Diminished carbonyl reactivity, a biochemical abnormality in aminonucleoside-nephrotic rat skin α- and ß-collagen

Abstract

Comparative study of chromatographically purified α- and ß-collagen fractions of normal and aminonucleoside-nephrotic rat skin collagen reveals that both the rate and extent of reactivity of such fractions with the highly specific and sensitive carbonyl reagent, N-methyl-benzothiazolone hydrazone hydrochloride (MBTH), are diminished in skin collagen from the aminonucleoside-treated rats. Mechanistically this might be explained either by (a) an inhibitory effect of aminonucleoside on the initial step in the formation of a type of collagen cross-linking, involving enzymatically catalyzed oxidative deamination of the ϵ-amino group of lysyl and/or hydroxylysyl residues in the collagen chains to their respective semi-aldehyde residues; or (b) by direct interaction of aminonucleoside with the semi-aldehyde groups, thus effectively reducing their reactivity with MBTH. Elimination of the latter from consideration would seem to be indicated, however, by the absence of any of the identifying spectral characteristics of the aminonucleoside in skin α- and ß-collagen of aminonucleoside-treated rats, as well as in spectra of exhaustively dialyzed normal rat skin α-collagen which had been incubated for several hours with aminonucleoside. Correlation of the diminished carbonyl reactivity with possible inhibitory effects of aminonucleoside on a specific skin lysyl oxidase activity has not yet been accomplished. Evidence suggesting in vitro inhibitory effects of aminonucleoside on thick embryo cartilage lysyl oxidase activity has been obtained, however, by a modified monoamine oxidase assay. Incubation of cartilage extracts, which contain lysyl oxidase activity (demonstrated by tritium release from [6- 3H]-elastin substrate), with 10 −3 M aminonucleoside prior to assay, in a system utilizing lysine-vasopressin as a substrate, resulted in a considerable reduction in the production of hydrogen peroxide, measured by a sensitive fluorometric procedure.