Functional characterization of the thylakoid Ndh complex phosphorylation by site-directed mutations in the ndhF gene.
To investigate the phosphorylation of the NDH-F subunit of the thylakoid Ndh complex, we constructed three site-directed mutant transgenic tobaccos (Nicotiana tabacum) (T181A, T181S and T181D) in which the (541)ACT(543) triplet encoding the Thr-181 has been substituted by GCT, TCT or GAT encoding alanine, serine and aspartic acid, respectively. Western blots with phospho-threonine antibody detected the 73 kD NDH-F phosphorylated polypeptide in control but not in mutant tobaccos. Differences in Ndh activity, chlorophyll fluorescence and photosynthesis among mutants and control plant demonstrate the key role of the phosphorylation of conserved Thr-181 in the activity and function of the Ndh complex. The substitution of aspartic acid for threonine in T181D mimics the presumable activation effects of the threonine phosphorylation in Ndh activity, post-illumination increase of chlorophyll fluorescence and photosynthesis rapid responses to changing light intensities. A tentative role of the phosphorylation-activated Ndh complex is suggested to poise the redox level and, consequently, optimizing the rate of cyclic electron transport under field conditions.
Martín
Mercedes
M
Departmento de Biología Vegetal. Universidad de Alcalá. Alcalá de Henares, 28871-Madrid, Spain.
Funk
Helena T
HT
Serrot
Patricia H
PH
Poltnigg
Peter
P
Sabater
Bartolomé
B
D000409:Alanine / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001224:Aspartic Acid / Q000378:metabolism; D002734:Chlorophyll / Q000378:metabolism; D017343:Genes, Plant; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D010788:Photosynthesis / Q000502:physiology; D030821:Plants, Genetically Modified; D017422:Sequence Analysis, DNA; D012694:Serine / Q000378:metabolism; D020524:Thylakoids / Q000235:genetics* / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism
D002734:Chlorophyll; D001224:Aspartic Acid; D012694:Serine; D000409:Alanine