Identification of plasma proteins binding oxidized phospholipids using pull-down proteomics and OxLDL masking assay‡
Oxidized phospholipids (OxPLs) are increasingly recognized as toxic and proinflammatory mediators, which raises interest in the mechanisms of their detoxification. Circulating OxPLs are bound and neutralized by plasma proteins, including both antibodies and non-immunoglobulin proteins. The latter gr...
Saved in:
| Main Authors: | , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Elsevier
2025-01-01
|
| Series: | Journal of Lipid Research |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S0022227524002098 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1832582269975396352 |
|---|---|
| author | Philipp Jokesch Lisa Holzer Lydia Jantscher Sebastian Guttzeit Rudolf Übelhart Olga Oskolkova Valery Bochkov Bernd Gesslbauer |
| author_facet | Philipp Jokesch Lisa Holzer Lydia Jantscher Sebastian Guttzeit Rudolf Übelhart Olga Oskolkova Valery Bochkov Bernd Gesslbauer |
| author_sort | Philipp Jokesch |
| collection | DOAJ |
| description | Oxidized phospholipids (OxPLs) are increasingly recognized as toxic and proinflammatory mediators, which raises interest in the mechanisms of their detoxification. Circulating OxPLs are bound and neutralized by plasma proteins, including both antibodies and non-immunoglobulin proteins. The latter group of proteins is essentially not investigated because only three OxPC-binding plasma proteins are currently known. The goal of this work was to characterize a broad spectrum of plasma proteins selectively binding OxPLs. Using pull-down-proteomic analysis, we found about 150 non-immunoglobulin proteins preferentially binding oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-phosphatidylcholine (OxPAPC) as compared to non-oxidized PAPC. To test if candidate proteins indeed can form a barrier isolating OxPLs from recognition by other proteins, we applied an immune masking assay. Oxidized LDL (OxLDL) immobilized in multiwell plates was used as a carrier of OxPLs, while mAbs recognizing OxPC or OxPE were used as “detectors” showing if OxPLs on the surface of OxLDL are physically accessible to external binding partners. Using an orthogonal combination of pull-down and masking assays we confirmed that previously described OxPL-binding proteins (non-fractionated IgM, CFH, and Apo-M) indeed can bind to and mask OxPC and OxPE on liposomes and OxLDL. Furthermore, we identified additional plasma proteins selectively binding and masking OxPC including Apo-D, Apo-H, pulmonary surfactant-associated protein B, and antithrombin-III. We hypothesize that in addition to circulating antibodies, multiple non-immunoglobulin plasma proteins can also bind OxPLs and modulate their recognition by innate and adaptive immunity. |
| format | Article |
| id | doaj-art-df05f9832d9b4d85ad924110ec2b7ca0 |
| institution | Kabale University |
| issn | 0022-2275 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Lipid Research |
| spelling | doaj-art-df05f9832d9b4d85ad924110ec2b7ca02025-01-30T05:12:36ZengElsevierJournal of Lipid Research0022-22752025-01-01661100704Identification of plasma proteins binding oxidized phospholipids using pull-down proteomics and OxLDL masking assay‡Philipp Jokesch0Lisa Holzer1Lydia Jantscher2Sebastian Guttzeit3Rudolf Übelhart4Olga Oskolkova5Valery Bochkov6Bernd Gesslbauer7Department of Pharmaceutical Chemistry, Institute of Pharmaceutical Sciences, University of Graz, Graz, AustriaDepartment of Pharmaceutical Chemistry, Institute of Pharmaceutical Sciences, University of Graz, Graz, AustriaDepartment of Pharmaceutical Chemistry, Institute of Pharmaceutical Sciences, University of Graz, Graz, AustriaVanudis GmbH, Heidelberg, GermanyVanudis GmbH, Heidelberg, GermanyDepartment of Pharmaceutical Chemistry, Institute of Pharmaceutical Sciences, University of Graz, Graz, AustriaDepartment of Pharmaceutical Chemistry, Institute of Pharmaceutical Sciences, University of Graz, Graz, Austria; Field of Excellence BioHealth - University of Graz, Graz, Austria; For correspondence: Bernd Gesslbauer; Valery BochkovDepartment of Pharmaceutical Chemistry, Institute of Pharmaceutical Sciences, University of Graz, Graz, Austria; For correspondence: Bernd Gesslbauer; Valery BochkovOxidized phospholipids (OxPLs) are increasingly recognized as toxic and proinflammatory mediators, which raises interest in the mechanisms of their detoxification. Circulating OxPLs are bound and neutralized by plasma proteins, including both antibodies and non-immunoglobulin proteins. The latter group of proteins is essentially not investigated because only three OxPC-binding plasma proteins are currently known. The goal of this work was to characterize a broad spectrum of plasma proteins selectively binding OxPLs. Using pull-down-proteomic analysis, we found about 150 non-immunoglobulin proteins preferentially binding oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-phosphatidylcholine (OxPAPC) as compared to non-oxidized PAPC. To test if candidate proteins indeed can form a barrier isolating OxPLs from recognition by other proteins, we applied an immune masking assay. Oxidized LDL (OxLDL) immobilized in multiwell plates was used as a carrier of OxPLs, while mAbs recognizing OxPC or OxPE were used as “detectors” showing if OxPLs on the surface of OxLDL are physically accessible to external binding partners. Using an orthogonal combination of pull-down and masking assays we confirmed that previously described OxPL-binding proteins (non-fractionated IgM, CFH, and Apo-M) indeed can bind to and mask OxPC and OxPE on liposomes and OxLDL. Furthermore, we identified additional plasma proteins selectively binding and masking OxPC including Apo-D, Apo-H, pulmonary surfactant-associated protein B, and antithrombin-III. We hypothesize that in addition to circulating antibodies, multiple non-immunoglobulin plasma proteins can also bind OxPLs and modulate their recognition by innate and adaptive immunity.http://www.sciencedirect.com/science/article/pii/S0022227524002098oxidized lipidsphospholipids/metabolismLDL/Oxidation/antioxidantsphospholipasesantibodiesglycosaminoglycan |
| spellingShingle | Philipp Jokesch Lisa Holzer Lydia Jantscher Sebastian Guttzeit Rudolf Übelhart Olga Oskolkova Valery Bochkov Bernd Gesslbauer Identification of plasma proteins binding oxidized phospholipids using pull-down proteomics and OxLDL masking assay‡ Journal of Lipid Research oxidized lipids phospholipids/metabolism LDL/Oxidation/antioxidants phospholipases antibodies glycosaminoglycan |
| title | Identification of plasma proteins binding oxidized phospholipids using pull-down proteomics and OxLDL masking assay‡ |
| title_full | Identification of plasma proteins binding oxidized phospholipids using pull-down proteomics and OxLDL masking assay‡ |
| title_fullStr | Identification of plasma proteins binding oxidized phospholipids using pull-down proteomics and OxLDL masking assay‡ |
| title_full_unstemmed | Identification of plasma proteins binding oxidized phospholipids using pull-down proteomics and OxLDL masking assay‡ |
| title_short | Identification of plasma proteins binding oxidized phospholipids using pull-down proteomics and OxLDL masking assay‡ |
| title_sort | identification of plasma proteins binding oxidized phospholipids using pull down proteomics and oxldl masking assay‡ |
| topic | oxidized lipids phospholipids/metabolism LDL/Oxidation/antioxidants phospholipases antibodies glycosaminoglycan |
| url | http://www.sciencedirect.com/science/article/pii/S0022227524002098 |
| work_keys_str_mv | AT philippjokesch identificationofplasmaproteinsbindingoxidizedphospholipidsusingpulldownproteomicsandoxldlmaskingassay AT lisaholzer identificationofplasmaproteinsbindingoxidizedphospholipidsusingpulldownproteomicsandoxldlmaskingassay AT lydiajantscher identificationofplasmaproteinsbindingoxidizedphospholipidsusingpulldownproteomicsandoxldlmaskingassay AT sebastianguttzeit identificationofplasmaproteinsbindingoxidizedphospholipidsusingpulldownproteomicsandoxldlmaskingassay AT rudolfubelhart identificationofplasmaproteinsbindingoxidizedphospholipidsusingpulldownproteomicsandoxldlmaskingassay AT olgaoskolkova identificationofplasmaproteinsbindingoxidizedphospholipidsusingpulldownproteomicsandoxldlmaskingassay AT valerybochkov identificationofplasmaproteinsbindingoxidizedphospholipidsusingpulldownproteomicsandoxldlmaskingassay AT berndgesslbauer identificationofplasmaproteinsbindingoxidizedphospholipidsusingpulldownproteomicsandoxldlmaskingassay |