Ca<sup>2+</sup>-regulated Photoprotein Obelin as N-terminal Partner in the Fusion Proteins
Ca<sup>2+</sup>-regulated photoproteins genetically fused with biospecific polipeptides have been extensively used in intracellular Ca<sup>2+</sup> measurements and in the development of binding assays. Gene fusions to aequorin have been limited to its N-terminus, since as pr...
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| Format: | Article |
| Language: | English |
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Siberian Federal University
2010-12-01
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| Series: | Журнал Сибирского федерального университета: Серия Биология |
| Subjects: | |
| Online Access: | http://elib.sfu-kras.ru/bitstream/2311/2248/1/Eremeeva.pdf |
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| author | Elena V. Eremeeva Ludmila A. Frank Svetlana V. Markova Eugene Vysotski |
| author_facet | Elena V. Eremeeva Ludmila A. Frank Svetlana V. Markova Eugene Vysotski |
| author_sort | Elena V. Eremeeva |
| collection | DOAJ |
| description | Ca<sup>2+</sup>-regulated photoproteins genetically fused with biospecific polipeptides have been extensively used in intracellular Ca<sup>2+</sup> measurements and in the development of binding assays. Gene fusions to aequorin have been limited to its N-terminus, since as previous studies indicated the protein loses bioluminescent activity upon modification of its C-terminus. To investigate this in regard to another photoprotein - obelin (OL) the one was elongated at its C-terminus with tyrosine (Y) and then fused with green fluorescent protein Clytia gregaria (cgreGFP) through the flexible 31 aa linker. Both proteins (OL-Y and OL-cgreGFP) were isolated and investigated. The OL-Y was found to form a stable photoprotein comlex, possessing 75 % of WT-OL bioluminescent activity. OL-cgreGFP activity preserves 46 % of WT-OL activity and demonstrates an effective resonance energy transfer, where OL-partner and cgreGFP-partner are energy donor and acceptor respectively. Thus, it was shown that the labels on the base of Ca<sup>2+</sup>-regulated photoprotein obelin may be obtained by fusing with biospecific polypeptides regardless its termini. |
| first_indexed | 2024-04-13T06:35:53Z |
| format | Article |
| id | doaj.art-bc63db1b86574fb8979a632fd213b929 |
| institution | Directory Open Access Journal |
| issn | 1997-1389 2313-5530 |
| language | English |
| last_indexed | 2024-04-13T06:35:53Z |
| publishDate | 2010-12-01 |
| publisher | Siberian Federal University |
| record_format | Article |
| series | Журнал Сибирского федерального университета: Серия Биология |
| spelling | doaj.art-bc63db1b86574fb8979a632fd213b9292022-12-22T02:57:56ZengSiberian Federal UniversityЖурнал Сибирского федерального университета: Серия Биология1997-13892313-55302010-12-0134372383http://elib.sfu-kras.ru/handle/2311/2248Ca<sup>2+</sup>-regulated Photoprotein Obelin as N-terminal Partner in the Fusion ProteinsElena V. Eremeeva0Ludmila A. Frank1Svetlana V. Markova2Eugene Vysotski3Institute of Biophysics SB RAS , Russia, 660036, Krasnoyarsk, AkademgorodokInstitute of Biophysics SB RAS Siberian Federal University , Russia, 660036, Krasnoyarsk, Akademgorodok Russia, 660041, Krasnoyarsk, Svobodny av., 79Institute of Biophysics SB RAS Siberian Federal University , Russia, 660036, Krasnoyarsk, Akademgorodok Russia, 660041, Krasnoyarsk, Svobodny av., 79Institute of Biophysics SB RAS , Russia, 660036, Krasnoyarsk, AkademgorodokCa<sup>2+</sup>-regulated photoproteins genetically fused with biospecific polipeptides have been extensively used in intracellular Ca<sup>2+</sup> measurements and in the development of binding assays. Gene fusions to aequorin have been limited to its N-terminus, since as previous studies indicated the protein loses bioluminescent activity upon modification of its C-terminus. To investigate this in regard to another photoprotein - obelin (OL) the one was elongated at its C-terminus with tyrosine (Y) and then fused with green fluorescent protein Clytia gregaria (cgreGFP) through the flexible 31 aa linker. Both proteins (OL-Y and OL-cgreGFP) were isolated and investigated. The OL-Y was found to form a stable photoprotein comlex, possessing 75 % of WT-OL bioluminescent activity. OL-cgreGFP activity preserves 46 % of WT-OL activity and demonstrates an effective resonance energy transfer, where OL-partner and cgreGFP-partner are energy donor and acceptor respectively. Thus, it was shown that the labels on the base of Ca<sup>2+</sup>-regulated photoprotein obelin may be obtained by fusing with biospecific polypeptides regardless its termini.http://elib.sfu-kras.ru/bitstream/2311/2248/1/Eremeeva.pdfcgreGFPobelinC-end fusingcgreGFP |
| spellingShingle | Elena V. Eremeeva Ludmila A. Frank Svetlana V. Markova Eugene Vysotski Ca<sup>2+</sup>-regulated Photoprotein Obelin as N-terminal Partner in the Fusion Proteins Журнал Сибирского федерального университета: Серия Биология cgreGFP obelin C-end fusing cgreGFP |
| title | Ca<sup>2+</sup>-regulated Photoprotein Obelin as N-terminal Partner in the Fusion Proteins |
| title_full | Ca<sup>2+</sup>-regulated Photoprotein Obelin as N-terminal Partner in the Fusion Proteins |
| title_fullStr | Ca<sup>2+</sup>-regulated Photoprotein Obelin as N-terminal Partner in the Fusion Proteins |
| title_full_unstemmed | Ca<sup>2+</sup>-regulated Photoprotein Obelin as N-terminal Partner in the Fusion Proteins |
| title_short | Ca<sup>2+</sup>-regulated Photoprotein Obelin as N-terminal Partner in the Fusion Proteins |
| title_sort | ca sup 2 sup regulated photoprotein obelin as n terminal partner in the fusion proteins |
| topic | cgreGFP obelin C-end fusing cgreGFP |
| url | http://elib.sfu-kras.ru/bitstream/2311/2248/1/Eremeeva.pdf |
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