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Publication Abstract from PubMed

Heparinase II (HepII) is an 85-kDa dimeric enzyme that depolymerizes both heparin and heparan sulfate glycosaminoglycans through a beta-elimination mechanism. Recently, we determined the crystal structure of HepII from Pedobacter heparinus (previously known as Flavobacterium heparinum) in complex with a heparin disaccharide product, and identified the location of its active site. Here we present the structure of HepII complexed with a heparan sulfate disaccharide product, proving that the same binding/active site is responsible for the degradation of both uronic acid epimers containing substrates. The key enzymatic step involves removal of a proton from the C5 carbon (a chiral center) of the uronic acid, posing a topological challenge to abstract the proton from either side of the ring in a single active site. We have identified three potential active site residues equidistant from C5 and located on both sides of the uronate product and determined their role in catalysis using a set of defined tetrasaccharide substrates. HepII H202A/Y257A mutant lost activity for both substrates and we determined its crystal structure complexed with a heparan sulfate-derived tetrasaccharide. Based on kinetic characterization of various mutants and the structure of the enzyme-substrate complex we propose residues participating in catalysis and their specific roles.

Catalytic mechanism of heparinase II investigated by site-directed mutagenesis and the crystal structure with its substrate.,Shaya D, Zhao W, Garron ML, Xiao Z, Cui Q, Zhang Z, Sulea T, Linhardt RJ, Cygler M J Biol Chem. 2010 Jun 25;285(26):20051-61. Epub 2010 Apr 19. PMID:20404324^[2]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.