Kinetic properties of Streptococcus pneumoniae hyaluronate lyase.
Streptococcus pneumoniae hyaluronate lyase is a surface antigen of this bacterial pathogen, which causes significant mortality and morbidity in human populations worldwide. The primary function of this enzyme is the degradation of hyaluronan, a major component of the extracellular matrix of the tissues of practically all vertebrates. The enzyme uses a processive mode of action to degrade hyaluronan to a final product, an unsaturated disaccharide hyaluronan unit. This catalysis proceeds via a five-step proton acceptance and donation mechanism that includes substrate binding, catalysis, release of the disaccharide product, translocation of the remaining hyaluronan substrate, and proton exchange with microenvironment. Based on the analysis of the three-dimensional structure of the native enzyme and its complexes with hexasaccharide substrate and disaccharide product, several residues have been chosen for mutation studies. These mutated residues included the catalytic residues Asn349, His399, Tyr408, and residues responsible for substrate binding and translocation, Arg243 and Asn580. The comparison of the kinetic properties of the wild-type with the mutant enzymes allowed for the characterization of every mutant and the correlation of the kinetic properties of the enzyme with its structure. The comparison of the wild-type hyaluronate lyase with other polysaccharide-degrading enzymes, the hydrolases endonuclease and glucoamylase, shows striking similarity of K(m)s for all of these different enzymes.
Kelly
S J
SJ
Department of Microbiology, 933 19th Street South, 545 CHSB-19, University of Alabama at Birmingham, Birmingham AL 35294, USA.
Taylor
K B
KB
Li
S
S
Jedrzejas
M J
MJ
D001665:Binding Sites; D002240:Carbohydrate Sequence; D006801:Humans; D006820:Hyaluronic Acid / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011133:Polysaccharide-Lyases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D013296:Streptococcus pneumoniae / Q000201:enzymology* / Q000235:genetics; D014470:Umbilical Cord
D006820:Hyaluronic Acid; D011133:Polysaccharide-Lyases; C100002:hyaluronate lyase