X-ray structures of the three Lactococcus lactis dihydroxyacetone kinase subunits and of a transient intersubunit complex

Abstract

Bacterial dihydroxyacetone (Dha) kinases do not exchange the ADP for ATP but utilize a subunit of the phosphoenolpyruvate carbohydrate phosphotransferase system for in situ rephosphorylation of a permanently bound ADP-cofactor. Here we report the 2.1-Å crystal structure of the transient complex between the phosphotransferase subunit DhaM of the phosphotransferase system and the nucleotide binding subunit DhaL of the Dha kinase of Lactococcus lactis, the 1.1-Å structure of the free DhaM dimer, and the 2.5-Å structure of the Dha-binding DhaK subunit. Conserved salt bridges and an edge-to-plane stacking contact between two tyrosines serve to orient DhaL relative to the DhaM dimer. The distance between the imidazole Nε2 of the DhaM His-10 and the β-phosphate oxygen of ADP, between which the γ-phosphate is transferred, is 4.9 Å. An invariant arginine, which is essential for activity, is appropriately positioned to stabilize the γ-phosphate in the transition state. The (βα)4α fold of DhaM occurs a second time as a subfold in the DhaK subunit. By docking DhaL-ADP to this subfold, the nucleotide bound to DhaL and the C1-hydroxyl of Dha bound to DhaK are positioned for in-line transfer of phosphate. © 2008 by The American Society for Biochemistry and Molecular Biology, Inc.