Paper disk assay for glycosaminoglycan sulfotransferases.
A method is described for the assay of sulfotransferases, which transfer sulfate from 3'-phosphoadenosine-5'-phosphosulfate (PAPS) to glycosaminoglycan acceptors. Following the sulfation reactions, the [35S]sulfate-labeled products are precipitated and then separated from a sulfate donor ([35S]PAPS) and its degradation products by a paper disk method, and then the radioactivity remaining on the paper disk is subsequently determined by liquid scintillation counting. The rapidity and simplicity of the method are advantageous for multiple assays and have allowed us to establish assay conditions for serum sulfotransferases which introduce sulfate at position 6 of the internal N-acetylgalactosamine units of chondroitin, position 2 (amino group) of the glucosamine units of heparan sulfate and sugar units of keratan sulfate, respectively. The assay method will be applicable with modification to the assay of other glycosaminoglycan sulfotransferases and glycoprotein sulfotransferases.
D000818:Animals; D002021:Buffers; D002645:Chickens; D004589:Electrophoresis, Disc; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000494:pharmacology; D013045:Species Specificity; D015238:Sulfotransferases; D013462:Sulfur Radioisotopes; D013466:Sulfurtransferases / Q000032:analysis* / Q000097:blood
D002021:Buffers; D008670:Metals; D013462:Sulfur Radioisotopes; C022321:PAPS sulfotransferase; D013466:Sulfurtransferases; D015238:Sulfotransferases; C019893:chondroitin 4-sulfotransferase