Trimeric ring-like structure of ArsA ATPase

ArsA protein is the soluble subunit of the Ars anion pump in the Escherichi a coli membrane which extrudes arsenite or antimonite from the cytoplasm, T he molecular weight of the subunit is 63 kDa. In the cell it hydrolyzes ATP , and the energy released is used by the membrane-bound subunit ArsB to tra nsport the substrates across the membrane. We have obtained two-dimensional crystals of ArsA in the presence of arsenite on negatively-charged lipid m onolayer composed of DMPS and DOPC. These crystals have been studied using electron microscopy of negatively-stained specimens followed by image proce ssing. The projection map obtained at 2.4 nm resolution reveals a ring-like structure with threefold symmetry. Many molecular assemblies with the same ring-shape and dimensions were also seen dispersed on electron microscopy grids, prepared directly from purified ArsA protein solution. Size-exclusio n chromatography of the protein sample with arsenite present revealed that the majority of the protein particles in solution have a molecular weight o f about 180 kDa. Based on these experiments, we conclude that in solution t he ArsA ATPase with substrate bound is mainly in a trimeric form. (C) 2000 Federation of European Biochemical Societies.