Abstract
The CO2-fixing enzyme ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO) catalyzes the complex multi-step reaction starting with enolization of ribulose 1,5-bisphosphate (RuBP). The RuBisCO-like protein from Bacillus subtilis (BsRLP) catalyzes the 2,3-diketo-5-methylthiopentyl-1-phosphate (DK-MTP-1-P) enolase reaction, which resembles the enolization of RuBP. Here we determine the general enzymatic properties of the BsRLP. BsRLP activity required Mg2+ for catalysis and was activated by CO2 as well as RuBisCO. The four residues Lys175, Lys201, Asp203 and Glu204, essential for enolization reaction of RuBisCO were conserved in BsRLP and essential for BsRLP catalysis. The Lys123, the residue conserved in DK-MTP-1-P enolases, was also essential. Interestingly, the substrate, the product and the transition-state analog of RuBisCO (RuBP, phosphoglycerate and 2-carboxy-D-arabinitol-1,5-bisphosphate, respectively) exhibited competitive inhibition with respect to DK-MTP-1-P for BsRLP activity. These results suggest that BsRLP utilizes the same amino acid residues for its enolase reaction in similar ways as does RuBisCO for RuBP enolization.
