A cysteine proteinase inhibitor was found in culture media of Candida albicans. Purification to homogeneity of the inhibitor was performed by carboxymethyl-papain-Sepharose affinity, DE-52 ion-exchange, and reverse- phase high performance liquid chromatographies. The purified inhibitor had an M(r) of 15 kDa and a pI of 4.9. It was more stable to heat and pH than most proteins. The N-terminal sequence of the first 30 residues demonstrated high similarity with that of human cystatin A. Thus, C. albicans cysteine proteinase inhibitor seems to belong to the cystatin superfamily. The inhibitor activity of the yeast cellular form was 4.0 times higher than that of the hyphal cellular form in 7-day culture media. It is suggested that the inhibitor has regulatory functions similar to those of its counterpart proteinases in the invasion of host cells.