Bombyx cysteine proteinase inhibitor (BCPI) is a novel cysteine proteinase inhibitor. The protein sequence is homologous to the proregions of certain cysteine proteinases. Here we report the mechanism of its inhibition of several cysteine proteinases. BCPI strongly inhibited Bombyx cysteine proteinase (BCP) activity with a Ki=5.9pM, and human cathepsin L with a Ki=6pM. The inhibition obeyedslow-binding kinetics. The inhibition of cathepsin H was much weaker (Ki=82nM), while inhibition of papain (Ki>1μm) and cathepsin B (Ki>4 μM) was negligible. Following incubation with BCP, BCPI was first truncated at the C-terminal end, and then gradually degraded over time. The truncation mainly involved twoC-terminal amino acid residues. Recombinant BCPI lacking the two C-terminal amino acid residues still retained substantial inhibitory activity. Our results indicate that BCPI is a stable and highly selective inhibitor of cathep-sin L-likecysteine proteinases.