Adenosine-5′-phosphosulfate kinase from Thermobifida fusca

Abstract

Sulfur, as a macronutrient, is essential for all kinds of organisms. Sulfate, the primary available source of sulfur, is firstly activated by adenylation catalyzed by ATP sulfurylase (ATPS) to form adenosine 5′-phosphosulfate (APS), which will be further phosphorylated into 3′-phosphoadenosine 5′-phosphosulfate (PAPS) by APS kinase (APSK). In some organisms, sulfate activating related enzymes are assembled to form sulfate-activating complex (SAC) to facilitate APS synthesis, the thermodynamically unfavorable reaction. In genome of a moderate thermophilic bacterium, Thermobifida fusca, there are presumably GTPasecoupled ATPS and one putative bifunctional ATPS/APSK type SAC. In this study, this putative SAC of T. fusca was prokaryotically expressed, purified and characterized. Activity assays showed that it contained APSK activity, while lacked ATPS activity. SAC of T. fusca was further used as a coupling enzyme to assay APS formation catalyzed by yeast ATPS. Based on the sequence alignment and modeled structure, we infer that the divergences of two conserved motifs and the missing of a loop and a helix-turn-helix motifs may contribute to the deficiency of ATPS activity.