Myeloperoxidase-dependent Inactivation of Surfactant Protein D in Vitro and in Vivo

Sadis Matalon,Barbara Mcdonald,Azzaq Belaaouaj,Rym Benabid,Rachel Boxio,Erika C Crouch,Baohai Shao,Tim O Hirche,Jay Heinecke,Julien Wartelle

doi:10.1074/jbc.m109.097048

Abstract

Surfactant protein D (SP-D) plays diverse and important roles in innate immunity and pulmonary homeostasis. Neutrophils and myeloperoxidase (MPO) colocalized with SP-D in a murine bacterial pneumonia model of acute inflammation, suggesting that MPO-derived reactive species might alter the function of SP-D. Exposure of SP-D to the complete MPO-H(2)O(2)-halide system caused loss of SP-D-dependent aggregating activity. Hypochlorous acid (HOCl), the major oxidant generated by MPO, caused a similar loss of aggregating activity, which was accompanied by the generation of abnormal disulfide-cross-linked oligomers. A full-length SP-D mutant lacking N-terminal cysteine residues and truncation mutants lacking the N-terminal domains were resistant to the oxidant-induced alterations in disulfide bonding. Mass spectroscopy of HOCl-treated human SP-D demonstrated several modifications, but none involved key ligand binding residues. There was detectable oxidation of cysteine 15, but no HOCl-induced cysteine modifications were observed in the C-terminal lectin domain. Together, the findings localize abnormal disulfide cross-links to the N-terminal domain. MPO-deficient mice showed decreased cross-linking of SP-D and increased SP-D-dependent aggregating activity in the pneumonia model. Thus, MPO-derived oxidants can lead to modifications of SP-D structure with associated alterations in its characteristic aggregating activity.

Highlights

Surfactant protein D (SP-D) consists of one or more trimeric subunits, each with four structurally distinct domains [5, 6]
SP-D and MPO Are Co-distributed at Sites of Acute Inflammation—In conjunction with previous studies of SP-D degradation by neutrophil serine proteases, we implemented a murine model of P. aeruginosa pneumonia to study neutrophil-dependent modifications
Mice were challenged with a sublethal dose of P. aeruginosa sufficient to elicit and activate neutrophils in SP-D containing regions of the lung [14]

Summary

Introduction

SP-D consists of one or more trimeric subunits, each with four structurally distinct domains [5, 6]. Hypochlorous acid (HOCl), the major oxidant generated by MPO, caused a similar loss of aggregating activity, which was accompanied by the generation of abnormal disulfide-cross-linked oligomers. MPO-deficient mice showed decreased cross-linking of SP-D and increased SP-D-dependent aggregating activity in the pneumonia model.

Results

Conclusion