We have identified a Ca2+-binding site of the 29-kDa chlorophyll a/b-bindin
g protein CP29, a light harvesting protein of photosystem II most likely in
volved in photoregulation. Ca-45(2+) binding studies and dot blot analyses
of CP29 demonstrate that CP29 is a Ca2+-binding protein. The primary sequen
ce of CP29 does not exhibit an obvious Ca2+-binding site therefore we have
used Yb3+ replacement to analyze this site. Near-infrared Yb3+ vibronic sid
e band fluorescence spectroscopy (Roselli, C., Boussac, A. and Mattioli, T.
A. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 12897-12901) of Yb3+-reconst
ituted CP29 indicated a single population of Yb3+-binding sites rich in car
boxylic 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, theref
ore we propose that the Ca2+ in CP29 is bound in the domain including the l
umenal loop between helices B and C.