title: Synthesis of heparosan oligosaccharides by Pasteurella multocida PmHS2 single-action transferases.
abstract: Pasteurella multocida heparosan synthase PmHS2 is a dual action glycosyltransferase that catalyzes the polymerization of heparosan polymers in a non-processive manner. The two PmHS2 single-action transferases, obtained previously by site-directed mutagenesis, have been immobilized on Ni(II)-nitrilotriacetic acid agarose during the purification step. A detailed study of the polymerization process in the presence of non-equal amounts of PmHS2 single-action transferases revealed that the glucuronyl transferase (PmHS2-GlcUA(+)) is the limiting catalyst in the polymerization process. Using experimental design, it was determined that the N-acetylglucosaminyl transferase (PmHS2-GlcNAc(+)) plays an important role in the control of heparosan chain elongation depending on the number of heparosan chains and the UDP-sugar concentrations present in the reaction mixture. Furthermore, for the first time, the synthesis of heparosan oligosaccharides alternately using PmHS2-GlcUA(+) and PmHS2-GlcNAc(+) is reported. It was shown that the synthesis of heparosan oligosaccharides by PmHS2 single-action transferases do not require the presence of template molecules in the reaction mixture.
authors.affiliation: Bioprocess Engineering Group, Wageningen University and Research Center, P.O. Box 8129, 6700 EV, Wageningen, the Netherlands.
authors.forename: Anaïs A E
authors.initials: AA
authors.lastname: Chavaroche
authors.forename: Lambertus A M
authors.initials: LA
authors.lastname: van den Broek
authors.forename: Carmen
authors.initials: C
authors.lastname: Boeriu
authors.forename: Gerrit
authors.initials: G
authors.lastname: Eggink
mesh_terms: D004187:Disaccharides / Q000378:metabolism*; D004800:Enzymes, Immobilized / Q000235:genetics / Q000378:metabolism; D016695:Glycosyltransferases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016979:Pasteurella multocida / Q000201:enzymology*
chemical_list: D004187:Disaccharides; D004800:Enzymes, Immobilized; C560363:heparosan; D016695:Glycosyltransferases; C450833:heparosan synthase