Receptor-binding domain of ephrin-A1: Production in bacterial expression system and activity
| Autor: | A. N. Wulfson, Oksana V. Nekrasova, Alexey V. Feofanov, Mikhail P. Kirpichnikov, A. I. Tagvey, R. V. Tikhonov, Olga V. Bocharova, S.A. Yakimov, M. V. Astapova, Peter M. Kolosov, A. A. Korchagina, George V. Sharonov |
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| Rok vydání: | 2012 |
| Předmět: |
Gene Expression
Biology Biochemistry EPH receptor B2 EPH receptor A3 Escherichia coli Humans Ephrin Cloning Molecular Phosphorylation Receptor Receptors Eph Family Receptor EphA2 Erythropoietin-producing hepatocellular (Eph) receptor Water Ephrin-A1 General Medicine Ligand (biochemistry) EPH receptor A2 biological factors Protein Structure Tertiary HEK293 Cells Solubility MCF-7 Cells Genetic Engineering |
| Zdroj: | Biochemistry (Moscow). 77:1387-1394 |
| ISSN: | 1608-3040 0006-2979 |
| Popis: | Eph receptor tyrosine kinases and their ligands, the ephrins, perform an important regulatory function in tissue organization, as well as participate in malignant transformation of cells. Ephrin-A1, a ligand of A class Eph receptors, is a modulator of tumor growth and progression, and the mechanism of its action needs detailed investigation. Here we report on the development of a system for bacterial expression of an ephrin-A1 receptor-binding domain (eA1), a procedure for its purification, and its renaturation with final yield of 50 mg/liter of culture. Functional activity of eA1 was confirmed by immunoblotting, laser scanning confocal microscopy, and flow cytometry. It is shown that monomeric non-glycosylated receptor-binding domain of ephrin-A1 is able to activate cellular EphA2 receptors, stimulating their phosphorylation. Ligand eA1 can be used to study the features of ephrin-A1 interactions with different A class Eph receptors. The created expression cassette is suitable for the development of ligands with increased activity and selectivity and experimental systems for the delivery of cytotoxins into tumor cells that overexpress EphA2 or other class A Eph receptors. |
| Databáze: | OpenAIRE |
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