Conformation and side chains environments of recombinant human interleukin-1 receptor antagonist (rh-IL-1ra) probed by raman, raman optical activity, and UV-resonance Raman spectroscopy.

The conformation and local environments of the side chains cysteines and aromatics of recombinant human interleukin-1 receptor antagonist (rh-IL-1ra) have been studied by visible Raman, Raman optical activity (ROA) and UVRR spectroscopy. The results reveal that the secondary structure of rh-IL-1ra is predominantly beta-sheet, which is consistent with conclusions from multinuclear NMR in solutions and X-ray diffraction analysis of crystals. It confirms that all four cysteines are in reduced state. Three cysteines are not hydrogen bonded and exposed. One cysteine is moderately hydrogen bonded and buried. This explains the earlier observation that only three cysteines were detectable using DTNB titration. No characteristic Raman band of disulfide bond was observed in the Raman spectra of rh-IL-1ra in both solution and in crystals. It rules out the supposition that there is one disulfide bond in rh-IL-1ra crystals based on X-ray diffraction. Raman and UVRR spectra of rh-IL-1ra exhibit canonical marker bands of tryptophan. They do not support the proposal that there is cation-pi interaction involving tryptophans in solutions and crystals. These results demonstrate that Raman spectroscopy offers certain advantages over X-ray diffraction for studies of detailed local environment and intermolecular interactions of side chains of proteins in solution and in crystals.