A monomeric Ca²⁺-dependent protease (CDP I) of 39 kDa active at neutral pH has been purified from the aquatic fungusAllomyces arbuscula.The enzyme elutes at NaCl molarity of 0.07 M from the DEAE (DE52)-cellulose columns in contrast to the second Ca²⁺-dependent protease (CDP II) characterized earlier which elutes at 0.18 M NaCl. The enzyme has no basal activity in the absence of Ca²⁺ and requires 1.7 mM Ca²⁺ for half maximum activation of thein vitroenzyme activity. The enzyme prefers substrates with Arg in P₁position but this specificity also depends strongly on the nature of the subsite residues, for example Pro in P₂ position. The enzyme is glycosylated and contains essential cysteine residues in the active site. It appears to be an atypical cysteine protease as it is inactivated to varying degree with some serine protease inhibitors.