Interaction of CPa-1 with the manganese-stabilizing protein of photosystem II: identification of domains on CPa-1 which are shielded from N-hydroxysuccinimide biotinylation by the manganese-stabilizing protein.
The structural organization of photosystem II proteins has been investigated by use of the amino group-labeling reagent N-hydroxysuccinimidobiotin (NHS-biotin) and calcium chloride-washed photosystem II membranes. We have previously shown that the presence of the extrinsic, manganese-stabilizing protein on photosystem II membranes prevents the modification of lysyl residues located on the chlorophyll protein CPa-1 (CP-47) by NHS-biotin [Bricker, T. M., Odom, W. R., & Queirolo, C. B. (1988) FEBS Lett. 231, 111-117]. Upon removal of the manganese-stabilizing protein by calcium chloride-washing, CPa-1 can be specifically modified by treatment with NHS-biotin. Preparative quantities of biotinylated CPa-1 were subjected to chemical cleavage with cyanogen bromide. Two major biotinylated peptides were identified with apparent molecular masses of 11.8 and 15.7 kDa. N-terminal sequence analysis of these peptides indicated that the 11.8-kDa peptide was 232G-330M and that the 15.7-kDa peptide was 360P-508V. The 15.7-kDa CNBr peptide was subjected to limited tryptic digestion. The two smallest tryptic fragments identified migrated at apparent molecular masses of 9.1 (nonbiotinylated) and 7.5 kDa (biotinylated). N-terminal sequence analysis and examination of the predicted amino acid sequences of these peptides suggest that the 9.1-kDa fragment was 422R-508V and that the 7.5-kDa fragment was 360P-421A. These results strongly suggest that two NHS-biotinylated domains, 304K-321K and 389K-419K, become exposed on CPa-1 when the manganese-stabilizing protein is removed by CaCl2 treatment. Both of these domains lie in the large extrinsic loop E of CPa-1.
Frankel
L K
LK
Department of Botany, Louisiana State University, Baton Rouge 70803.
Bricker
T M
TM
D000595:Amino Acid Sequence; D001710:Biotin / Q000031:analogs & derivatives* / Q000737:chemistry; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D045342:Light-Harvesting Protein Complexes; D008345:Manganese / Q000378:metabolism*; D008969:Molecular Sequence Data; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry*; D045332:Photosystem II Protein Complex; D010940:Plant Proteins / Q000378:metabolism*; D011487:Protein Conformation; D013388:Succinimides / Q000737:chemistry*
D045342:Light-Harvesting Protein Complexes; D045322:Photosynthetic Reaction Center Complex Proteins; D045332:Photosystem II Protein Complex; D010940:Plant Proteins; D013388:Succinimides; C033788:biotinyl N-hydroxysuccinimide ester; C070970:photosystem II, chlorophyll-binding protein, CP-47; D008345:Manganese; D001710:Biotin