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The α5β1 integrin heterodimer is involved in many cellular processes and is an anti-cancer therapeutic target. Therefore, access to quantities of protein suitable for studies aimed at understanding its biological functions is important. To this end, a large-scale protein expression system, utilizing the recombinant baculovirus/SF9 insect cell expression system, was created to produce the extracellular domain of the α5β1 integrin. An incorporated 8X-histidine tag enabled one-step nickel-column purification. Following sequence confirmation by LC-MS/MS, the conformation of the heterodimer was characterized by native dot blot and negative stain electron microscopy. Cellular transduction inhibition studies confirmed biological activity. The system allows expression and purification of α5β1 integrin in quantities suitable for an array of different experiments including structural biology., (Copyright © 2013 Elsevier Inc. All rights reserved.)

A recombinant integrin expression system has been created for the large-scale production of αVβ5 integrin extracellular domains that take advantage of Fos and Jun dimerization for expression in bacterial, insect, and mammalian cells. This utilizes an all-in-one vector, pQE-TriSystem, with molecular machinery for parallel expression without the need of additional subcloning. Optimal expression in HEK293 cells was determined by a time course analysis. The heterodimer was purified in a one-step nickel column purification scheme, and the sequence and functional state were confirmed by mass spectrometry and inhibition assays, respectively. The yields of αVβ5 integrin obtained are in quantities suitable for multiple applications including structural biology and functional assays., (Copyright © 2013 Elsevier Inc. All rights reserved.)