Light-dependent fragmentation of the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase in chloroplasts isolated from wheat leaves.
The large subunit (LSU) of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco; EC 4.1.1.39) is degraded into an N-terminal side fragment of 37 kDa and a C-terminal side fragment of 16 kDa by the hydroxyl radical in the lysates of chloroplasts in light (H. Ishida et al. 1997, Plant Cell Physiol 38: 471-479). In the present study, we demonstrate that this fragmentation of the LSU also occurs in the same manner in intact chloroplasts, and discuss the mechanisms of the fragmentation. The fragmentation of the LSU was observed when intact chloroplasts from wheat leaves were incubated under illumination in the presence of KCN or NaN3, which is a potent inhibitor of active oxygenscavenging enzyme(s). The properties, such as molecular masses and cross-reactivities against the site-specific anti-LSU antibodies, of the fragments found in the chloroplasts were the same as those found in the lysates. These results indicate that, as in the lysates, the fragmentation of the LSU in the intact chloroplasts was also caused by the hydroxyl radical generated in light. The fragmentation of the LSU was completely inhibited by 3-(3',4'-dichlorophenyl)-1,1-dimethylurea (DCMU), and only partially inhibited by methyl viologen in the lysates. The addition of hydrogen peroxide to the lysates stimulated LSU fragmentation in light, but did not induce any fragmentation in darkness. Thus, we conclude that both production of hydrogen peroxide and generation of the reducing power at thylakoid membranes in light are essential requirements for fragmentation of the LSU.
Ishida
H
H
Department of Applied Biological Chemistry, Faculty of Agriculture, Tohoku University, Sendai, Japan.
Shimizu
S
S
Makino
A
A
Mae
T
T
D000906:Antibodies / Q000276:immunology; D002736:Chloroplasts / Q000201:enzymology / Q000528:radiation effects*; D003429:Cross Reactions; D008027:Light; D018515:Plant Leaves / Q000201:enzymology / Q000528:radiation effects*; D012273:Ribulose-Bisphosphate Carboxylase / Q000276:immunology / Q000378:metabolism*; D014908:Triticum / Q000201:enzymology / Q000378:metabolism*
D000906:Antibodies; D012273:Ribulose-Bisphosphate Carboxylase