Single-Step Purification of Monomeric l-Selectin via Aptamer Affinity Chromatography
l-selectin is a transmembrane receptor expressed on the surface of white blood cells and responsible for the tethering of leukocytes to vascular endothelial cells. This initial intercellular contact is the first step of the complex leukocyte adhesion cascade that ultimately permits extravasation of...
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| Format: | Article |
| Language: | English |
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MDPI AG
2017-01-01
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| Series: | Sensors |
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| Online Access: | http://www.mdpi.com/1424-8220/17/2/226 |
| _version_ | 1798041419653316608 |
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| author | Christian Kuehne Stefanie Wedepohl Jens Dernedde |
| author_facet | Christian Kuehne Stefanie Wedepohl Jens Dernedde |
| author_sort | Christian Kuehne |
| collection | DOAJ |
| description | l-selectin is a transmembrane receptor expressed on the surface of white blood cells and responsible for the tethering of leukocytes to vascular endothelial cells. This initial intercellular contact is the first step of the complex leukocyte adhesion cascade that ultimately permits extravasation of leukocytes into the surrounding tissue in case of inflammation. Here we show the binding of a soluble histidine tagged l-selectin to a recently described shortened variant of an l-selectin specific DNA aptamer with surface plasmon resonance. The high specificity of this aptamer in combination with its high binding affinity of ~12 nM, allows for a single-step protein purification from cell culture supernatants. In comparison to the well-established Ni-NTA based technology, aptamer affinity chromatography (AAC) was easier to establish, resulted in a 3.6-fold higher protein yield, and increased protein purity. Moreover, due to target specificity, the DNA aptamer facilitated binding studies directly from cell culture supernatant, a helpful characteristic to quickly monitor successful expression of biological active l-selectin. |
| first_indexed | 2024-04-11T22:21:17Z |
| format | Article |
| id | doaj.art-b51a468365474a9c810951ff201448a2 |
| institution | Directory Open Access Journal |
| issn | 1424-8220 |
| language | English |
| last_indexed | 2024-04-11T22:21:17Z |
| publishDate | 2017-01-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Sensors |
| spelling | doaj.art-b51a468365474a9c810951ff201448a22022-12-22T04:00:06ZengMDPI AGSensors1424-82202017-01-0117222610.3390/s17020226s17020226Single-Step Purification of Monomeric l-Selectin via Aptamer Affinity ChromatographyChristian Kuehne0Stefanie Wedepohl1Jens Dernedde2Institute of Laboratory Medicine, Clinical Chemistry and Pathobiochemistry, Charité-Universitätsmedizin Berlin, 13353 Berlin, GermanyInstitute of Chemistry and Biochemistry, Freie Universität Berlin, 14195 Berlin, GermanyInstitute of Laboratory Medicine, Clinical Chemistry and Pathobiochemistry, Charité-Universitätsmedizin Berlin, 13353 Berlin, Germanyl-selectin is a transmembrane receptor expressed on the surface of white blood cells and responsible for the tethering of leukocytes to vascular endothelial cells. This initial intercellular contact is the first step of the complex leukocyte adhesion cascade that ultimately permits extravasation of leukocytes into the surrounding tissue in case of inflammation. Here we show the binding of a soluble histidine tagged l-selectin to a recently described shortened variant of an l-selectin specific DNA aptamer with surface plasmon resonance. The high specificity of this aptamer in combination with its high binding affinity of ~12 nM, allows for a single-step protein purification from cell culture supernatants. In comparison to the well-established Ni-NTA based technology, aptamer affinity chromatography (AAC) was easier to establish, resulted in a 3.6-fold higher protein yield, and increased protein purity. Moreover, due to target specificity, the DNA aptamer facilitated binding studies directly from cell culture supernatant, a helpful characteristic to quickly monitor successful expression of biological active l-selectin.http://www.mdpi.com/1424-8220/17/2/226">l-selectinaptamerDNArecombinant proteinpurificationaffinitySPR |
| spellingShingle | Christian Kuehne Stefanie Wedepohl Jens Dernedde Single-Step Purification of Monomeric l-Selectin via Aptamer Affinity Chromatography Sensors ">l-selectin aptamer DNA recombinant protein purification affinity SPR |
| title | Single-Step Purification of Monomeric l-Selectin via Aptamer Affinity Chromatography |
| title_full | Single-Step Purification of Monomeric l-Selectin via Aptamer Affinity Chromatography |
| title_fullStr | Single-Step Purification of Monomeric l-Selectin via Aptamer Affinity Chromatography |
| title_full_unstemmed | Single-Step Purification of Monomeric l-Selectin via Aptamer Affinity Chromatography |
| title_short | Single-Step Purification of Monomeric l-Selectin via Aptamer Affinity Chromatography |
| title_sort | single step purification of monomeric l selectin via aptamer affinity chromatography |
| topic | ">l-selectin aptamer DNA recombinant protein purification affinity SPR |
| url | http://www.mdpi.com/1424-8220/17/2/226 |
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