Crystal structure of chondroitin AC lyase, a representative of a family ofglycosaminoglycan degrading enzymes

Glycosaminoglycans (GAGs), highly sulfated polymers built of hexosamine-uro nic acid disaccharide units, are major components of the extracellular matr ix, mostly in the form of proteoglycans. They interact with a large array o f proteins, in particular of the blood coagulation cascade. Degradation of GAGs in mammalian systems occurs by the action of GAG hydrolases. Bacteria express a large number of GAG-degrading lyases that break the hexosamine-ur onic acid bond to create an unsaturated sugar ring. Flavobacterium heparinu m produces at least five GAG lyases of different specificity. Chondroitin A C lyase (chondroitinase AC, 75 kDa) is highly active toward chondroitin 4-s ulfate and chondroitin-6 sulfate. Its crystal structure has been determined to 1.9 Angstrom resolution. The enzyme is composed of two domains. The N-t erminal domain of approximately 300 residues contains mostly alpha-helices which form a doubly-layered horseshoe (a subset of the (alpha/alpha)(6) tor oidal topology). The similar to 370 residues long C-terminal domain is made of beta-strands arranged in a four layered beta-sheet sandwich, with the f irst two sheets having nine strands each. This fold is novel and has no cou nterpart in full among known structures. The sequence of chondroitinase AC shows low level of homology to several hyaluronate lyases, which likely sha re its fold. The shape of the molecule, distribution of electrostatic poten tial, the pattern of conservation of the amino acids and the results of mut agenesis of hyaluronate lyases, indicate that the enzymatic activity reside s primarily within the N-terminal domain. The most likely candidate for the catalytic base is His225. Other residues involved in catalysis and/or subs trate binding are Arg288, Arg292, Lys298 and Lys299. (C) 1999 Academic Pres s.