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Cloning, purification, and polymerization of Capsicum annuum recombinant α and β tubulin
- Title
- Cloning, purification, and polymerization of Capsicum annuum recombinant α and β tubulin
- Authors
- Jang M.-H.; Kim J.; Kalme S.; Han J.-W.; Yoo H.-S.; Koo B.-S.; Kim S.-K.; Yoon M.-Y.
- Ewha Authors
- 김진흥
- SCOPUS Author ID
- 김진흥
- Issue Date
- 2008
- Journal Title
- Bioscience, Biotechnology and Biochemistry
- ISSN
- 0916-8451
- Citation
- Bioscience, Biotechnology and Biochemistry vol. 72, no. 4, pp. 1048 - 1055
- Indexed
- SCIE; SCOPUS
- Document Type
- Article
- Abstract
- α and β tubulin genes were cloned from the Capsicum annuum leaves using rapid amplification of cDNA ends (RACE)-PCR. Nucleotide sequence analysis revealed that 1,353 bp Capsicum annuum α/β-tubulin (CAnm α/β-TUB) encodes a protein of 450 amino acids (aa) each. The recombinant α/β tubulin was overexpressed mainly as an inclusion body in Escherichia coli BL21 (DE3), upon induction with 0.2 mM isopropyl-β-D- thiogalactopyranoside (IPTG), and its content was as high as 50% of the total protein content. Effective fusion protein purification and refolding are described. The average yields of α and β tubulin were 2.0 and 1.3 mg/l of culture respectively. The apparent molecular weight of each tubulin was estimated to be 55 kDa by SDS-polyacrylamide gel electrophoresis (PAGE). The tubulin monomers were found to be assembly competent using a standard dimerization assay, and also retained antigenicity with anti-His/T7 antibodies. The purified tubulins were polymerized to microtubule-like structures in the presence of 2 mM guanosine 5′-triphosphate (GTP).
- DOI
- 10.1271/bbb.70794
- Appears in Collections:
- 자연과학대학 > 화학·나노과학전공 > Journal papers
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