Calcium binding to the photosystem II subunit CP29

We have identified a Ca2+-binding site of the 29-kDa chlorophyll a/b-binding protein CP29, a light harvesting protein of photosystem II most likely involved in photoregulation. Ca-45(2+) binding studies and dot blot analyses of CP29 demonstrate that CP29 is a Ca2+-binding protein. The primary sequence of CP29 does not exhibit an obvious Ca2+-binding site therefore we have used Yb3+ replacement to analyze this site. Near-infrared Yb3+ vibronic side band fluorescence spectroscopy (Roselli, C., Boussac, A. and Mattioli, T. A. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 12897-12901) of Yb3+-reconstituted CP29 indicated a single population of Yb3+-binding sites rich in carboxylic acids, characteristic of Ca2+-binding sites. A structural model of CP29 presents two purported extra-membranar loops which are relatively rich in carboxylic acids, one on the stromae side and one on the lumenal side. The loop on the lumenal side is adjacent to glutamic acid 166 in helix C of CP29, which is known to be the binding site for dicyclohexylcarbodiimide (Pesaresi, P., Sandona, D., Giuffra, E., and Bassi, R. (1997) FEBS Lett. 402, 151-156). Dicyclohexylcarbodiimide binding prevented Ca2+ binding, therefore we propose that the Ca2+ in CP29 is bound in the domain including the lumenal loop between helices B and C.