Structural and Functional Characterization of Recombinant Interleukin-10 from Indian Major Carp Labeo rohita
Interleukin-10, an important regulator of both the innate and adaptive immune systems, is a multifunctional major cytokine. Though it is one of the major cytokines, IL-10 from the Indian major carp, Labeo rohita, has not yet been characterized. In the present study, we report large scale production...
Saved in:
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Wiley
2016-01-01
|
| Series: | Journal of Immunology Research |
| Online Access: | http://dx.doi.org/10.1155/2016/3962596 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1832547541714993152 |
|---|---|
| author | Sweta Karan Pujarini Dash Himani Kaushik Pramoda K. Sahoo Lalit C. Garg Aparna Dixit |
| author_facet | Sweta Karan Pujarini Dash Himani Kaushik Pramoda K. Sahoo Lalit C. Garg Aparna Dixit |
| author_sort | Sweta Karan |
| collection | DOAJ |
| description | Interleukin-10, an important regulator of both the innate and adaptive immune systems, is a multifunctional major cytokine. Though it is one of the major cytokines, IL-10 from the Indian major carp, Labeo rohita, has not yet been characterized. In the present study, we report large scale production and purification of biologically active recombinant IL-10 of L. rohita (rLrIL-10) using a heterologous expression system and its biophysical and functional characterization. High yield (~70 mg/L) of soluble rLrIL-10 was obtained at shake flask level. The rLrIL-10 was found to exist as a dimer. Far-UV CD spectroscopy showed presence of predominantly alpha helices. The tertiary structure of the purified rLrIL-10 was verified by fluorescence spectroscopy. Two-dimensional gel analysis revealed the presence of six isoforms of the rLrIL-10. The rLrIL-10 was biologically active and its administration significantly reduced serum proinflammatory cytokines, namely, interleukin 1β, TNFα, and IL-8, and augmented the NKEF transcript levels in spleen of L. rohita. Anti-inflammatory role of the rLrIL-10 was further established by inhibition of phagocytosis using NBT reduction assay in vitro. The data indicate that the dimeric alpha helical structure and function of IL-10 of L. rohita as a key regulator of anti-inflammatory response have remained conserved during evolution. |
| format | Article |
| id | doaj-art-fd446bbcbc694d5d8e3b48e725b7effc |
| institution | Kabale University |
| issn | 2314-8861 2314-7156 |
| language | English |
| publishDate | 2016-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | Journal of Immunology Research |
| spelling | doaj-art-fd446bbcbc694d5d8e3b48e725b7effc2025-02-03T06:44:25ZengWileyJournal of Immunology Research2314-88612314-71562016-01-01201610.1155/2016/39625963962596Structural and Functional Characterization of Recombinant Interleukin-10 from Indian Major Carp Labeo rohitaSweta Karan0Pujarini Dash1Himani Kaushik2Pramoda K. Sahoo3Lalit C. Garg4Aparna Dixit5Gene Regulation Laboratory, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, IndiaFish Health Management Division, Central Institute of Freshwater Aquaculture, Kausalyaganga, Bhubaneswar 751 002, IndiaGene Regulation Laboratory, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, IndiaFish Health Management Division, Central Institute of Freshwater Aquaculture, Kausalyaganga, Bhubaneswar 751 002, IndiaGene Regulation Laboratory, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, IndiaSchool of Biotechnology, Jawaharlal Nehru University, New Delhi 110067, IndiaInterleukin-10, an important regulator of both the innate and adaptive immune systems, is a multifunctional major cytokine. Though it is one of the major cytokines, IL-10 from the Indian major carp, Labeo rohita, has not yet been characterized. In the present study, we report large scale production and purification of biologically active recombinant IL-10 of L. rohita (rLrIL-10) using a heterologous expression system and its biophysical and functional characterization. High yield (~70 mg/L) of soluble rLrIL-10 was obtained at shake flask level. The rLrIL-10 was found to exist as a dimer. Far-UV CD spectroscopy showed presence of predominantly alpha helices. The tertiary structure of the purified rLrIL-10 was verified by fluorescence spectroscopy. Two-dimensional gel analysis revealed the presence of six isoforms of the rLrIL-10. The rLrIL-10 was biologically active and its administration significantly reduced serum proinflammatory cytokines, namely, interleukin 1β, TNFα, and IL-8, and augmented the NKEF transcript levels in spleen of L. rohita. Anti-inflammatory role of the rLrIL-10 was further established by inhibition of phagocytosis using NBT reduction assay in vitro. The data indicate that the dimeric alpha helical structure and function of IL-10 of L. rohita as a key regulator of anti-inflammatory response have remained conserved during evolution.http://dx.doi.org/10.1155/2016/3962596 |
| spellingShingle | Sweta Karan Pujarini Dash Himani Kaushik Pramoda K. Sahoo Lalit C. Garg Aparna Dixit Structural and Functional Characterization of Recombinant Interleukin-10 from Indian Major Carp Labeo rohita Journal of Immunology Research |
| title | Structural and Functional Characterization of Recombinant Interleukin-10 from Indian Major Carp Labeo rohita |
| title_full | Structural and Functional Characterization of Recombinant Interleukin-10 from Indian Major Carp Labeo rohita |
| title_fullStr | Structural and Functional Characterization of Recombinant Interleukin-10 from Indian Major Carp Labeo rohita |
| title_full_unstemmed | Structural and Functional Characterization of Recombinant Interleukin-10 from Indian Major Carp Labeo rohita |
| title_short | Structural and Functional Characterization of Recombinant Interleukin-10 from Indian Major Carp Labeo rohita |
| title_sort | structural and functional characterization of recombinant interleukin 10 from indian major carp labeo rohita |
| url | http://dx.doi.org/10.1155/2016/3962596 |
| work_keys_str_mv | AT swetakaran structuralandfunctionalcharacterizationofrecombinantinterleukin10fromindianmajorcarplabeorohita AT pujarinidash structuralandfunctionalcharacterizationofrecombinantinterleukin10fromindianmajorcarplabeorohita AT himanikaushik structuralandfunctionalcharacterizationofrecombinantinterleukin10fromindianmajorcarplabeorohita AT pramodaksahoo structuralandfunctionalcharacterizationofrecombinantinterleukin10fromindianmajorcarplabeorohita AT lalitcgarg structuralandfunctionalcharacterizationofrecombinantinterleukin10fromindianmajorcarplabeorohita AT aparnadixit structuralandfunctionalcharacterizationofrecombinantinterleukin10fromindianmajorcarplabeorohita |