Crystallization and preliminary X-ray diffraction analysis of the 1,3-1,4-beta-D-glucanase from Fibrobacter succinogenes

The truncated 1,3-1,4-beta -glucanase (1,3-1,4-beta -D-glucan 4-glucanohydr olase; E.C. 3.2.1.73) from Fibrobacter succinogenes was crystallized in fou r different forms by the vapour-diffusion method. Form A crystals have the largest trigonal P321 unit cell, diffracting to 3.0 Angstrom resolution wit h four to six molecules per asymmetric unit. Form B and C crystals belong t o the same monoclinic space group P2(1), but the form B unit cell is twice as large as the unit cell of form C. Form B crystals diffract to 2.5 Angstr om resolution and contain four molecules per asymmetric unit. Form C crysta ls diffract to 2.1 Angstrom resolution and contain two molecules per asymme tric unit. Form D crystals have the smallest orthorhombic P2(1)2(1)2(1) uni t cell, containing only one molecule per asymmetric unit, and diffract beyo nd 2.1 Angstrom resolution. The crystallization conditions for form B and C crystals are almost identical, except that form C crystals were grown in t he presence of 2 mM Ca2+ ions. It is likely that Ca2+ directly binds to the glucanase, leading to unit-cell shrinkage as observed in other Bacillus gl ucanase crystals. A self-rotation search identified non-crystallographic tw ofold axes that combine with the crystallographic twofold dyads to give 222 symmetry for both form A and form B crystals, indicating that the glucanas e has a tendency to pack in 222 symmetry.