Hydrolysis of S-aryl-cysteinylglycine conjugates catalyzed by porcine kidney cortex membrane dipeptidase

Abstract

We gratefully acknowledge the assistance of Nikole Freeman; Aaron Witham

The mercapturic acid pathway

“…Three dipeptidases have been demonstrated to catalyze the hydrolysis of Cys-Gly S-conjugates to Cys S-conjugates and Gly: membrane dipeptidase (DPEP1), aminopeptidase N (APN), and leucyl aminopeptidase (LAP) (Tate 1980;Commandeur et al 1995;Cooper and Hanigan 2010;Barrett et al 2012;Poon and Josephy 2012). DPEP1 and APN are membrane-bound, homodimeric proteins, whereas LAP is a soluble homohexamer.…”

Experimental colitis and malnutrition differentially affect the metabolism of glutathione and related sulfhydryl metabolites in different tissues

“…The intracellular concentrations of CYS-GLY have been thought to be important in the control of GSH metabolism. The hydrolysis of CYS-GLY has previously been reported in pig kidney [48] because the kidney is the tissue responsible for the majority of GSH breakdown, removing approximately 80-90 % of first-pass plasma GSH [49][50][51]. Our study appears to be the first to quantitate the amount of CYS-GLY prior to hydrolysis in vivo and in multiple tissues.…”

Donor substrate specificity of bovine kidney gamma-glutamyltransferase

“…GSH conjugates were prepared and sent for ESI-LC-MS as described previously (Poon and Josephy, 2012). Liquid chromatography-mass spectrometry analyses were performed at the Advanced Analysis Centre, University of Guelph, on a Dionex UHPLC UltiMate® 3000 liquid chromatograph interfaced to an AmaZon SL® ion trap mass spectrometer (Bruker Daltonics, Billerica, MA, USA).…”

“…GSH conjugates dissolved in 100 mM Tris-HCl (pH 8.0) buffer were used for UV-visible spectrometry; crude products were used to determine yield; recrystallized conjugates were used for melting point analysis. ESI-LC-MS analysis was performed as described previously (Poon and Josephy, 2012); the mass spectra of the GSH conjugates are found in Appendix A.…”

“…Recrystallized GSH conjugates were used for melting point determinations. ESI-LC-MS analysis was performed as described previously (Poon and Josephy, 2012). Single point determinations were performed for yields, UV absorbance peak wavelengths, melting points and mass-to charge ratios.…”