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X‐ray crystallographic and enzymatic analyses of shikimate dehydrogenase from Staphylococcus epidermidis
Author(s) -
Han Cong,
Hu Tiancen,
Wu Dalei,
Qu Su,
Zhou Jiahai,
Ding Jianping,
Shen Xu,
Qu Di,
Jiang Hualiang
Publication year - 2009
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/j.1742-4658.2008.06856.x
Subject(s) - shikimate pathway , staphylococcus epidermidis , enzyme , chemistry , staphylococcus , dehydrogenase , crystallography , biochemistry , microbiology and biotechnology , stereochemistry , staphylococcus aureus , biology , bacteria , biosynthesis , genetics
Shikimate dehydrogenase (SDH) catalyzes the NADPH‐dependent reduction of 3‐dehydroshikimate to shikimate in the shikimate pathway. In this study, we determined the kinetic properties and crystal structures of Staphylococcus epidermidis SDH (SeSDH) both in its ligand‐free form and in complex with shikimate. SeSDH has a k cat of 22.8 s −1 and a K m of 73 μ m towards shikimate, and a K m of 100 μ m towards NADP. The overall folding of SeSDH comprises the N‐terminal α/β domain for substrate binding and the C‐terminal Rossmann fold for NADP binding. The active site is within a large groove between the two domains. Residue Tyr211, normally regarded as important for substrate binding, does not interact with shikimate in the binary SeSDH–shikimate complex structure. However, the Y211F mutation leads to a significant decrease in k cat and a minor increase in the K m for shikimate. The results indicate that the main function of Tyr211 may be to stabilize the catalytic intermediate during catalysis. The NADP‐binding domain of SeSDH is less conserved. The usually long helix specifically recognizing the adenine ribose phosphate is substituted with a short 3 10 helix in the NADP‐binding domain. Moreover, the interdomain angle of SeSDH is the widest among all known SDH structures, indicating an inactive ‘open’ state of the SeSDH structure. Thus, a ‘closing’ process might occur upon NADP binding to bring the cofactor close to the substrate for catalysis.