We have identified and characterized a gene family comprising at least four genes encoding cathepsin L-like cysteine proteinases (SCPs) in Sitophilus zeamais, Coleoptera. A cDNA clone corresponding to a major mRNA species of the gene family in adult insects codes for a cysteine proteinase (CP) of 338 amino acid residues, and two genomic clones code for similar CPs of 331 residues whose COOH-terminal structures are seven amino acids shorter. These three SCPs show high sequence similarity to one another as well as to other insect and mammalian cathepsin L-like proteinases. A polyclonal antibody raised against bacterially expressed SCP was used as a probe to examine the molecular forms and distribution of the enzyme. SCP exists in both proenzyme and mature forms in larvae, pupae, and adults, and the proenzyme converts in vitro into the mature form at acidic pH. Immunohistochemical analysis showed that SCP is present in several tissues including alimentary organs and germ cells. In alimentary organs, SCP is distributed in the gastric caeca, but not the midgut. SCP is also present in genital organs, especially in oocytes and nurse cells, where it exists at high levels. These results indicate that SCP plays a variety of physiological roles including a role in food digestion.