Construction, expression, and purification of recombinant αVß5 integrin.

ABSTRACT

A recombinant integrin expression system has been created for the large-scale production of αVß5 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 αVß5 integrin obtained are in quantities suitable for multiple applications including structural biology and functional assays.