In vivo fragmentation of the large subunit of ribulose-1,5-bisphosphate carboxylase by reactive oxygen species in an intact leaf of cucumber under chilling-light conditions.
Previous studies have demonstrated that the large subunit (LSU) of ribulose-1,5-bisphosphate carboxylase (Rubisco) is site-specifically cleaved by a hydroxyl radical (*OH) generated in the illuminated chloroplast lysates or by an artificial *OH-generating system. However, it is not known whether such cleavage of the LSU by reactive oxygen species (ROS) actually occurs in an intact leaf. When leaf discs of chilling-sensitive cucumber (Cucumis sativus L.) were illuminated at 4 degrees C, five major fragments of the LSU were observed. This fragmentation was completely inhibited by ROS scavengers, such as n-propyl gallate (for *OH) and 1,2-dihydroxybenzene-3,5-disulfonic acid (Tiron) (for superoxide). FeSO4 stimulated this fragmentation, whereas an iron-specific chelator, deferoxamine, suppressed it. Furthermore, such fragments were identical to those generated from the purified Rubisco by an *OH-generating system in vitro on two-dimensional PAGE. These results indicate that the direct fragmentation of the LSU by reactive oxygen species also occurs in an intact leaf.
Nakano
Ryouhei
R
Department of Applied Plant Science, Graduate School of Agricultural Science, Tohoku University, Tsutsumidori-Amamiyamachi, Sendai, 981-8555 Japan.
Ishida
Hiroyuki
H
Makino
Amane
A
Mae
Tadahiko
T
D014013:1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt / Q000494:pharmacology; D003080:Cold Temperature; D018553:Cucumis sativus / Q000201:enzymology*; D003676:Deferoxamine / Q000494:pharmacology; D015180:Electrophoresis, Gel, Two-Dimensional; D005290:Ferric Compounds / Q000494:pharmacology; D008027:Light; D010446:Peptide Fragments / Q000032:analysis; D018515:Plant Leaves / Q000201:enzymology*; D011435:Propyl Gallate / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012273:Ribulose-Bisphosphate Carboxylase / Q000737:chemistry / Q000378:metabolism*; D013696:Temperature
D005290:Ferric Compounds; D010446:Peptide Fragments; D017382:Reactive Oxygen Species; C024823:ferric sulfate; D014013:1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; D011435:Propyl Gallate; D012273:Ribulose-Bisphosphate Carboxylase; D003676:Deferoxamine