TY - JOUR
T1 - Elucidation of the phosphate binding mode of DING proteins revealed by subangstrom X-ray crystallography
AU - Liebschner, Dorothee
AU - Elias, Mikael
AU - Moniot, Sèbastien
AU - Fournier, Bertrand
AU - Scott, Ken
AU - Jelsch, Christian
AU - Guillot, Benoit
AU - Lecomte, Claude
AU - Chabrière, Eric
PY - 2009/6/10
Y1 - 2009/6/10
N2 - PfluDING is a bacterial protein isolated from Pseudomonas fluorescens that belongs to the DING protein family, which is ubiquitous in eukaryotes and extends to prokaryotes. DING proteins and PfluDING have very similar topologies to phosphate Solute Binding Proteins (SBPs). The three-dimensional structure of PfluDING was obtained at subangstrom resolution (0.88 and 0.98 Å) at two different pH's (4.5 and 8.5), allowing us to discuss the hydrogen bond network that sequesters the phosphate ion in the binding site. From this high resolution data, we experimentally elucidated the molecular basis of phosphate binding in phosphate SBPs. The phosphate ion is tightly bound to the protein via 12 hydrogen bonds between phosphate oxygen atoms and OH and NH groups of the protein. The proton on one oxygen atom of the phosphate dianion forms a 2.5 Å low barrier hydrogen bond with an aspartate, with the energy released by forming this strong bond ensuring the specificity for the dianion even at pH 4.5. In particular, contrary to previous theories on phosphate SBPs, accurate electrostatic potential calculations show that the binding cleft is positively charged. PfluDING structures reveal that only dibasic phosphate binds to the protein at both acidic and basic phosphate, suggesting that the protein binding site environment stabilizes the HPO42- form of phosphate.
AB - PfluDING is a bacterial protein isolated from Pseudomonas fluorescens that belongs to the DING protein family, which is ubiquitous in eukaryotes and extends to prokaryotes. DING proteins and PfluDING have very similar topologies to phosphate Solute Binding Proteins (SBPs). The three-dimensional structure of PfluDING was obtained at subangstrom resolution (0.88 and 0.98 Å) at two different pH's (4.5 and 8.5), allowing us to discuss the hydrogen bond network that sequesters the phosphate ion in the binding site. From this high resolution data, we experimentally elucidated the molecular basis of phosphate binding in phosphate SBPs. The phosphate ion is tightly bound to the protein via 12 hydrogen bonds between phosphate oxygen atoms and OH and NH groups of the protein. The proton on one oxygen atom of the phosphate dianion forms a 2.5 Å low barrier hydrogen bond with an aspartate, with the energy released by forming this strong bond ensuring the specificity for the dianion even at pH 4.5. In particular, contrary to previous theories on phosphate SBPs, accurate electrostatic potential calculations show that the binding cleft is positively charged. PfluDING structures reveal that only dibasic phosphate binds to the protein at both acidic and basic phosphate, suggesting that the protein binding site environment stabilizes the HPO42- form of phosphate.
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U2 - 10.1021/ja901900y
DO - 10.1021/ja901900y
M3 - Article
C2 - 19445459
AN - SCOPUS:67650540884
VL - 131
SP - 7879
EP - 7886
JO - Journal of the American Chemical Society
JF - Journal of the American Chemical Society
SN - 0002-7863
IS - 22
ER -