Effect of lipids on glycoprotein sulphotransferase activity in rat submandibular salivary glands.

Although glycoprotein sulphation has been implicated in the processing of salivary mucin, little is known about the regulation of the enzyme responsible for this event. Using desulphated glycoprotein as sulphate acceptor, the glycoprotein sulphotransferase (GPST) from Golgi membranes of submandibular salivary gland was used to study the effect of various lipids on its activity. The GPST activity in the Golgi membrane was 0.7 pmol/mg protein per min and the activity was extractable by Triton S-100. The Km of the solubilized GPST for glycoprotein and 3'-phosphoadenosine 5'-phosphosulphate (PAPS) were 11 and 0.2 microM, respectively. Among the various lipids tested, phosphatidylinositol and sphingosine stimulated the GPST activity, while other lipids such as sphingomyelin, phosphatidylcholine and phosphatidylserine did not produce a significant effect. At 12 mol% (when expressed as mol% of sphingosine to total phospholipids plus Triton X-100) of sphingosine concentration, the enzyme activity was increased nearly 1.7-fold. The stimulatory effect of sphingosine was accompanied by a significant decrease in Km for glycoprotein from 11 to 2 microM but the increase in Vmax was small. In contrast, the sphingosine effect did not change the Km for PAPS but increased the Vmax nearly two fold. Of the two sphingosine analogues tested, threosphinganine and erythrosphinganine had a lesser stimulatory effect than sphingosine. Stearylamine was partially active, whereas the amino acids (glutamate, aspartate, glutamine, asparagine and serine) were not. These observations and our earlier finding of tyrosylprotein sulphotransferase inhibition by sphingosine demonstrate diverse sphingosine effects on the post-translational sulphation involved in the processing of salivary proteins and suggest an important role for sphingosine in the regulation of salivary protein sulphation.