1. Interaction of Surfactant Protein A with Peroxiredoxin 6 Regulates Phospholipase A2 Activity
- Author
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Yefim Manevich, Chandra Dodia, Aron B. Fisher, Yongzheng Wu, Kevin Yu, Sheldon I. Feinstein, James L. Baldwin, University of Pennsylvania [Philadelphia], and This work was supported by Grant HL19737 from the National Institutes of Health. The results of this study have been presented in part at EB2004 in Washington, DC and EB2005 in San Diego, CA. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked 'advertisement' in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
- Subjects
Male ,MESH: Hydrogen-Ion Concentration ,MESH: Pulmonary Surfactant-Associated Protein A ,MESH: Peroxiredoxins ,MESH: Peroxiredoxin VI ,Biochemistry ,law.invention ,MESH: Peroxidases ,MESH: Recombinant Proteins ,Rats, Sprague-Dawley ,Mice ,[SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases ,MESH: Animals ,Lung ,MESH: Immunoblotting ,Pulmonary Surfactant-Associated Protein A ,Sepharose ,Hydrogen-Ion Concentration ,MESH: Surface Plasmon Resonance ,MESH: Epithelial Cells ,Recombinant DNA ,1,2-Dipalmitoylphosphatidylcholine ,MESH: Chromatography ,Gene Expression Regulation, Enzymologic ,Phospholipases A ,Phospholipase A2 ,MESH: Protein Binding ,Humans ,MESH: Scattering, Radiation ,Molecular Biology ,MESH: Phospholipids ,MESH: Humans ,Dose-Response Relationship, Drug ,Macrophages ,[SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology ,Epithelial Cells ,Peroxiredoxins ,Surface Plasmon Resonance ,MESH: Light ,MESH: Cell Line ,Mice, Inbred C57BL ,Phospholipases A2 ,Microscopy, Fluorescence ,chemistry ,Dipalmitoylphosphatidylcholine ,MESH: Glutathione Peroxidase ,Peroxiredoxin VI ,Time Factors ,Light ,MESH: 1,2-Dipalmitoylphosphatidylcholine ,MESH: Microscopy, Fluorescence ,MESH: Rats, Sprague-Dawley ,MESH: Dose-Response Relationship, Drug ,chemistry.chemical_compound ,Pulmonary surfactant ,Nickel ,law ,MESH: Nickel ,Scattering, Radiation ,Surface plasmon resonance ,MESH: Phospholipases A2 ,Phospholipids ,Chromatography ,MESH: Kinetics ,biology ,respiratory system ,MESH: Gene Expression Regulation ,Recombinant Proteins ,Cross-Linking Reagents ,Peroxidases ,MESH: Calcium ,Agarose ,MESH: Phospholipases A ,lipids (amino acids, peptides, and proteins) ,Dimerization ,Protein Binding ,MESH: Sepharose ,MESH: Rats ,MESH: Cross-Linking Reagents ,Immunoblotting ,Phospholipid ,Cell Line ,MESH: Mice, Inbred C57BL ,Animals ,Immunoprecipitation ,MESH: Lung ,MESH: Mice ,Glutathione Peroxidase ,MESH: Immunoprecipitation ,MESH: Time Factors ,MESH: Macrophages ,Cell Biology ,MESH: Male ,Rats ,Surfactant protein A ,Kinetics ,MESH: Dimerization ,biology.protein ,Calcium - Abstract
International audience; Peroxiredoxin 6 (Prdx6) is a "moonlighting" protein with both GSH peroxidase and phospholipase A(2) (PLA(2)) activities. This protein is responsible for degradation of internalized dipalmitoylphosphatidylcholine, the major phospholipid component of lung surfactant. The PLA(2) activity is inhibited by surfactant protein A (SP-A). We postulate that SP-A regulates the PLA(2) activity of Prdx6 through direct protein-protein interaction. Recombinant human Prdx6 and SP-A isolated from human alveolar proteinosis fluid were studied. Measurement of kinetic constants at pH 4.0 (maximal PLA(2) activity) showed K(m)0.35 mm and V(max) 138 nmol/min/mg of protein. SP-A inhibited PLA(2) activity non-competitively with K(i) 10 mug/ml and was Ca(2+) -independent. Activity at pH 7.4 was approximately 50% less, and inhibition by SP-A was partially dependent on Ca(2+). Interaction of SP-A and Prdx6 at pH 7.4 was shown by Prdx6-mediated inhibition of SP-A binding to agarose beads, a pull-down assay using His-tagged Prdx6 and Ni(2) -chelating beads, co-immunoprecipitation from lung epithelial cells and from a binary mixture of the two proteins, binding after treatment with a trifunctional cross-linker, and size-exclusion chromatography. Analysis by static light scattering and surface plasmon resonance showed calcium-independent SP-A binding to Prdx6 at pH 4.0 and partial Ca(2+) dependence of binding at pH 7.4. These results indicate a direct interaction between SP-A and Prdx6, which provides a mechanism for regulation of the PLA(2) activity of Prdx6 by SP-A.
- Published
- 2006