11] Two-dimensional protein crystals in aid of three-dimensional protein crystal growth

Abstract

Publisher Summary The possibility of two-dimensional crystals serving as seeds for three-dimensional crystal growth is suggested by electron micrographs showing multilayered crystals of antihaptenic antibodies formed on lipid hapten and RNA polymerase II formed on charged lipids. This apparent tendency toward epitaxial crystal growth with dilute protein solutions (50–100 μg/mL) in the absence of a precipitant was pursued by the use of conditions more conducive to three-dimensional crystallization. The approach has so far been tried and has proved successful with three protein and lipid combinations, streptavidin with biotinylated lipids, RNA polymerase II with charged lipids, and the Epstein–Barr virus nuclear antigen-1 (EBNA1) with charged lipids. In the case of streptavidin, a thorough study was performed, thereby demonstrating the formation of three-dimensional crystals of a size and quality suitable for X-ray diffraction analysis and establishing that these crystals arose by epitaxial growth from two-dimensional crystals. Such epitaxial growth occurred at very low protein concentrations and yielded three-dimensional crystals in space groups corresponding to those of the two-dimensional crystals from which they were derived rather than those normally obtained under the ionic conditions used. Three procedures for epitaxial growth of three-dimensional crystals are described in this chapter: the first illustrates the use of lipid layers without preformed two-dimensional crystals in the conventional hanging drop procedure and the second and third procedures utilize preformed two-dimensional crystals to nucleate three-dimensional crystal growth.