Human Tissue Plasminogen Activator Expression in Escherichia coli using Cytoplasmic and Periplasmic Cumulative Power.

Abstract : Tissue plasminogen activator (tPA) is a serine protease, which is composed of five distinct structural domains with 17 disulfide bonds, representing a model of high-disulfide proteins in human body. One of the most important limitations for high yield heterologous protein production in Escherichia coli (E. coli) is the expression of complex proteins with multiple disulfide bridges. In this study the combination of two distinct strategies, manipulated cytoplasm and native periplasm, was applied to produce the functional full length tPA enzyme in E. coli. Using a PelB signal peptide sequence at 5' site of tPA gene, the expression cassette was prepared and subsequently was transformed into a strain with manipulated oxidizing cytoplasm. Then the induction was made to express the protein of interest. The SDS-PAGE analysis and gelatin hydrolysis confirmed the successful expression of functional tPA. The results of this study showed that complex proteins can be produced in E. coli using the cumulative power of both cytoplasm and periplasm.
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Keivan Majidzadeh-A, Fereidoun Mahboudi, Mahdi Hemayatkar, Fatemeh Davami, Farzaneh Barkhordary, et al.. Human Tissue Plasminogen Activator Expression in Escherichia coli using Cytoplasmic and Periplasmic Cumulative Power.. Avicenna J Med Biotechnol, 2010, 2 (3), pp.131-6. ⟨pasteur-00829123⟩

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