Overexpression of Nicotiana tabacum acetolactate synthase as an inducible fusion protein in Escherichia coli: Production of a polyclonal antibody to Nicotiana tabacum acetolactate synthase

Chang, S.-Ik; Kang, M.-Kyeong; Kim, H.-Ju; Choi, J.-Do; Namgoong, S.-Keon

Journal of Biochemistry and Molecular Biology 29(5): 462-467

1996


ISSN/ISBN: 1225-8687
Accession: 009144684

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Abstract
Acetolactate synthase (ALS, EC 4.1.3.18) is the first common enzyme in the biosynthesis of leucine, isoleucine, and valine. It is the target enzyme for several classes of herbicides, including the sulfonylureas, the imidazolinones, the triazolopyrimidines, the pyrimidyl-oxy-benzoates, the Pyrimidyl-thio-benzens, and the 4,6-dimethoxypyrimidines. An amino-terminal fragment of the sulfonylurea-resistant ALS gene (SurB) from Nicotiana tabaccum was cloned into the bacterial expression vector pGEX-2T. The resulting recombinant plasmid pGEX-ALS1 was used to transform Escherichia coli strain BL21, and the tobacco ALS was expressed in the bacteria as a protein fused with glutathione S-transferase (GST). Polyclonal antibodies against the fusion product (GST-ALS) were produced, and the sensitivity of GST-ALS with the rabbit anti-GST-ALS IgG was up to 50 ng. This antibody was used for Western blot analysis of the partially purified ALS from barley shoots. The results suggest that the polygonal antibody produced in this study can be used to detect plant ALS.