INFLUENCE OF DIVALENT-CATIONS IN PROTEIN CRYSTALLIZATION

We have tested the effect of several cations in attempts to crystalliz e the ligand-bound forms of the leucine/isoleucine/valine-binding prot ein (LIVBP) (M(r) = 36,700) and leucine-specific binding protein (LBP) (M(r) = 37,000), which act as initial periplasmic receptors for the h igh-affinity osmotic-shock-sensitive active transport system in bacter ial cells. Success was achieved with Cd2+ promoting the most dramatic improvement in crystal size, morphology, and diffraction quality. This comes about 15 years after the ligand-free proteins were crystallized . Nine other different divalent cations were tried as additives in the crystallization of LIVBP with polyethylene glycol 8000 as precipitant , and each showed different effects on the crystal quality and morphol ogy. Cd2+ produced large hexagonal prism crystals of LIVBP, whereas a majority of the cations resulted in less desirable needle-shaped cryst als. Zn2+ gave crystals that are long rods with hexagonal cross sectio ns, a shape intermediate between the hexagonal prism and needle forms. The concentration of Cd2+ is critical. The best crystals of the LIVBP were obtained in the presence of 1 mM CdCl2, whereas those of LBP, wi th trigonal prism morphology, were obtained at a much higher concentra tion of 100 mM. Both crystals diffract to at least 1.7 Angstrom resolu tion using a conventional X-ray source.