Identification of zinc binding proteins of wheat seed
Gökçe, Onur (2007) Identification of zinc binding proteins of wheat seed. [Thesis]
Zinc content of the wheat seed has significance for agronomy, environment and also for human health. Seeds containing higher amounts of micronutrients, especially zinc, are better at germination and show high resistance to pests. Experiments on the mechanisms of micronutrient uptake and their accumulation in the plant body and the seeds showed a correlation between zinc and protein amount in the seed, which suggested some proteins, mostly abundant in the embryo and the aleurone layer, might be sinks for zinc. There is a lack of knowledge on zinc binding proteins of wheat seed. In this project, we aimed to determine the effect of environmental conditions on the protein and micronutrient content of the durum wheat seed and to screen the proteome of the wheat seed to identify the proteins with metalin particular Zn-binding propensity. Protein analyses were carried out on seeds with extreme N and Zn content. Extraction methods available in the literature were modified to obtain reproducible profiles of the total protein content and to separate seed proteins into fractions of different storage protein classes. Extracts were analyzed using polyacrylamide gel electrophoresis and a procedure for detection of Zn-binding proteins was developed. Results showed Zn-binding by Cys rich proteins in the molecular mass range 30 to 50 kDa. The method was extensively tested to confirm the association of Zn with specific proteins, however, if those proteins bind metal in vivo remains to be shown. Microscopic analysis of in situ staining for protein and Zn localization showed preferential staining in the embryo and aleurone layers of the seed and led to determination of the protein profiles from embryo and endosperm tissues, separately. The methods developed during this and further studies, shall be used to do the same screening for different wild-type and cultivated genotypes of wheat.
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