Rapid and Efficient Purification of Functional Collectin-12 and Its Opsonic Activity against Fungal Pathogens
Collectin-12 (collectin placenta 1, CL-P1, or CL-12) is a newly identified pattern recognition molecule of the innate immune system. Recent evidences show that CL-12 plays important roles not only in innate immune protection against certain clinically important pathogens but also in scavenging of ho...
Saved in:
| Main Authors: | , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Wiley
2019-01-01
|
| Series: | Journal of Immunology Research |
| Online Access: | http://dx.doi.org/10.1155/2019/9164202 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1832550092799737856 |
|---|---|
| author | Jie Zhang Anna Li Chang-qing Yang Peter Garred Ying Jie Ma |
| author_facet | Jie Zhang Anna Li Chang-qing Yang Peter Garred Ying Jie Ma |
| author_sort | Jie Zhang |
| collection | DOAJ |
| description | Collectin-12 (collectin placenta 1, CL-P1, or CL-12) is a newly identified pattern recognition molecule of the innate immune system. Recent evidences show that CL-12 plays important roles not only in innate immune protection against certain clinically important pathogens but also in scavenging of host molecules, leukocyte recruitment, and cancer metastasis. Furthermore, CL-12 has been shown to be associated with the pathogenesis of human diseases such as Alzheimer’s disease and multiple sclerosis lesion development. Therefore, the functional consequence of CL-12 remains intriguing and awaits further elucidation. However, available protocols for the purification of recombinant CL-12 with high purity are laborious and inefficient and hamper further functional studies. Here, we report a simple, rapid, and efficient solution to obtain biologically active CL-12 with high purity. We established stable transfected Flp-In™-CHO cells expressing the recombinant CL-12 extracellular domain in high amounts. Recombinant CL-12 was purified from cell culture supernatants using a 3-step rapid purification procedure utilizing disposable affinity and ion exchange minicolumns. Purified recombinant CL-12 adopted an oligomeric structure with monomers, dimers, and trimers and retained its binding capacity towards the A. fumigatus strain that has been described before. Furthermore, we demonstrated the opsonic properties towards eight clinical isolates of A. fumigatus strains and diverse clinically important fungal pathogens. Purified recombinant CL-12 revealed a differential binding capacity towards selected fungal pathogens in vitro. In conclusion, we demonstrate a rapid and efficient purification solution for further biochemical and functional characterization of CL-12 and reveal opsonic properties of CL-12 towards diverse fungal pathogens. |
| format | Article |
| id | doaj-art-af4d2130930d4fc2839cbf7b279bdb95 |
| institution | Kabale University |
| issn | 2314-8861 2314-7156 |
| language | English |
| publishDate | 2019-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | Journal of Immunology Research |
| spelling | doaj-art-af4d2130930d4fc2839cbf7b279bdb952025-02-03T06:07:49ZengWileyJournal of Immunology Research2314-88612314-71562019-01-01201910.1155/2019/91642029164202Rapid and Efficient Purification of Functional Collectin-12 and Its Opsonic Activity against Fungal PathogensJie Zhang0Anna Li1Chang-qing Yang2Peter Garred3Ying Jie Ma4The Laboratory of Molecular Medicine, Department of Clinical Immunology, Section 7631, Rigshospitalet, Faculty of Health and Medical Sciences, University of Copenhagen, Ole Maaloesvej 26, 2200 Copenhagen N, DenmarkThe Laboratory of Molecular Medicine, Department of Clinical Immunology, Section 7631, Rigshospitalet, Faculty of Health and Medical Sciences, University of Copenhagen, Ole Maaloesvej 26, 2200 Copenhagen N, DenmarkDepartment of Clinical Pharmacy, School of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, 211198 Nanjing, ChinaThe Laboratory of Molecular Medicine, Department of Clinical Immunology, Section 7631, Rigshospitalet, Faculty of Health and Medical Sciences, University of Copenhagen, Ole Maaloesvej 26, 2200 Copenhagen N, DenmarkThe Laboratory of Molecular Medicine, Department of Clinical Immunology, Section 7631, Rigshospitalet, Faculty of Health and Medical Sciences, University of Copenhagen, Ole Maaloesvej 26, 2200 Copenhagen N, DenmarkCollectin-12 (collectin placenta 1, CL-P1, or CL-12) is a newly identified pattern recognition molecule of the innate immune system. Recent evidences show that CL-12 plays important roles not only in innate immune protection against certain clinically important pathogens but also in scavenging of host molecules, leukocyte recruitment, and cancer metastasis. Furthermore, CL-12 has been shown to be associated with the pathogenesis of human diseases such as Alzheimer’s disease and multiple sclerosis lesion development. Therefore, the functional consequence of CL-12 remains intriguing and awaits further elucidation. However, available protocols for the purification of recombinant CL-12 with high purity are laborious and inefficient and hamper further functional studies. Here, we report a simple, rapid, and efficient solution to obtain biologically active CL-12 with high purity. We established stable transfected Flp-In™-CHO cells expressing the recombinant CL-12 extracellular domain in high amounts. Recombinant CL-12 was purified from cell culture supernatants using a 3-step rapid purification procedure utilizing disposable affinity and ion exchange minicolumns. Purified recombinant CL-12 adopted an oligomeric structure with monomers, dimers, and trimers and retained its binding capacity towards the A. fumigatus strain that has been described before. Furthermore, we demonstrated the opsonic properties towards eight clinical isolates of A. fumigatus strains and diverse clinically important fungal pathogens. Purified recombinant CL-12 revealed a differential binding capacity towards selected fungal pathogens in vitro. In conclusion, we demonstrate a rapid and efficient purification solution for further biochemical and functional characterization of CL-12 and reveal opsonic properties of CL-12 towards diverse fungal pathogens.http://dx.doi.org/10.1155/2019/9164202 |
| spellingShingle | Jie Zhang Anna Li Chang-qing Yang Peter Garred Ying Jie Ma Rapid and Efficient Purification of Functional Collectin-12 and Its Opsonic Activity against Fungal Pathogens Journal of Immunology Research |
| title | Rapid and Efficient Purification of Functional Collectin-12 and Its Opsonic Activity against Fungal Pathogens |
| title_full | Rapid and Efficient Purification of Functional Collectin-12 and Its Opsonic Activity against Fungal Pathogens |
| title_fullStr | Rapid and Efficient Purification of Functional Collectin-12 and Its Opsonic Activity against Fungal Pathogens |
| title_full_unstemmed | Rapid and Efficient Purification of Functional Collectin-12 and Its Opsonic Activity against Fungal Pathogens |
| title_short | Rapid and Efficient Purification of Functional Collectin-12 and Its Opsonic Activity against Fungal Pathogens |
| title_sort | rapid and efficient purification of functional collectin 12 and its opsonic activity against fungal pathogens |
| url | http://dx.doi.org/10.1155/2019/9164202 |
| work_keys_str_mv | AT jiezhang rapidandefficientpurificationoffunctionalcollectin12anditsopsonicactivityagainstfungalpathogens AT annali rapidandefficientpurificationoffunctionalcollectin12anditsopsonicactivityagainstfungalpathogens AT changqingyang rapidandefficientpurificationoffunctionalcollectin12anditsopsonicactivityagainstfungalpathogens AT petergarred rapidandefficientpurificationoffunctionalcollectin12anditsopsonicactivityagainstfungalpathogens AT yingjiema rapidandefficientpurificationoffunctionalcollectin12anditsopsonicactivityagainstfungalpathogens |