Isolation of phosphorylated and dephosphorylated forms of the CP43 internal antenna of photosystem II in Hordeum vulgare L.
As a consequence of variation in environmental factors, light being the most important one, a number of photosystem II polypeptides may be reversibly phosphorylated by thylakoid-bound kinase(s). Among them, the reaction centre D1 and D2 polypeptides, the PsbH subunit, and the inner antenna CP43. Here, the separation of two forms of CP43 by high-resolution denaturing polyacrylamide gel electrophoresis is reported. By means of immunoblotting with antibody to phosphothreonine-containing proteins and authentic CP43 and limited proteolysis, these two bands could be identified as the phosphorylated and dephosphorylated forms of CP43. Using non-denaturing isoelectrofocusing, a chromatographically derived CP43-enriched fraction could be resolved into three different native forms of CP43. Among them, one was found to be a phosphorylated form, whereas the other two were dephosphorylated forms of the protein. With respect to other methods, the procedure described here allows the isolation, for the first time, of a fully homogeneous population of this chlorophyll-protein complex, opening the way to the study of the role of phopshorylation on functional properties of this core antenna protein.
Andreucci
Flora
F
Dipartimento di Scienze Ambientali e della Vita, Università del Piemonte Orientale Amedeo Avogadro, Via Bellini 25/G, I-15100 Alessandria, Italy.
Barbato
Roberto
R
Picollo
Chiara
C
Segalla
Anna
A
D001467:Hordeum / Q000378:metabolism*; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D045322:Photosynthetic Reaction Center Complex Proteins / Q000302:isolation & purification / Q000378:metabolism*; D045332:Photosystem II Protein Complex / Q000302:isolation & purification / Q000378:metabolism*; D013053:Spectrophotometry
D045322:Photosynthetic Reaction Center Complex Proteins; D045332:Photosystem II Protein Complex; C083100:photosystem II, chlorophyll binding protein, CP-43; D010770:Phosphotransferases