Structural aspects of nucleotide ligand binding by a bacterial 2H phosphoesterase

Myllykoski, Matti; Kursula, Petri

Another product analogue of the reaction catalysed by LigT, NADP + , was also trapped in the active site, being well defined in electron density (Fig 2B) . In the NADP + complex, there are 4 monomers in the asymmetric unit, and the nicotinamide end of the ligand binds to each of them differently, having different conformations or being disordered-depending on crystal contacts. The adenosine 2 0 ,5 0 -bisphosphate moiety in each monomer binds identically, however, and in a mode highly similar to that seen in 2 0 -AMP. The 5 0 -phosphate mimics the next phosphodiester in an RNA molecule, and it can be used to deduce further binding determinants for RNA substrates. This phosphate moiety is bound by Arg6 from the N-terminal strand β1 in the crystal structure, and it is likely that this residue plays a direct role in RNA A. Overall structure of LigT. Secondary structure elements and the N and C termini are labeled, and the two active site HxT motif side chains are also shown. B. Stereo view of the organization and conservation of the active site between LigT (white) and mouse CNPase (blue) [18] . The four water molecules between strands 2 and 6 are conserved (LigT, red; CNPase, blue). The nucleophilic water molecule in LigT (green) is coordinated by His125 and Arg130, while the corresponding water molecule in CNPase (magenta) interacts with the N terminus of helix 7 (right) and the catalytic histidine. substrate binding also. An additional NADP + fragment is seen stacked on top of Trp82 in one monomer.