The chloroplastic glutamine synthetase (GS-2) of tobacco is phosphorylated and associated with 14-3-3 proteins inside the chloroplast.
The chloroplastic isoform of glutamine synthetase (GS-2, EC 6.3.1.2) from Nicotiana tabacum L. is phosphorylated at the serine residues. At least three of the six GS-2 subunits separated by two-dimensional polyacrylamide gel electrophoresis cross-reacted with an antibody raised against phosphoserine. This provoked the question as to whether 14-3-3 proteins might be present in the chloroplast and bind to chloroplastic GS-2. Although two different 14-3-3 proteins of 32 and 30 kDa were present in total leaf extracts, in the soluble fraction of chloroplasts, only the 32-kDa 14-3-3 protein was immunodetected with an antibody raised against a conserved region of 14-3-3 protein from corn. This demonstrates the presence of a chloroplast-located isoform of 14-3-3 proteins in tobacco. To examine a putative binding of GS-2 to these 14-3-3 proteins in vivo, the native GS-2 holoenzyme was probed with a 14-3-3 antibody. The strong cross-reaction between GS-2 and the 14-3-3 antibody clearly points to a binding of GS-2 and 14-3-3 in tobacco chloroplasts. Only those oligomers of GS-2 that were strongly associated with 14-3-3 proteins were catalytically active.
Riedel
J
J
Albrecht-von-Haller-Institut für Pflanzenwissenschaften, Göttingen, Germany.
Tischner
R
R
Mäck
G
G
D048948:14-3-3 Proteins; D002736:Chloroplasts / Q000201:enzymology / Q000378:metabolism*; D005974:Glutamate-Ammonia Ligase / Q000378:metabolism*; D015151:Immunoblotting; D008970:Molecular Weight; D009565:Nitrate Reductases / Q000378:metabolism; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D010940:Plant Proteins / Q000378:metabolism*; D011485:Protein Binding; D020033:Protein Isoforms; D014026:Nicotiana / Q000201:enzymology* / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism*
D048948:14-3-3 Proteins; D010940:Plant Proteins; D020033:Protein Isoforms; D010768:Phosphoserine; D014446:Tyrosine 3-Monooxygenase; D009565:Nitrate Reductases; C083818:glutamine synthetase 2; D005974:Glutamate-Ammonia Ligase