Cationic activation of galactosyltransferase from rat mammary Golgi membranes by polyamines and by basic peptides and proteins

Abstract

Galactosyltransferase (EC 2.4.1.22) requires bivalent metal ions for its activity. However, preparations of this enzyme solubilized from Golgi membranes of lactating rat mammary gland were shown to be activated not only by Mn2+, Ca2+ and Mg2+, but also by spermine, spermidine, lysyl-lysine, ethylenediamine and other diaminoalkanes, and by a range of basic proteins and peptides, including clupeine, histone, polylysine, ribonuclease, pancreatic trypsin inhibitor, cytochrome c, melittin, avidin and myelin basic protein. Both N-acetyl-lactosamine synthetase and lactose synthetase activities were enhanced. A basic protein fraction was isolated from bovine milk and shown to activate galactosyltransferase at low concentrations. The polyanions ATP, casein, chondroitin sulphate and heparin reversed the activation of galactosyltransferase by several of the above substances. Galactosyltransferase, assayed as a lactose synthetase, showed a 10-fold greater affinity for glucose when Mn2+ ions were replaced by clupeine or by ribonuclease as cationic activator. Evidence was obtained for the presence of an endogenous cationic activator in solubilized Golgi membrane preparations which evoked a similar low apparent Km, glucose. The findings are discussed in the light of cationic activations of glycosyltransferases generally, of the porous nature of the Golgi membrane, and of the unlikelihood of bivalent metal ions being the physiological activators of galactosyltransferase. It is suggested that the natural cationic activator of lactose synthetase may be a secretory protein acting in a manner analogous to the enzyme's activation by alpha-lactalbumin. A scheme is proposed for the two-stage synthesis of lactose and phosphorylation of casein within the cell, to accommodate the apparent incompatibility of these two processes.