Calprotectin is a protein in neutrophil cytoplasm and abscess fluids that appears to inhibit microbial growth through competition for zinc. This study was undertaken to identify specific sites that might be responsible for the protein's zinc-binding antimicrobial activity. A review of published calprotectin amino acid sequences revealed the HEXXH motif of thermolysin-type metalloproteases and an HHH polyhistidine sequence near the C-terminus of the protein's heavy chain. Reagent polyhistidine had antimicrobial activity against Candida albicans similar to that of calprotectin. Also, one type of HEXXH-containing thermolysin was inactive in the C. albicans assay, whereas a protein tagged with six C-terminal histidines did have calprotectin-like zinc-reversible antimicrobial activity. The activity of polyhistidine, as well as that of calprotectin itself, was reversed by addition of zinc or treatment with the histidine-modifying compound diethylpyrocarbonate. These results suggest that calprotectin's antimicrobial activity may be related to certain histidine-based zinc-binding sequences.