Three-dimensional structure of the catalytic domain of chitinase A1 from Bacillus circulans WL-12 at a very high resolution

The mature form of chitinase A1 from Bacillus circulans WL-12 (one of the family 18 glycosyl hydrolases) comprises an N-terminal catalytic domain, two fibronectin type III domains and a C-terminal chitin-binding domain. In the efforts for crystallizing each domain, combined domains and the intact mature form, we have obtained crystals of the catalytic domain which diffract to a truly atomic resolution (1.13 Å). The catalytic domain consists of an (α/β)₈-TIM-barrel. Two small β-domains attached on top of the TIM-barrel provide a deep cleft for substrate binding. Crystals of an inactivated (through Glu204Gln mutation) catalytic domain complexed with a heptameric N-acetyl glucosamine (7NAG (N^I, N^II, N^III, N^IV, N^V, N^VI, N^VII-hepta-acetyl-chitoheptaose which is a β-1, 4-linked oligosaccharide of N-acetylglucosamine with a polymerization degree of seven.), which is a true substrate) diffracted up to 1.5 Å resolution. The bound 7NAG substrate showed a marked kink between the sugar units at positions -1 and +1. The proximity of Glu204 to the scissile site strongly supports the hitherto conceived notion that Glu204 plays a role of catalytic acid in the so-called "substrate-assisted catalysis" mechanism which is distinct from the general acid-base mechanism long accepted for lysozyme and related glycosyl hydrolases.