Structure function relationship of vanadate bound to a single site in chloroplast CF1-ATPase as determined by X-ray absorption

Irit Sagi, Yehosua Hochman, Grant Bunker, Shmuel Carmeli, Chanoch Carmeli

Research output: Contribution to journalArticlepeer-review

Abstract

The structure of vanadate, a phosphate analogue which was suggested to function in the presence of tightly bound ADP and divalent cations as a transition state inhibitor of CF1-ATPase, was investigated by X-ray absorption spectroscopy. Analysis of the vanadium K-edge was used for determination of the structure of vanadate bound to a single site in CF1-ATPase containing a single tightly bound ADP. There was a decrease in the intensity of the 1s-3d pre-edge transition and a change in the shape of two other shoulders at the edge region upon binding of vanadate to CF1 in the presence of Mg2+ ions. The changes are due to alteration in the structure of vanadium from tetrahedral to a five-coordinated trigonal bipyramidal geometry. Comparison of the pre-edge peak intensity of ADP-vanadate complex, and model compound resolved by crystallography support the proposed structure of CF1-bound vanadate. 51V NMR measurements were used to verify the pentacoordinated structure of ADP-vanadate complex used as a model in the X-ray absorption studies. The inhibition of a single and multiple site activity by vanadate and by MgADP was measured. Vanadate inhibition of CF1-ATPase activity decreased more than 90 fold in the presence of MgADP. A differential specificity of the inhibition in single and multiple mode of activity was observed. It is suggested that ADP-vanadate binds to the active sites of the enzyme as a pentacoordinated vanadium having approximate trigonal bipyramidal geometry. This structure is analogous to the proposed transition state of the phosphate during the synthesis and the hydrolysis of ATP by CF1.

Original languageEnglish
Pages (from-to)275-285
Number of pages11
JournalPhotosynthesis Research
Volume57
Issue number3
DOIs
StatePublished - 1998

Keywords

  • ATP synthase
  • EXAFS
  • K-edge
  • Kinetics
  • MgADP
  • Transition state

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