title: In vitro UDP-sugar:undecaprenyl-phosphate sugar-1-phosphate transferase assay and product detection by thin layer chromatography.
abstract: In vitro assays are invaluable for the biochemical characterization of UDP-sugar:undecaprenyl-phosphate sugar-1-phosphate transferases. These assays typically involve the use of a radiolabeled substrate and subsequent extraction of the product, which resides in a lipid environment. Here, we describe the preparation of bacterial membranes containing these enzymes, a standard in vitro transferase assay with solvents containing chloroform and methanol, and two methods to measure product formation: scintillation counting and thin layer chromatography.
authors.affiliation: Centre for Infection and Immunity, Queen's University Belfast, Belfast, UK.
authors.forename: Kinnari B
authors.initials: KB
authors.lastname: Patel
authors.forename: Miguel A
authors.initials: MA
authors.lastname: Valvano
mesh_terms: D001419:Bacteria / Q000201:enzymology* / Q000378:metabolism; D002855:Chromatography, Thin Layer / Q000379:methods*; D057075:Enzyme Assays / Q000379:methods*; D011106:Polyisoprenyl Phosphates / Q000378:metabolism*; D014166:Transferases / Q000378:metabolism*; D014530:Uridine Diphosphate / Q000378:metabolism*
chemical_list: D011106:Polyisoprenyl Phosphates; C009621:undecaprenyl phosphate; D014530:Uridine Diphosphate; D014166:Transferases