Cloning and expression of Bombyx mori silk gland elongation factor 1gamma in Escherichia coli |
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Authors: | Kamiie Katsuyoshi Nomura Yoshitaka Kobayashi Satoru Taira Hideharu Kobayashi Kohmei Yamashita Tetsuro Kidou Shin-ichiro Ejiri Shin-ichiro |
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Affiliation: | Department of Bioscience and Biotechnology, Faculty of Engineering, Aomori University, Japan. kamiie@aomori-u.ac.jp |
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Abstract: | Elongation factor 1 (EF-1) from the silk gland of Bombyx mori consists of alpha-, beta-, gamma-, and delta-subunits. EF-1alpha GTP catalyzes the binding of aminoacyl-tRNA to ribosomes concomitant with the hydrolysis of GTP. EF-1betagammadelta catalyzes the exchange of EF-1alpha-bound GDP for exogenous GTP and stimulates the EF-1alpha-dependent binding of aminoacyl-tRNA to ribosomes. EF-1gamma cDNA, which contains an open reading frame (ORF) encoding a polypeptide of 423 amino acid residues, was amplified and cloned by PCR from a silk gland cDNA library. The calculated molecular mass and predicted pI of the product were 48,388 Da and 5.84, respectively. The silk gland EF-1gamma shares 67.3% amino acid identity with Artemia salina EF-lgamma. The N-terminal domain (amino acid residues 1-211) of silk gland EF-lgamma is 29.3% identical to maize glutathione S-transferase. We demonstrated that silk gland EF-lgamma bound to glutathione Sepharose, suggesting that the N-terminal domain of EF-1gamma may have the capacity to bind to glutathione. |
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