6U1K

Thermus thermophilus D-alanine-D-alanine ligase in complex with ADP, carbonate, D-alanine-D-alanine, Mg2+ and K+

Summary for 6U1K

• Entry DOI: 10.2210/pdb6u1k/pdb
• Related: 6U1C 6U1D 6U1E 6U1F 6U1G 6U1H 6U1I 6U1J
• Descriptor: D-alanine--D-alanine ligase, D-ALANINE, MAGNESIUM ION, ... (7 entities in total)
• Functional Keywords: atp-grasp domain, atp binding, metal binding, ligase
• Biological source: Thermus thermophilus (strain HB8 / ATCC 27634 / DSM 579)
• Total number of polymer chains: 4
• Total formula weight: 146168.37

Authors

Pederick, J.L.,Bruning, J.B. (deposition date: 2019-08-15, release date: 2020-05-06, Last modification date: 2023-10-11)

Primary citation

Pederick, J.L.,Thompson, A.P.,Bell, S.G.,Bruning, J.B.
d-Alanine-d-alanine ligase as a model for the activation of ATP-grasp enzymes by monovalent cations.
J.Biol.Chem., 295:7894-7904, 2020

PubMed Abstract: The ATP-grasp superfamily of enzymes shares an atypical nucleotide-binding site known as the ATP-grasp fold. These enzymes are involved in many biological pathways in all domains of life. One ATP-grasp enzyme, d-alanine-d-alanine ligase (Ddl), catalyzes ATP-dependent formation of the d-alanyl-d-alanine dipeptide essential for bacterial cell wall biosynthesis and is therefore an important antibiotic drug target. Ddl is activated by the monovalent cation (MVC) K, but despite its clinical relevance and decades of research, how this activation occurs has not been elucidated. We demonstrate here that activating MVCs bind adjacent to the active site of Ddl from and used a combined biochemical and structural approach to characterize MVC activation. We found that Ddl is a type II MVC-activated enzyme, retaining activity in the absence of MVCs. However, the efficiency of Ddl increased ∼20-fold in the presence of activating MVCs, and it was maximally activated by K and Rb ions. A strict dependence on ionic radius of the MVC was observed, with Li and Na providing little to no Ddl activation. To understand the mechanism of MVC activation, we solved crystal structures of Ddl representing distinct catalytic stages in complex with K, Rb, or Cs Comparison of these structures with apo Ddl revealed no evident conformational change on MVC binding. Of note, the identified MVC binding site is structurally conserved within the ATP-grasp superfamily. We propose that MVCs activate Ddl by altering the charge distribution of its active site. These findings provide insight into the catalytic mechanism of ATP-grasp enzymes.

PubMed: 32335509
DOI: 10.1074/jbc.RA120.012936

Experimental method

X-RAY DIFFRACTION (1.67 Å)