Glucan synthase complex of Aspergillus fumigatus

The glucan synthase complex of the human pathogenic mold Aspergillus fumiga tus has been investigated. The genes encoding the putative catalytic subuni t Fks1p and four Rho proteins of A. fumigatus were cloned and sequenced. Se quence analysis showed that AfFks1p was a transmembrane protein very simila r to other Fksp proteins in yeasts and in Aspergillus nidulans. Heterologou s expression of the conserved internal hydrophilic domain of AfFks1p aas ac hieved in Escherichia coli. Anti-Fks1p antibodies labeled the apex of the g erm tube, as did aniline blue fluorochrome, which was specific for beta (1- 3) glucans, showing that AfFks1p colocalized with the newly synthesized bet a (1-3) glucans. AfRHO1, the most homologous gene to RHO1 of Saccharomyces cerevisiae, was studied for the first time in a filamentous fungus. AfRho p roteins have GTP binding and hydrolysis consensus sequences identical to th ose of yeast Rho proteins and have a slightly modified geranylation site in AfRho1p and AfRho3p. Purification of the glucan synthase complex by produc t entrapment led to the enrichment of four proteins: Fks1p, Rho1p, a 100-kD a protein homologous to a membrane Hf-ATPase, and a 160-kDa protein which w as labeled by an anti-beta (13) glucan antibody and was homologous to ABC b acterial beta (1-2) glucan transporters.