown that migration inhibitory factor (MIF) from guinea pigs can be separated into two subfractions. These are pH3-MIF (isoelectric point of 3.0 to 4.5 and apparent m.w. of 65,000) and pH5-MIF (isoelectric point of 5.0 to 5.5 and apparent m.w. of 25,000 to 40,000). Upon centrifugation of pH3-MIF in CsCl gradients, activity was found in the density range of 1.38 to 1.46 g/ml. pH5-MIF banded in CsCl gradients in fractions with a density ranging from 1.29 to 1.38 g/ml. Thus, pH3-MIF bands with glycoproteins having a high carbohydrate content. pH5-MIF has a density similar to pure proteins or to glycoprotein with low carbohydrate content. This interpretation is reinforced by the finding that pH3-MIF is neuraminidase sensitive, whereas pH5-MIF is neuraminidase resistant. Furthermore, the activity of pH5-MIF but not that of pH3-MIF was destroyed by treatment with trypsin. In contrast, chymotrypsin inactivated both MIF species. Thrombin and plasmin, on the other hand, did not inactivate pH5-MIF or pH3-MIF. The activity of pH5-MIF was also destroyed by incubation with peritoneal exudate cells, whereas the activity of pH3-MIF was unaltered after incubation with peritoneal exudate cells. Reaction of the peritoneal exudate cells with diisopropylfluorophosphate abolished their ability to inactivate pH5-MIF. Thus, inactivation of MIF is caused by a cell-associated serine protease(s) that may play a role in the regulation of the macrophage's response to one MIF species.