Expression, Purification, Identification And Biological Activity Of Human Thymosin Beta 4 Concatemer

Thymosin beta 4 (Tβ₄) is originally isolated from the thymus gland. It consists of 43-amino acid residues and is found in high concentration in blood platelets, wound fluid, white blood cells, and in most other tissues of the body. Tβ₄ is neither a growth factor nor a cytokine. Rather, Tβ₄ is the major actin regulating molecule in most mammalian cells and plays an important role in the regeneration, remodeling, and healing of injured or damaged tissues. Independent researchers have shown that Tβ₄ is multi-faceted and involved in a wide variety of biological activities including: regulation of actin, epithelialization, angiogenesis, and anti-inflammation, so it can promote wound healing and treat septic shock with MSOF and plays a significant role in the military medicine.Although Tβ₄ is likely to be used widely, it cannot be expressed in E.coli directly because of its low molecular weight. Furthermore, Tβ₄ used in the laboratory investigation and clinical trials now is mainly produced synthetically with a high cost, meanwhile, isolating Tβ₄ from the mammalian tissue is a complex technics and with low yield, so the application of Tβ₄ is restricted. How to get the polypeptide economically is the problem that we try to solve.In the study, we synthesize Tβ₄ gene using E.coli preferred codon and construct expression vector pET-22b(+)-Tβ₄â‘¡, then transform recombinant plasmid to E.coli BL21(DE3) and induced with IPTG. The expressed protein Tβ₄â‘¡can be purified by salting-out, hydrophobic chromatography and ion exchange chromatography and identified by Western blot and MALDI-TOF-MS. The purified Tβ₄â‘¡concatemer is able to promote lymphocyte differentiation and proliferation as effectively as synthetic one, and has weak immunogenicity detected by ELISA.The purified Tβ₄â‘¡concatemer has established a foundation for its function and application. It will be a new first-aid medicine in the war one day.