title: Dual selective nitration in Arabidopsis: Almost exclusive nitration of PsbO and PsbP, and highly susceptible nitration of four non-PSII proteins, including peroxiredoxin II E.
abstract: Protein tyrosine nitration is a selective process, as revealed in studies of animals. However, evidence for selective protein nitration in plants is scarce. In this study, Arabidopsis plants were exposed to air with or without nitrogen dioxide at 40 ppm for 8 h in light. Proteins extracted from whole leaves or isolated chloroplasts were subjected to 2D PAGE followed by SYPRO Ruby staining and immunoblotting using an anti-3-nitrotyrosine antibody. We determined the relative intensity of a spot on an immunoblot (designated RISI), and relative intensity of the corresponding spot on SYPRO Ruby gel (designated RISS). Proteins that exhibited a high RISI value and/or a high RISI/RISS ratio were considered selectively nitrated. In whole leaf proteins from exposed plants, all immunopositive spots were identified as PsbO1, PsbO2 or PsbP1 by PMF. Thus, nitration was exclusive to PsbO and PsbP, extrinsic proteins of photosystem II (PSII). Their RISI/RISS ratio was ≤1.5. Non-exposed plants showed very faint nitration. In purified chloroplast proteins, PsbO and PsbP accounted for >80% of the total RISI values, while four non-PSII proteins, including peroxiredoxin II E, exhibited high RISI/RISS ratios (2.5∼6.6). Tyr(9) of PsbO1 was identified as a nitration site. Thus, nitration is selective for two PSII and four non-PSII proteins in Arabidopsis.
authors.affiliation: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Hiroshima, Japan.
authors.forename: Misa
authors.initials: M
authors.lastname: Takahashi
authors.affiliation: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Hiroshima, Japan.
authors.forename: Jun
authors.initials: J
authors.lastname: Shigeto
authors.affiliation: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Hiroshima, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Sakamoto
authors.affiliation: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Hiroshima, Japan.
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Izumi
authors.affiliation: Faculty of Engineering, Fukuyama University, Fukuyama, Japan.
authors.forename: Kozi
authors.initials: K
authors.lastname: Asada
authors.affiliation: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Hiroshima, Japan.
authors.forename: Hiromichi
authors.initials: H
authors.lastname: Morikawa
mesh_terms: D029681:Arabidopsis Proteins / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism*; D060365:Chloroplast Proteins; D015180:Electrophoresis, Gel, Two-Dimensional; D009574:Nitro Compounds / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism*; D045332:Photosystem II Protein Complex / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014443:Tyrosine / Q000031:analogs & derivatives / Q000032:analysis / Q000737:chemistry / Q000378:metabolism
chemical_list: D029681:Arabidopsis Proteins; D060365:Chloroplast Proteins; D009574:Nitro Compounds; D045332:Photosystem II Protein Complex; C002744:3-nitrotyrosine; D014443:Tyrosine
keywords: Arabidopsis; Peroxiredoxin II E; Protein tyrosine nitration; PsbO; PsbP