Search

Electrostatic interactions between charged gatekeeper residues of the enzyme and approaching substrates can effectively discriminate between similar substrates. We investigated whether the charged gatekeeper residues play any role in the substrate specificity of P. falciparum succinyl‐CoA synthetase (PfSCS). Our results demonstrated that charged gatekeeper residues in PfSCS did alter the ATP binding but could not alter its substrate specificity, pointing toward other factors affecting the substate specificity of PfSCS., Substrate specificity of an enzyme is an important characteristic of its mechanism of action. Investigation of the nucleotide specificity of Plasmodium falciparum succinyl‐CoA synthetase (SCS; PfSCS) would provide crucial insights of its substrate recognition. Charged gatekeeper residues have been shown to alter the substrate specificity via electrostatic interactions with approaching substrates. The enzyme kinetics of recombinant PfSCS (wild‐type), generated by refolding of the individual P. falciparum SCSβ and Blastocystis SCSα subunits, demonstrated ADP‐forming activity (K mATP = 48 µm). Further, the introduction of charged gatekeeper residues, either positive (Lys and Lys) or negative (Glu and Asp), resulted in significant reductions in the ATP affinity of PfSCS. It is interesting to note that the recombinant PfSCSβ subunit can be refolded to a functional enzyme conformation using Blastocystis SCSα, indicating the possibility of subunits swapping among different organisms. These results concluded that electrostatic interactions at the gatekeeper region alone are insufficient to alter the substrate specificity of PfSCS, and further structural analysis with a particular focus on binding site architecture is required.