Y. Ikehara, S. Ogata, Y. Misumi
1994
Citations
1
Influential Citations
22
Citations
Journal
Methods in enzymology
Abstract
Publisher Summary This chapter describes assay, purification, and properties of dipeptidyl-peptidase IV (DPPIV) from rat liver. DPPIV is a serine protease that cleaves N-terminal dipeptides from oligo- and polypeptides with a penultimate prolyl residue. It is a membrane-bound glycoprotein localized on the cell surface (ectoenzyme), in contrast to other DPPs localized in the lysosome and in the cytoplasm. The release of p -nitroaniline from dipeptidyl- p -nitroanilides is photometrically determined at 385 nm after the indicated incubation period. Gly-Pro- p -nitroanilide tosylate is used as a routine substrate, but any Xaa-Pro- p -nitroanilide can be used for the assay. Two forms of DPPIV, papain-cleaved soluble form and Triton X-100- solubilized membrane form, are purified from plasma membranes of rat liver. DPPIV is known to survive autolysis for up to 24 hr at pH 4 and 37°, with a recovery of more than 50% of the activity. DPPIV also exhibits a strong preference for substrates having a penultimate prolyl residue, which can be replaced only by alanine and hydroxyproline with much lower rates of hydrolysis.