title: Characterization of the active site of group B streptococcal hyaluronan lyase.
abstract: Hyaluronan lyase is secreted by most strains of the human pathogen, group B streptococcus. Site-directed mutagenesis of the enzyme identified three amino acid residues important for enzyme activity, H479, Y488, and R542. These three residues are in close proximity in the putative active site of a homology model of group B streptococcal hyaluronan lyase. The homology model was based on the crystal structure of another related glycosaminoglycan lyase, chondroitin AC lyase, which exhibits different substrate specificity. Two asparagine residues in the active site groove, N429 and N660, were also found to be essential for enzyme activity. In addition, conversion of two adjacent tryptophan residues in the groove to alanines abolished activity. All amino acids found to be essential in GBS hyaluronan lyase are conserved in both enzymes. However, several amino acids in the active site groove of the two enzymes are not conserved. In the 18 cases in which one of these amino acids in GBS hyaluronan lyase was replaced with its corresponding amino acid in chondroitin AC lyase, no major loss of activity or change in substrate specificity was observed.
authors.affiliation: Department of Biochemistry & Molecular Genetics, University of Alabama at Birmingham, 35294-0007, USA. davidp1@uab.edu
authors.forename: D G
authors.initials: DG
authors.lastname: Pritchard
authors.forename: J O
authors.initials: JO
authors.lastname: Trent
authors.forename: X
authors.initials: X
authors.lastname: Li
authors.forename: P
authors.initials: P
authors.lastname: Zhang
authors.forename: M L
authors.initials: ML
authors.lastname: Egan
authors.forename: J R
authors.initials: JR
authors.lastname: Baker
mesh_terms: D002118:Calcium / Q000737:chemistry; D002240:Carbohydrate Sequence; D020134:Catalytic Domain; D002808:Chondroitin Lyases / Q000737:chemistry; D020832:EF Hand Motifs; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011133:Polysaccharide-Lyases / Q000737:chemistry* / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D013292:Streptococcus agalactiae / Q000737:chemistry*
chemical_list: D002808:Chondroitin Lyases; D011133:Polysaccharide-Lyases; C100002:hyaluronate lyase; D002118:Calcium