Enzymes are critical for the biochemistry of life, as they make chemical reactions happen that could not otherwise occur. The Proteolysis Lab studies a family of enzymes that trims individual amino acids from the carboxy-terminal ends of peptides and proteins. This can be for the purpose of digestion, but is often to regulate the activity of important peptides and proteins. Currently we are exploring the mechanism and function of two such enzymes. Ecm14L is derived from the mushroom Agaricus bisporus. Its function is unknown, but it appears to function through a unique mechanism, in which a tyrosine plays the catalytic role of general base. Carboxypeptidase O (CPO) is an enzyme found in most vertebrates. Several lines of evidence suggest that CPO may be a folate hydrolase, including expression in immune cells, ability to cleave glutamate-extended folate, and activation by glutamate-extended folates. Results from these two lines of inquiry will lead to a deeper understanding of proteolytic function and diversity.