Nucleotide Binding States of Subunit A of the A-ATP Synthase and the Implication of P-Loop Switch in Evolution |
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Authors: | Anil Kumar Asha Manikkoth Balakrishna Gerhard Grüber |
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Institution: | 1 School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551, Republic of Singapore 2 National Synchrotron Radiation Research Center, 101 Hsin-Ann Road, Hsinchu Science Park, Hsinchu 30076, Taiwan 3 Bioinformatics Institute (A?STAR), 30 Biopolis Street, 07-01 Matrix, Singapore 138671, Republic of Singapore |
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Abstract: | The crystal structures of the nucleotide-empty (AE), 5′-adenylyl-β,γ-imidodiphosphate (APNP)-bound, and ADP (ADP)-bound forms of the catalytic A subunit of the energy producer A1AO ATP synthase from Pyrococcus horikoshii OT3 have been solved at 2.47 Å and 2.4 Å resolutions. The structures provide novel features of nucleotide binding and depict the residues involved in the catalysis of the A subunit. In the AE form, the phosphate analog SO42− binds, via a water molecule, to the phosphate binding loop (P-loop) residue Ser238, which is also involved in the phosphate binding of ADP and 5′-adenylyl-β,γ-imidodiphosphate. Together with amino acids Gly234 and Phe236, the serine residue stabilizes the arched P-loop conformation of subunit A, as shown by the 2.4-Å structure of the mutant protein S238A in which the P-loop flips into a relaxed state, comparable to the one in catalytic β subunits of F1FO ATP synthases. Superposition of the existing P-loop structures of ATPases emphasizes the unique P-loop in subunit A, which is also discussed in the light of an evolutionary P-loop switch in related A1AO ATP synthases, F1FO ATP synthases, and vacuolar ATPases and implicates diverse catalytic mechanisms inside these biological motors. |
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Keywords: | P-loop phosphate binding loop A-ATP synthase A1AO ATP synthase F-ATP synthase F1FO ATP synthase V-ATPase vacuolar ATPase PMF proton motive force AMP-PNP 5&prime -adenylyl-β γ-imidodiphosphate MPD 2-methyl-2 4 pentanediol WT-A wild-type subunit A ITC isothermal titration calorimetry r m s d root-mean-square distance PDB Protein Data Bank NSRRC National Synchrotron Radiation Research Center |
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