BLEOMYCIN HYDROLASE (BLH1P), A MULTI-SITED THIOL PROTEASE IN SEARCH OF A DISTINCT PHYSIOLOGICAL-ROLE

Bleomycin hydrolase, Blh1p, from yeast was co-purified with Gce1p, a c AMP-binding ectoprotein, anchored to the plasma membrane by a glycosyl -phosphatidylinositol (GPI) anchor. Blh1p is a hydrophilic thiol prote ase lacking transmembrane domains. We have used polyclonal antibodies to study the topology of the over-expressed protein in yeast and have found that it is amphitropic. Part of Blh1p is associated with plasma membranes and most of the rest occurs in the cytosol. Both the growth conditions and calcium were found to have minor influences on the topo logy of Blh1p, in that glucose and the earth-alkali ion slightly enhan ced recruitment to the membrane. We have examined the possibility that co-purification of Blh1p with Gce1p has a functional basis, and have observed that over-expression of BLH1 in yeast leads to an acceleratio n of the glucose-induced amphiphilic to hydrophilic conversion of Gce1 p, wherein Blh1p could either directly catalyse the proteolytic remova l of the polar headgroup of the GPI anchor subsequent to an initial li polytic cleavage by a GPI-specific phospholipase C or indirectly modul ate the reaction. The data show that a thiol protease is involved, but point to an indirect role of Blh1p in GPI processing. Proteases with similar or overlapping substrate specificity are likely to exist, sinc e deletion of BLH1 neither entails a growth defect on any carbon sourc e tested, nor the loss of proteolytic processing of the GPI anchor of Gce1p. Reduced proteolytic GPI processing is, however, observed in the blh1 mutant and the corresponding acceleration in the respective BLH1 multi-copy transformant.