Induced Long-Range Attractive Potentials of Human Serum Albumin by Ligand Binding

Small-angle x-ray scattering and dielectric spectroscopy investigation on the solutions of recombinant human serum albumin and its heme hybrid revealed that heme incorporation induces a specific long-range attractive potential between protein molecules. This is evidenced by the enhanced forward intensity upon heme binding, despite no hindrance to rotatory Brownian motion, unbiased colloid osmotic pressure, and discontiguous nearest-neighbor distance, confirming monodispersity of the proteins. The heme-induced potential may play a trigger role in recognition of the ligand-filled human serum albumins in the circulatory system.

Figure 1

(a) The normalized x-ray scattered intensities, $I(q)/c$, and (b) the pair-distance distribution functions, $p(r)$, of rHSA in $0.15M$ PBS solutions in $3.0\le c/\mathrm{mg}{\mathrm{ml}}^{-1}\le 50$. The black solid curve shown in (b) represents $p(r)$ calculated from the crystallography data on HSA [3].

Figure 2

(a) Variation of $I(q)$ and (b) the effective structure factors $S^{\mathrm{eff}}(q)$ for rHSA and rHSA-heme in aqueous and $0.15M$ PBS solutions of a fixed concentration $c=30\mathrm{mg}{\mathrm{ml}}^{-1}$. In the inset, the low-$q$ intensities for $c=3.5\mathrm{mg}{\mathrm{ml}}^{-1}$ and $c=10\mathrm{mg}{\mathrm{ml}}^{-1}$ are compared. Arrows on $S^{\mathrm{eff}}(q)$ highlight the monomer-monomer correlation peak positions $q^{*}$.

Figure 3

Effects of concentration, ionic strength, and heme binding on $S^{\mathrm{eff}}(q)$. (a) rHSA-heme and (b) rHSA in $0.15M$ PBS solutions and (c) rHSA-heme and (d) rHSA in aqueous solutions in $10\le c/\mathrm{mg}{\mathrm{ml}}^{-1}\le 50$ (an increment of $10\mathrm{mg}{\mathrm{ml}}^{-1}$). (e) The protein-protein correlation peak position $q^{*}$ in $S^{\mathrm{eff}}(q)$ for aqueous rHSA (□) and rHSA-heme (○) as a function of $c$.

Figure 4

Complex dielectric spectra of aqueous solutions of (a) rHSA and (b) rHSA-heme in $10\le c/\mathrm{mg}{\mathrm{ml}}^{-1}\le 50$ (an increment of $10\mathrm{mg}{\mathrm{ml}}^{-1}$ from the bottom) at $25°\mathrm{C}$.