Determination of leucine aminopeptidase using phenylalanyl-3-thia-phenylalanine as substrate

The peptide mimetic L-phenylalanyl-L-3-thiaphenylalanine has been shown to facilitate a sensitive and simple determination of leucine aminopeptidase. A colorimetric assay, employing Ellman's reagent to detect the thiophenol released upon hydrolysis of the dipeptide, has been developed. Under the experimental conditions employed the substrate has a Km of 0.054 mM and a kcat of 5800 min-1 and can distinguish sharply between leucine aminopeptidase and aminopeptidase M.