Molecular and biochemical characterizations of three fructose-1,6-bisphosphate aldolases from Clonorchis sinensis

Abstract Fructose-1,6-bisphosphate aldolase (FbA) is a ubiquitous enzyme in glycolysis. In the present study, we screened out three distinct genes encoding FbA isozymes ( Cs FbAs, Cs FbA-1/2/3) from Clonorchis sinensis ( C. sinensis ) and characterized their sequences and structures profiles as well as biochemical properties. The amino acid sequences of Cs FbAs shared homology with those of Class I FbAs from other species. The putative quaternary structures revealed that Cs FbA-2 and Cs FbA-3 were tetramers, while Cs FbA-1 was dimer. Recombinant Cs FbA-2 and Cs FbA-3 (r Cs FbA-2/3) were confirmed to be Class I FbAs for their stable enzymatic activities in the presence of EDTA or metal ions. However, recombinant Cs FbA-1 (r Cs FbA-1) did not show the catalytic activity, which might be due to the inappropriate fold and interaction between its subunits. Both r Cs FbA-2 and r Cs FbA-3 showed similar enzymatic properties such as optimal temperatures and broad pH ranges that similar to human FbA isozymes. They showed relatively higher affinities for fructose-1,6-bisphosphate (FBP) than fructose-1-phosphate (F-1-P). Their k cat ratios of FBP to F-1-P were in accordance with those of human FbA-A or C. In addition, Cs FbAs were differentially transcribed in the developmental stages of C. sinensis , suggesting their essential roles throughout the life stages. Extensive distribution of Cs FbAs in adult worms indicated that ubiquitous activities of Cs FbAs took place in these organs. Collectively, these results suggested that long-term parasitic environment might adapt these isozymes similar to host FbAs for metabolic requirement. Our study will provide new insight into Cs FbAs in the glycometabolism of C. sinensis and relationship between the host and the parasite.