The 6-Å Crystal Structure of Delta5-3-Ketosteroid Isomerase. Architecture and Location of the Active Center

E. M. WESTBROOK; O. E. PIRO; P. B. SIGLER

JOURNAL OF BIOLOGICAL CHEMISTRY

American Society for Biochemistry and Molecular Biology

Año: 1984 vol. 259 p. 9096 - 9103

0021-9258

The crystal structure of the dimeric steroid metabolizing enzyme, delta 5-3-ketosteroid isomerase (EC 5.3.3.1), has been solved to 6-A resolution by multiple isomorphous replacement, augmented by real space direct methods. The unit cell is hexagonal (space group P6122) with dimensions a = b = 65.4 A, c = 504 A, and contains four identical 13,400-dalton protomers in each of its 12 asymmetric units. The 504-A c axis required double focusing mirrors (Franks optics) to resolve the reflections. The complexity of the combined local and lattice symmetry necessitated direct methods to establish the positions of heavy atoms in even the simplest of the isomorphous derivatives. The electron density map clearly showed both (a) the elaborate packing scheme of protomers, which accounts for this large and complicated unit cell, and (b) the coarse features of the functional dimer. The steroid-binding site has been established by imaging the bound inhibitor, 4- acetoxymercuriestradiol, in a difference Fourier map. Each of the dimer´s two steroid-binding sites lies completely within one subunit but close enough to the opposing subunit that functional interactions may be possible.