An SPR‐based fragment mapping study on interleukin 2 (IL‐2) towards the development of small molecule inhibitors of protein‐protein interactions (PPIs)
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E-Artikel
Titel:
An SPR‐based fragment mapping study on interleukin 2 (IL‐2) towards the development of small molecule inhibitors of protein‐protein interactions (PPIs)
Beschreibung:
<jats:p>To better understand how to inhibit PPI targets with small molecules, we have developed a fragment‐based method for probing the IL‐2/IL‐2 receptor alpha (IL‐2Rα) interface using surface plasmon resonance (SPR) spectroscopy. Previous SPR fragment‐binding studies used IL‐2 randomly coupled to the sensor surface. In our hands random coupling resulted in partial loss of IL‐2 activity, as measured by its ability to bind IL2Rα. We therefore prepared an IL‐2 mutant containing Ser‐Gly‐Cys at the C‐terminus to enable uniform capture of functional IL‐2 through thiol chemistry. The identity of the purified Cys‐IL‐2 was confirmed by MALDI‐TOF MS, and the absence of covalent or noncovalent aggregates was shown by gel filtration chromatography. Proper folding and functional activity were established by showing that the Cys‐IL‐2 was able to bind soluble IL‐2Rα comparably to wild‐type IL‐2, as measured by Biacore and by ELISA. The Cys‐IL‐2 is being used to measure binding of fragments derived from the 60 nM IL‐2 inhibitor reported by Sunesis Pharmaceuticals, to map how binding energy is generated by interaction with each portion of the compound in this PPI inhibitor complex. This research was supported by GM064700‐07S1 from NIGMS.</jats:p>