Expression and Hydroxylamine Cleavage of Thymosin Alpha 1 Concatemer

• Published in: BioMed research international Vol. 2008; no. 2008; pp. 1 - 8
• Main Authors: Gong, Xing-Guo, Lu, Min-Kan, Lai, Zong-Teng, Zhou, Liang, Xie, Yi
• Format: Journal Article
• Language: English
• Published: Cairo, Egypt Hindawi Publishing Corporation 01.01.2008; John Wiley & Sons, Inc; Hindawi Limited
• Subjects: Amino Acid Sequence; Animals; Apoptosis; Cancer cells; Cell Line, Tumor; Cell Proliferation; Cells, Cultured; Deoxyribonucleic acid; DNA; E coli; Escherichia coli - genetics; Formazans - metabolism; Gene Expression - drug effects; Humans; Hydroxylamine - pharmacology; Inclusion Bodies - chemistry; Infections; Isopropyl Thiogalactoside - pharmacology; Life sciences; Lymphocytes - cytology; Lymphocytes - drug effects; Lymphocytes - physiology; Mice; Mice, Inbred BALB C; Mitochondria - drug effects; Mitochondria - physiology; Molecular Sequence Data; Molecular Weight; Peptide Fragments - chemistry; Peptide Fragments - genetics; Peptide Fragments - metabolism; Peptide Fragments - pharmacology; Peptides; Physiological aspects; Plasmids; Protein Structure, Secondary; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - metabolism; Tetrazolium Salts - metabolism; Thymosin; Thymosin - analogs & derivatives; Thymosin - chemical synthesis; Thymosin - chemistry; Thymosin - genetics; Thymosin - metabolism; Viral infections; China
• ISSN: 2314-6133; 1110-7243; 1110-7251; 2314-6141; 1110-7251
• DOI: 10.1155/2008/736060

Human thymosin alpha 1 (Tα1) is an important peptide in the development and senescence of immunological competence in human, and many studies have reported the expression of this peptide. In this study, we designed and synthesized the Tα1 gene according to the E. coli codon usage preference and constructed a 6×Tα1 concatemer. The latter was inserted into an E. coli expression vector pET-22b (+), and transformed into E. coli BL21 (DE3). After induction with IPTG, the concatemer protein was successfully expressed in E. coli then cleaved by hydroxylamine to release the Tα1 monomer. Gly-SDS-PAGE and mass spectrometry confirmed that the recombinant protein was cleaved as intended. The bioactivity of the Tα1 monomer was analyzed by lymphocyte proliferation and by mitochondrial activity in two different tumor cell lines. This study provides a description of the preparation of a bioactive Tα1, which may prove useful in future biomedical research.