Solubilization and Analysis of Mannoprotein Molecules from the Cell Wall of Saccharomyces cerevisiae

Purified walls from Saccharomyces cerevisiae were treated chemically to release intrinsic mannoproteins. Boiling in 2% SDS gave the best results, although treatment in 6 M-urea at room temperature also released significant amounts of mannoprotein radioactivity. Triton X-100, sodium deoxycholate and EDTA were poor solubilizers. Electrophoretic patterns of SDS- or urea-released mannoproteins in SDS-acrylamide gels indicated a great heterogeneity of molecular species, with more than 60 bands. Zymolyase, a glucan-digesting complex, released about half of the mannoproteins, but these species showed an altered mobility on SDS-acrylamide gels and had a lowered capacity for precipitation by ethanol. Action of the enzyme on isolated walls was favoured by dithiothreitol, as is the case with whole cells, and repeated treatments with SDS and Zymolyase released all of the mannoproteins from the wall. Solubilizing treatments other than SDS had a differential effect on recently or formerly incorporated mannoproteins in the wall. The results suggest an asymmetrical arrangement of molecules in the envelope and point to dynamic changes inside the wall as it thickens as a result of cell aging.