Kinetic characterization and structural modeling of an NADP+-dependent succinic semialdehyde dehydrogenase from Anabaena sp. PCC7120.

Succinic semialdehyde dehydrogenases (SSADH) of cyanobacteria played a pivotal role in completing the cyanobacterial tricarboxylic acid cycle. The structural information of cofactor preference and catalysis for SSADH from cyanobacteria is currently available. However, the detailed kinetics of SSADH from cyanobacteria were not characterized yet. In this study, an all3556 gene encoding SSADH from Anabaena sp. PCC7120 (ApSSADH) was amplified and the recombinant ApSSADH was purified homogenously. Kinetic analysis showed that ApSSADH was an NADP+-dependent SSADH, which utilized NADP+ and succinic semialdehyde (SSA) as its preferred substrates and the activity of ApSSADH was inhibited by its substrate of SSA. At the same time, the Ser157 residue was found to function as the determinant of cofactor preference. Further study demonstrated that activity and substrate inhibition of ApSSADH would be greatly reduced by the mutation of the residues at the active site. Bioinformatic analysis indicated that those residues were highly conserved throughout the SSADHs. To our knowledge this is the first report exploring the detailed kinetics of SSADH from cyanobacteria.