Comparative structure and function analyses of native and his-tagged forms of dihydrodipicolinate reductase from methicillin-resistant Staphylococcus aureus

Given the rise of multi drug resistant bacterial strains, such as methicillin-resistant Staphylococcus aureus   (MRSA), there is an urgent need to discover new antimicrobial agents. A validated but as yet unexplored target for new antibiotics is dihydrodipicolinate reductase (DHDPR), an enzyme that catalyzes the second step of the lysine biosynthesis pathway in bacteria. We report here the cloning, expression and purification of N-terminally his-tagged recombinant DHDPR from MRSA (6H-MRSA–DHDPR) and compare its secondary and quaternary structure with the wild type (MRSA–DHDPR) enzyme. Comparative analyses demonstrate that recombinant 6H-MRSA–DHDPR is folded and adopts the native tetrameric quaternary structure in solution. Furthermore, kinetic studies show 6H-MRSA–DHDPR is functional, displaying parameters for KmNADH of 6.0 μM, KmDHDP of 22 μM, and kcat of 21 s−1, which are similar to those reported for the native enzyme. The solution properties and stability of the 6H-MRSA–DHDPR enzyme are also reported in varying physicochemical conditions.

► This study compares the solution properties of native and his-tagged MRSA–DHDPR.
► The cloning, expression and purification of his-tagged DHDPR is described.
► This is the first study to employ AUC to examine the quaternary structure of DHDPR.
► Native and his-tagged enzymes display similar kinetic and structure characteristics.
► This work will provide insight into drug design strategies targeting MRSA–DHDPR.