The adsorption of Rb+, Cs+, Mn2+, Co2+ and Yb3+ onto the positively charged hen egg-white lysozyme (HEWL) has been investigated by solving 13 X-ray structures of HEWL crystallized with their chlorides and by applying electrospray ionization mass spectrometry (ESI-MS) first to dissolved protein crystals and then to the protein in buffered salt solutions. The number of bound cations follows the order Cs+ < Mn2+ ≃ Co2+ < Yb3+ at 293 K. HEWL binds less Rb+ (qtot = 0.7) than Cs+ (qtot = 3.9) at 100 K. Crystal flash-cooling drastically increases the binding of Cs+, but poorly affects that of Yb3+, suggesting different interactions. The addition of glycerol increases the number of bound Yb3+ cations, but only slightly increases that of Rb+. HEWL titrations with the same chlorides, followed by ESI-MS analysis, show that only about 10% of HEWL binds Cs+ and about 40% binds 1–2 Yb3+ cations, while the highest binding reaches 60–70% for protein binding 1–3 Mn2+ or Co2+ cations. The binding sites identified by X-ray crystallography show that the monovalent Rb+ and Cs+ preferentially bind to carbonyl groups, whereas the multivalent Mn2+, Co2+ and Yb3+ interact with carboxylic groups. This work elucidates the basis of the effect of the Hofmeister cation series on protein solubility.
Supporting information
PDB references: HEWL in 0.5 M MnCl2, data collected at room temperature, 4neb; HEWL in 1.1 M MnCl2, data collected at room temperature, 4nfv; HEWL in 1.9 M CsCl, data collected at room temperature, 4ng1; dialyzed HEWL batch-crystallized in 1.9 M CsCl, data collected at 100 K, 4ng8; previously deionized HEWL crystallized in 1.0 M RbCl, data collected at 125 K, 4ngi; dialyzed HEWL batch-crystallized in 1.0 M RbCl, data collected at 100 K, 4ngj; previously deionized HEWL batch-crystallized in 0.2 M CoCl2, 4ngk; previously deionized HEWL batch-crystallized in 1.0 M CoCl2, 4ngo; previously deionized HEWL batch-crystallized in 0.6 M CoCl2, 4ngl; previously deionized HEWL batch-crystallized in 0.5 M YbCl3, 4ngv; dialyzed HEWL batch-crystallized in 0.5 M YbCl3, data collected at 100 K, 4ngw; dialyzed HEWL batch-crystallized in 0.75 M YbCl3, data collected at 100 K, 4ngy; previously deionized HEWL crystallized in 0.48 M YbCl3/30%(v/v) glycerol, data collected at 125 K, 4ngz