Figure 2.

Spectroscopic studies on heme binding to agonistic IL-36 family members and IL-36α protein mutants. (A) SPR signal (RU) of trIL-36α, β, and γ with five consecutive heme injections of increasing heme concentrations (80 nM to 20 μM). The single-cycle kinetics method was employed (fit is displayed in red). (B) UV/Vis differential spectra of heme-incubated IL-36α and protein mutants. Arrows denote UV/Vis maxima whereas dashed lines at 400 and 500 nm indicate UV/Vis band width in order to illustrate band broadening which was previously found for unspecific heme binding³⁰. (C) Raman spectra of heme (in black), and pentacoordinated wild-type IL-36α (in red) including wavenumber fingerprint region with assignment of prominent normal-mode frequencies ν₇ (681 cm⁻¹), ν₄ (1374 cm⁻¹), ν₃ (1492 cm⁻¹), ν₂ (1571 cm⁻¹), and ν₁₀ (1628 cm⁻¹) for heme.