Download MendeleyConformational Itinerary of Pseudomonas aeruginosa 1,6-Anhydro-N-acetylmuramic Acid Kinase during Its Catalytic Cycle.
Bacik, J.P., Tavassoli, M., Patel, T.R., McKenna, S.A., Vocadlo, D.J., Khajehpour, M., Mark, B.L.(2014) J Biological Chem 289: 4504-4514
- PubMed: 24362022
- DOI: https://doi.org/10.1074/jbc.M113.521633
- Primary Citation of Related Structures:
4MO4, 4MO5 - PubMed Abstract:
Anhydro-sugar kinases are unique from other sugar kinases in that they must cleave the 1,6-anhydro ring of their sugar substrate to phosphorylate it using ATP. Here we show that the peptidoglycan recycling enzyme 1,6-anhydro-N-acetylmuramic acid kinase (AnmK) from Pseudomonas aeruginosa undergoes large conformational changes during its catalytic cycle, with its two domains rotating apart by up to 32¡ã around two hinge regions to expose an active site cleft into which the substrates 1,6-anhydroMurNAc and ATP can bind. X-ray structures of the open state bound to a nonhydrolyzable ATP analog (AMPPCP) and 1,6-anhydroMurNAc provide detailed insight into a ternary complex that forms preceding an operative Michaelis complex. Structural analysis of the hinge regions demonstrates a role for nucleotide binding and possible cross-talk between the bound ligands to modulate the opening and closing of AnmK. Although AnmK was found to exhibit similar binding affinities for ATP, ADP, and AMPPCP according to fluorescence spectroscopy, small angle x-ray scattering analyses revealed that AnmK adopts an open conformation in solution in the absence of ligand and that it remains in this open state after binding AMPPCP, as we had observed for our crystal structure of this complex. In contrast, the enzyme favored a closed conformation when bound to ADP in solution, consistent with a previous crystal structure of this complex. Together, our findings show that the open conformation of AnmK facilitates binding of both the sugar and nucleotide substrates and that large structural rearrangements must occur upon closure of the enzyme to correctly align the substrates and residues of the enzyme for catalysis.
Organizational Affiliation:
From the Departments of Microbiology and.
