Two-dimensional crystals of enzyme-effector complexes: ribonucleotide reductase at 18-.ANG. resolution

Abstract

The B1 subunit of ribonucleotide reductase formed two-dimensional cystals when bound to an effector nucleotide linked to lipids in planar layers at the air/water interface. The effector lipid consisted of dATP coupled through the .gamma.-phosphoryl group and an .epsilon.-aminocaproyl linker to phosphatidylethanolamine. Two-dimensional crystals of B1 reductase, like those of antibodies and cholera toxin obtained previously, formed under physiologic conditions of pH and ionic strength, with no precipitant added to the solution. There was, however, a requirement for dTTP in the solution, presumably to ensure binding of the dATP-lipid at only one of two effector sites on the enzyme. Diffraction from the crystals extended to 18-.ANG. resolution in negative stain, with unit cell parameters a = 110 .ANG., b = 227 .ANG., and .gamma. = 90.degree.. Image analysis revealed the B1 dimer as a pair of roughly cylindrical objects, each 105-109 .ANG. in length and 31-34 .ANG. in diameter.