Spectrofluorometric assay for monoamine-preferring phenol sulfotransferase (SULT1A3)

Abstract

A continuous and real-time fluorometric assay for monoamine-preferring phenol sulfotransferase (SULT1A3) was developed. The methodology was based on the coupling of SULT1A1 to regenerate 3'-phosphoadenosine-5'-phosPhosulfate (PAPS) using 4-methylumbelliferyl sulfate (MUS) as a sulfuryl group donor. The fluorophore product (4-methylumbelliferone. MU) was continuously produced and monitored when SULT1A3 catalyzed dopamine sulfation with PAPS. The optimal conditions of this turnover reaction and substrate inhibition of SULT1A3 were also determined. This coupled-enzyme assay allows the continuous measurement of initial reaction velocity and the sensitivity is comparable to that of end-point radioactive isotope assay. (C) 2010 Elsevier Inc. All rights reserved.