Abstract

The two-component phosphorelay system is the most pre-valent mechanism for sensing and transducing environ-mental signals in bacteria. Spore formation, which relies on the two-component phosphorelay system, enables the long- term survival of the glacial bacterium Paenisporosarcina sp. TG-14 in the extreme cold environment. Spo0A is a key re-sponse regulator of the phosphorelay system in the early stage of spore formation. The protein is composed of a regu-latory N-terminal phospho-receiver domain and a DNA- binding C-terminal activator domain. We solved the three- dimensional structure of the unphosphorylated (inactive) form of the receiver domain of Spo0A (PaSpo0A-R) from Paenisporosarcina sp. TG-14. A structural comparison with phosphorylated (active form) Spo0A from Bacillus stearo-thermophilus (BsSpo0A) showed minor notable differences. A molecular dynamics study of a model of the active form and the crystal structures revealed significant differences in the α4 helix and the preceding loop region where phosphorylation occurs. Although an oligomerization study of PaSpo0A-R by analytical ultracentrifugation (AUC) has shown that the protein is in a monomeric state in solution, both crosslinking and crystal-packing analyses indicate the possibility of weak dimer formation by a previously undocumented mechanism. Collectively, these observations provide insight into the me-chanism of phosphorylation-dependent activation unique to Spo0A.

• Keywords: nalytical ultracentrifugation;Paenisporosarcina sp. TG-14;spore formation;Spo0A;X-ray crystallography