Photoaffinity labeling probe for the substrate binding site of human phenol sulfotransferase (SULT1A1): 7‐Azido‐4‐methylcoumarin

• DOI: 10.1110/ps.8.10.2151
• Publication Date: 2009-02-17T21:25:15Z
• Authors: Guangping Chen; Eric Battaglia; Claire Senay; Charles N. Falany; Anna Radominska‐P...
• Keywords: Maltose-binding protein; Photoaffinity labeling; Moiety; Substrate analog; Cleavage (geology)

Abstract A novel fluorescent photoactive probe 7‐azido‐4‐methylcoumarin (AzMC) has been characterized for use in photoaffinity labeling of the substrate binding site human phenol sulfotransferase (SULT1A1 or P‐PST‐1). For experiments, SULT1A1 cDNA was expressed Escherichia coli as a fusion protein to maltose (MBP) and purified apparent homogeneity over an amylose column. The moiety removed by Factor Xa cleavage. Both MBSULT1A1 were efficiently photolabeled with AzMC. This concentration dependent. In absence light, AzMC competitively inhibited sulfation 4MU catalyzed ( K i = 0.47 ± 0.05 mM). Moreover, enzyme activity toward 2‐naphthol inactivated timeand concentration‐dependent manner. inactivation protected but not cosubstrate. These results indicate that is highly suitable identification SULT1A1. Further studies are aimed at identifying which amino acids modified localized site.