Enzymatic Degradation of Poly[N5-(2-Hydroxyethyl)-L-Glutamine] and Poly[N5-(2-Hydroxyethyl)-L-Glutamine-Stat-L-Glutamic Acid]: Analysis of Final Degradation Products

Abstract

The enzymatic degradation of N⁵-(2-hydroxyethyl)-L-glutamine homopolymer (PHEG) and its statistical copolymer with L-glutamic acid, (P[HEG-stat-Glu]) by papain, pronase and leucine aminopeptidase (LAP) was investigated with the aim to evaluate the role of endopeptidase and exopeptidase mechanisms of cleavage and to identify ultimate degradation products. The degradation products were analysed by size-exclusion chromatography, using amino end-groups labelled with fluorescamine, and by thin-layer chromatography. Papain cleaved both polymers by the endopeptidase mechanism only and the smallest degradation fragments thus produced were the size of tetramers. These fragments were susceptible to further degradation by an exopeptidase, i.e., leucine aminopeptidase. A combined treatment of polymers by papain and LAP ultimately yielded monomers, HEG only or HEG and glutamic acid for PHEG or P[HEG-stat-Glu] copolymer, respectively. Both endopeptidase and exopeptidase mechanisms were active in the degradation of the polymers under study by pronase. As the enzymes with analogous specificities are usually present in mammalian tissues, the feasibility of complete degradation of these polymers in vivo is supported.