The Highly Conserved Cys95 Residue of Fructose-1,6-Bisphosphatase 1 Mediates the pH-Driven Structure and Activity of the Enzyme and Photosynthesis.

In Arabidopsis, exposure to microbial volatile compounds promotes thiol reduction of the Cys95 residue of the photosynthetic enzyme fructose-1,6-bisphosphatase (cFBP1). Although highly conserved in plants, the Cys95 function still remains unknown. We characterised recombinant wild-type (WT) cFBP1 and a variant (C95S) in which the Cys95 residue was replaced by serine. Furthermore, we characterised cFBP1-lacking cfbp1 transgenic plants expressing WT or C95S cFBP1. Cys95 replacement by serine reduced cFBP1 activity and its Mg binding affinity and cooperativity. Although it is widely assumed that active cFBP1 is strictly homotetrameric, WT and C95S cFBP1 were present as inactive tetramers at pH 7.0 and active dimers at pH 8.3. At pH 7.8, WT and C95S cFBP1 were predominantly present as dimers and tetramers, respectively. WT cFBP1 expression totally reverted to WT the reduced photosynthetic activity of cfbp1 plants grown in the absence or presence of microbial volatiles, but that of C95S cFBP1 only partially did it. Artificial intelligence-based AlphaFold protein structure analyses predicted that the replacement of Cys95 by serine promotes cFBP1 conformational changes. We conclude that (i) active cFBP1 is strictly dimeric at pH values occurring in illuminated chloroplasts and (ii) Cys95 is an important determinant of the stromal pH-driven structure and activity of cFBP1 and photosynthesis.