Molecular cloning, overexpression and characterization of human interleukin 1α

Interleukin-1 alpha (IL-1α) regulates a wide range of important cellular processes. In this study for the first time, we report the cloning, expression, biophysical, and biological characterization of the human interleukin-1α. Human IL-1α has been expressed in Escherichia coli in high yields (∼4 mg per liter of the bacterial culture). The protein was purified to homogeneity (∼98% purity) using affinity chromatography and size exclusion chromatography. Results of the steady-state fluorescence and 2D NMR experiments show that the recombinant IL-1α is in a folded conformation. Far-UV circular dichroism (CD) data suggest that IL-1α is an all β-sheet protein with a β-barrel architecture. Isothermal titration calorimetry (ITC) experiments show that the recombinant IL-1α binds strongly (Kd ∼ 5.6 × 10−7 M) to S100A13, a calcium binding protein that chaperones the in vivo release of IL-1α into the extracellular compartment. Recombinant IL-1α was observed to exhibit strong cytostatic effect on human umbilical vascular endothelial cells. The findings of the present study not only pave way for an in-depth structural investigation of the molecular mechanism(s) underlying the non-classical release of IL-1α but also provide avenues for the rational design of potent inhibitors against IL-1α mediated pathogenesis.