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-angstroms 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-angstroms structure of the free DhaM dimer, and the 2.5-angstroms 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 Nepsilon2 of the DhaM His-10 and the beta-phosphate oxygen of ADP, between which the gamma-phosphate is transferred, is 4.9 angstroms. An invariant arginine, which is essential for activity, is appropriately positioned to stabilize the gamma-phosphate in the transition state. The (betaalpha)4alpha 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.
Highlights
Dha kinases can be grouped in two structurally homologous but functionally different families
The ADP moiety is not exchanged for ATP [6] but remains permanently bound to the DhaL subunit where it is rephosphorylated in situ by the third subunit, DhaM
This second family of Dha kinases that are exclusively bacterial have switched from ATP to PEP as a source of high energy phosphate
Summary
This second family of Dha kinases that are exclusively bacterial have switched from ATP to PEP as a source of high energy phosphate. Besides covalent substrate binding by DhaK [7] and containing ADP as cofactor, bacterial Dha kinases display another noteworthy property. The DhaK and DhaL subunits or paralogues thereof function as coactivators of dha operon transcription. In Escherichia coli, DhaL-ADP (but not DhaL-ATP), and DhaK without bound Dha act antagonistically as coactivators and corepressor, respectively, of the transcription factor DhaR [8], an enhancer-binding protein [9] consisting of a receiver, an AAAϩ (ATPases associated with various cellular activities), and a DNA binding domain. In Lactococcus lactis, DhaQ, a paralogue of DhaK, acts as coactivator of the transcription factor DhaS, and Dha acts as inducer by binding to DhaQ. The same scaffolds control the phosphate transfer from DhaM phosphohistidine to DhaL-ADP, and from DhaL-ATP to the hydroxyl group of Dha
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
