Growth-subinhibitory nonlytic doses of cilofungin (lipopeptide antibiotic affecting (1,3)-beta-D-glucan synthesis) inhibited the incorporation of 46- to 48-kDa glucan-associated (46K) protein into the growing cell wall of Candida albicans. The purified 46K protein constituent strongly reacted with a monoclonal antibody against enolase, a major cytoplasmic enzyme of the fungus. In addition, two internal fragments of 12- and 15-amino acid residues from a tryptic digest of 46K protein showed 100% identity with amino acids in positions 34-45 and 66-80 of enolase. By immunoelectron microscopy with polyclonal and monoclonal anti-enolase antibodies, the 46K protein was clearly detected in the inner layers of the fungal cell wall. Thus, consistent with the proposed immunogenic and diagnostic roles of enolase in candidiasis, biochemical, immunochemical, and ultrastructural evidence strongly suggest that the cilofungin-susceptible 46K protein is a cell wall-associated form of this enzyme.

The effect of the cilofungin, a beta 1-3 glucan synthase inhibitor, on the incorporation of the glucan associated proteins (GAP) into the mycelial wall of Candida albicans was investigated. For this study sub-inhibitory (< 2 micrograms/ml) doses of cilofungin were employed during the yeast to mycelial transition in a defined chemical medium, at 37 degrees C for 24 hours. Under these conditions, and particularly at the dose of 0.50 micrograms/ml cilofungin exerted a marked effect on GAP incorporation into the mycelial cell wall. The changes were essentially the absence of the two prominent bands of 46 and 31 kDa of the untreated cell wall coupled with an apparent increase in the amount of 55-56 kDa constituent, as well as of a minor constituent of 27-28 kDa. Radiolabel incorporation experiments demonstrated increased synthesis of a 34 kDa GAP, in addition to confirming the absence of the 46 kDa constituent, in mycelial cells under cilofunging treatment. Thus, sub-inhibitory doses of cilofungin may greatly alter the pattern of essential cell wall constituents such as the glucan-associated proteins, suggesting that this drug also has important effects on cell wall structure and fine organization, independent of, or prior to, its principal lytic effect on the fungal organism.