title: Chemoenzymatic synthesis with distinct Pasteurella heparosan synthases: monodisperse polymers and unnatural structures.
abstract: Heparosan (-GlcUA-beta1,4-GlcNAc-alpha1,4-)(n) is a member of the glycosaminoglycan polysaccharide family found in the capsule of certain pathogenic bacteria as well as the precursor for the vertebrate polymers, heparin and heparan sulfate. The two heparosan synthases from the Gram-negative bacteria Pasteurella multocida, PmHS1 and PmHS2, were efficiently expressed and purified using maltose-binding protein fusion constructs. These relatively homologous synthases displayed distinct catalytic characteristics. PmHS1, but not PmHS2, was able to produce large molecular mass (100-800 kDa) monodisperse polymers in synchronized, stoichiometrically controlled reactions in vitro. PmHS2, but not PmHS1, was able to utilize many unnatural UDP-sugar analogs (including substrates with acetamido-containing uronic acids or longer acyl chain hexosamine derivatives) in vitro. Overall these findings reveal potential differences in the active sites of these two Pasteurella enzymes. In the future, these catalysts should allow the creation of a variety of heparosan and heparinoids with utility for medical applications.
authors.affiliation: Department of Biochemistry and Molecular Biology, Oklahoma Center for Medical Glycobiology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
authors.forename: Alison E
authors.initials: AE
authors.lastname: Sismey-Ragatz
authors.affiliation: Department of Biochemistry and Molecular Biology, Oklahoma Center for Medical Glycobiology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
authors.forename: Dixy E
authors.initials: DE
authors.lastname: Green
authors.affiliation: Department of Biochemistry and Molecular Biology, Oklahoma Center for Medical Glycobiology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
authors.forename: Nigel J
authors.initials: NJ
authors.lastname: Otto
authors.affiliation: Department of Biological Chemistry, John Innes Centre, Norwich Research Park, Colney Lane, Norwich NR4 7UH, United Kingdom.
authors.forename: Martin
authors.initials: M
authors.lastname: Rejzek
authors.affiliation: Department of Biological Chemistry, John Innes Centre, Norwich Research Park, Colney Lane, Norwich NR4 7UH, United Kingdom.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Field
authors.affiliation: Department of Biochemistry and Molecular Biology, Oklahoma Center for Medical Glycobiology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104. Electronic address: paul-deangelis@ouhsc.edu.
authors.forename: Paul L
authors.initials: PL
authors.lastname: DeAngelis
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics; D006025:Glycosaminoglycans / Q000138:chemical synthesis* / Q000737:chemistry; D016695:Glycosyltransferases / Q000737:chemistry* / Q000235:genetics; D058619:Maltose-Binding Proteins; D016979:Pasteurella multocida / Q000201:enzymology* / Q000235:genetics; D011108:Polymers / Q000138:chemical synthesis* / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics; D014539:Uridine Diphosphate Sugars / Q000737:chemistry; D014574:Uronic Acids / Q000737:chemistry
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D006025:Glycosaminoglycans; D058619:Maltose-Binding Proteins; D011108:Polymers; D011993:Recombinant Fusion Proteins; D014539:Uridine Diphosphate Sugars; D014574:Uronic Acids; D016695:Glycosyltransferases; C450833:heparosan synthase