Cloning of the Full-length Gene for Tobacco Ethylene Receptor NTHK2 and Characterization of Its Kinase Domain
ZHANG Zhi-Gang , GONG Yan , HE Xin-Jian, WANG Yu-Jun, SUN Zhong-Xu, ZHANG Jin-Song , CHEN Shou-Yi(1. Plant Biotechnology Laboratory, Institute of Genetics and Developmental Biology, The Chinese Academy of Sciences, Beijing 100101 , China;2. College of Horticulture, Shandong Agriculture University, Taian 271018, China; 3. College of Marine Life Sciences , Ocean University of Qingdao,Qindao 266003, China)
Previously the partial sequence of an ethylene receptor gene NTHK2 was isolated from tobacco ( Nicotiana tabacum L. var. Xanthi) plants and it was wound and drought inducible. In the present stundy full-length cDNA of NTHK2 was cloned by 5'-RACE method. NTHK2 gene has 3 216 bp, with 509 bp of 5'-non-coding region and 427 bp of 3'-non-coding region, and encodes an ethylene-receptor homolog of 760 amino acids. NTHK2 protein has a putative signal peptide, three transmembrane domains, a histidine kinase domain and a receiver domain. In the putative histidine kinase domain, the histidine at the phosphorylation s ite was replaced by an asparagine. To study the biochemical property of NTHK2, its kinase domain was expre ssed as a fusion protein with glutathione S-transferase (GST) using yeast Schizosaccharomyces pombe as an expression system. In vitro kinase assay showed that NTHK2 kinase domain can autophosphorylate in the presence of Mg2 + , indicating that NTHK2 may function as a kinase. Further studies will elucidate the function of NTHK2 in plant.