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Synthesis and proof‐of‐function of a [ 14 C]‐labelled form of the plant iron chelator nicotianamine using recombinant nicotianamine synthase from barley
Author(s) -
Schmiedeberg Lars,
Krüger Claudia,
Stephan Udo W.,
Bäumlein Helmut,
Hell Rüdiger
Publication year - 2003
Publication title -
physiologia plantarum
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.351
H-Index - 146
eISSN - 1399-3054
pISSN - 0031-9317
DOI - 10.1034/j.1399-3054.2003.00128.x
Subject(s) - chemistry , biochemistry , atp synthase , complementation , lycopersicon , chelation , micronutrient , mutant , botany , biology , enzyme , organic chemistry , gene
The amino acid nicotianamine (NA) is essential for micronutrient metabolism in plants. Lack of NA results in a chlorotic phenotype and oxidative stress, since NA is a chelator of iron and other metal nutrients. To investigate the precise cellular function of NA in micronutrient transport and homeostasis, a protocol for the production of [ 14 C]‐labelled NA was developed. Recombinant NA synthase was used to generate [ 14 C]‐NA from [ 14 C]‐ S ‐adenosylmethionine. After purification by solid‐phase ion exchange about 66% yield was achieved. The identity of the [ 14 C]‐NA with chemically synthesized NA was demonstrated by several independent methods, including two TLC systems, two HPLC systems and immuno‐detection. Moreover, biological function was shown by complementation of the Lycopersicon esculentum mutant chloronerva that is free of NA due to a defect in NA synthase. Proof‐of‐function for the produced [ 14 C]‐NA as a suitable tool for transport studies was provided monitoring the distribution of [ 14 C]‐NA after feeding to tomato and Ricinus communis seedlings.