Abstract
Glutathionylspermidine synthetase of Helicobacter pylori (HpGspS), which belongs to the ATP-grasp superfamily, catalyzes glutathionylspermidine (Gsp) formation by coupling glutathione (GSH) and spermidine with ATP hydrolysis. HpGspS was cloned into the pQE30 vector and overexpressed in E. coli. HpGspS is monomer in solution, with molecular weight 43 kDa and the theoretical pI is 4.6. AMPPNP, a nonhydrolyzable ATP analogue, was used to imitate the ATP binding site in HpGspS. According to apo_HpGspS, HpGspS_AMPPNP and HpGspS_ADP_Pi structures, HpGspS comprises three conserved structural units, including lid domain, antiparallel β-sheet and parallel β-sheet and the structure displays the ATP-grasp fold. The ATP binding site of HpGspS structure is located at the central antiparallel β-sheet and is surrounded by five loops. The adenine ring and ribose of AMPPNP are buried in a hydrophobic pocket in HpGspS. We find some conserved residues participating in ATP binding, Arg98, Asp100, Glu114, Asn116, Lys276, Lys308 and Arg372.