Basement membranes provide structural support and convey regulatory signals to cells in diverse tissues. Assembly of collagen IV into a sheet-like network is a fundamental mechanism during the formation of basement membranes. Peroxidasin (PXDN) was recently described to catalyze crosslinking of collagen IV through the formation of sulfilimine bonds.Despite the significance of this pathway in tissue genesis, our understanding of PXDN function is far from complete. In this work we demonstrate that collagen IV crosslinking is a physiological function of mammalian PXDN. Moreover, we carried out structure-function analysis of PXDN to get a better insight into its role in collagen IV synthesis. We identify conserved cysteines in PXDN which mediate the oligomerization of the protein into a trimeric complex. We also demonstrate that oligomerization is not an absolute requirement for enzymatic activity but optimal collagen IV coupling is only catalyzed by the PXDN trimers.Localization experiments of different PXDN mutants in two different cell models revealed that PXDN oligomers, but not monomers, adhere on cell-surface in "hot spots", which represent previously unknown locations of collagen IV crosslinking.

PXDNL is a unique component of the heart with a recently evolved inactivation of peroxidase function. The elevation of PXDNL levels in the failing heart may contribute to ECM dysregulation due to its antagonism of PXDN function.

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