title: A structural and biochemical model of processive chitin synthesis.
abstract: Chitin synthases (CHS) produce chitin, an essential component of the fungal cell wall. The molecular mechanism of processive chitin synthesis is not understood, limiting the discovery of new inhibitors of this enzyme class. We identified the bacterial glycosyltransferase NodC as an appropriate model system to study the general structure and reaction mechanism of CHS. A high throughput screening-compatible novel assay demonstrates that a known inhibitor of fungal CHS also inhibit NodC. A structural model of NodC, on the basis of the recently published BcsA cellulose synthase structure, enabled probing of the catalytic mechanism by mutagenesis, demonstrating the essential roles of the DD and QXXRW catalytic motifs. The NodC membrane topology was mapped, validating the structural model. Together, these approaches give insight into the CHS structure and mechanism and provide a platform for the discovery of inhibitors for this antifungal target.
authors.affiliation: Division of Molecular Microbiology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom. Electronic address: hczdorfmueller@dundee.ac.uk.
authors.forename: Helge C
authors.initials: HC
authors.lastname: Dorfmueller
authors.affiliation: Division of Molecular Microbiology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom.
authors.forename: Andrew T
authors.initials: AT
authors.lastname: Ferenbach
authors.affiliation: MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom.
authors.forename: Vladimir S
authors.initials: VS
authors.lastname: Borodkin
authors.affiliation: Division of Molecular Microbiology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom; MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom. Electronic address: dmfvanaalten@dundee.ac.uk.
authors.forename: Daan M F
authors.initials: DMF
authors.lastname: van Aalten
mesh_terms: D020816:Amino Acid Motifs; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D002686:Chitin / Q000096:biosynthesis / Q000737:chemistry; D002687:Chitin Synthase / Q000737:chemistry / Q000378:metabolism; D005964:Glucosyltransferases / Q000737:chemistry / Q000378:metabolism; D008954:Models, Biological; D008958:Models, Molecular; D016962:Sinorhizobium meliloti / Q000201:enzymology*
chemical_list: D001426:Bacterial Proteins; D002686:Chitin; D005964:Glucosyltransferases; C478648:cellulose synthase; D002687:Chitin Synthase
keywords: Carbohydrate Biosynthesis; Enzyme Inhibitor; Enzyme Mechanism; Glycosyltransferase; Protein Structure