Pustaka Ilmiah Universitas Padjadjaran

Abstrak

Extracellular Secretion of Recombinant Human Epidermal Growth Factor by Using Trimethylamine N-Oxide Reductase A (TORA) Signal Peptide in Escherichia Coli BL21 (DE3)

Human Epidermal Growth Factor (hEGF) is a protein that helps in the healing process of wounds, in the process of proliferation, migration and cell differentiation. hEGF composed by 53 amino acid residues, with a molecular weight of 6.2 kDa and has a single chain conformation, and resistant to high temperatures up to 70°C. hEGF is already very widely produced in an industrial scale using Escherichia coli as a host cell, but the majority of hEGF recombinant protein is produced intracellularly, so it has a lot of disadvantages such as; the accumulation of non-functional protein, the amount of protein inclusion bodies are formed as a result of misfolding, and the high rate of degradation due to intracellular protease enzymes. Therefore the purpose of this study was to express recombinant proteins in the extracellular hEGF using a signal peptide TorA, via the Twin Arginine translocation (TAT) pathway, measure the levels of hEGF recombinant protein secreted into the culture medium with the use of a signal peptide TorA via TAT. This research using Escherichia coli BL21 as the host cell to be transformed by using recombinant plasmid pD881-TorA the consensus already containing the signal peptide TorA and also hEGF gene, then expressed by L-rhamnose induction, then the recombinant protein hEGF purified by heat treatment and then the results were characterized using SDS-PAGE, and ELISA. The result is hEGF protein can be secreted into culture medium and periplasm with concentration 0.9625 µg/ mL.