Incorporation of copper into lysyl oxidase

T. Kosonen, J. Y. Uriu-Hare, M. S. Clegg, Carl L Keen, R. B. Rucker

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33 Scopus citations

Abstract

Lysyl oxidase is a copper-dependent enzyme involved in extracellular processing of collagens and elastin. Although it is known that copper is essential for the functional activity of the enzyme, there is little information on the incorporation of copper. In the present study we examined the insertion of copper into lysyl oxidase using 67Cu in cell-free transcription/translation assays and in normal skin fibroblast culture systems. When a full-length lysyl oxidase cDNA was used as a template for transcription/translation reactions in vitro, unprocessed prolysyl oxidase appeared to bind copper. To examine further the post-translational incorporation of copper into lysyl oxidase, confluent skin fibroblasts were incubated with inhibitors of protein synthesis (cycloheximide, 10 μg/ml), glycosylation (tunicamycin, 10 μg/ml), protein secretion (brefeldin A, 10 μg/ml) and prolysyl oxidase processing (procollagen C-peptidase inhibitor, 2.5 μg/ml) together with 300 μCi of carrier-free 67Cu. It was observed that protein synthesis was a prerequisite for copper incorporation, but inhibition of glycosylation by tunicamycin did not affect the secretion of 67Cu as lysyl oxidase. Brefeldin A inhibited the secretion of 67Cu-labelled lysyl oxidase by 46%, but the intracellular incorporation of copper into lysyl oxidase was not affected. In addition, the inhibition of the extracellular proteolytic processing of prolysyl oxidase to lysyl oxidase had minimal effects on the secretion of protein-bound 67Cu. Our results indicate that, similar to caeruloplasmin processing, copper is inserted into prolysyl oxidase independently of glycosylation.

Original languageEnglish (US)
Pages (from-to)283-289
Number of pages7
JournalBiochemical Journal
Volume327
Issue number1
StatePublished - 1997

ASJC Scopus subject areas

  • Biochemistry

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    Kosonen, T., Uriu-Hare, J. Y., Clegg, M. S., Keen, C. L., & Rucker, R. B. (1997). Incorporation of copper into lysyl oxidase. Biochemical Journal, 327(1), 283-289.