P-271 - Nitration alters laminin polymerization

Abstract

Reactive nitrogen species (RNS) are produced by cells at sites of inflammation and can modify proteins. Nitration is used as a biomarker of damage and may contribute to multiple pathologies. We have shown that the extracellular matrix protein laminin (LN) co-localizes with nitrative damage in human atherosclerotic lesions, and that nitration decreases cell binding to LN in vitro. Study aim: To investigate how nitration affects laminin self-assembly and polymerization. Methods: Sites of nitration were determined by LC-MS. Polymerization was induced by pH changes and CaCl2. Dynamic light scattering was used to examine the extent of polymerization. Structure of LN polymers was investigated with scanning electron microscopy (SEM). Results: Extensive nitration was observed on the protein at specific sites. Nitrated LN underwent polymerization, but displayed increased light scattering, indicating altered particle size. SEM showed that untreated LN formed a homogenous ordered structure, whereas nitrated LN displayed a segmented, patchy and disordered pattern. Conclusions: While the extent of LN polymerization is not affected by nitration, the structure of LN polymers is altered. Such alterations in polymer structure could lead to decreased cell adhesion and altered ECM integrity.