Please use this identifier to cite or link to this item: https://hdl.handle.net/10356/54302
Title: Designing endosomal escape ability on protein cage by viral mimicking mechanism.
Authors: Kuniyil Ambili.
Keywords: DRNTU::Engineering::Bioengineering
Issue Date: 2013
Abstract: Protein cages are gaining lot of interest in the field of drug delivery research. The E2 protein cage used in this study is a self- assembled structure of 60 identical subunits. It is derived from pyruvate dehydrogenase enzyme complex from Geobacillus Stearothermophilus. The structure can be multifunctionalized by modifying the subunits both genetically and chemically. The E2 protein cages can be internalized by cancer cells by the endosomal uptake through clathrin coated pits. In this study we have engineered 81 amino acid (34-114) peptide derived from the adenovirus capsid protein VI (P VI) on the N- terminus of E2 subunit. This recombinant protein is expected to escape from the endosome to deliver its contents. For the optimization of protein production, recombinant gene expression was carried out under different conditions: induction time ( 0.5, 1, 1.5, 2, 4& 16 hour), temperature (37°C, 20°C &30°C), IPTG concentrations (1mM, 2mM & multiple additions), expression vector (pET-11a & pET 28a) and expression systems (BL21(DE3) cells & BL21(DE3)pLysS cells). Results of these experiments suggested that the PVI protein retains its membrane lytic ability even after binding with the E2 protein by showing a marked reduction in the growth of cells after induction of protein production. Since it is a toxic protein it may be produced in very low amount and also there is presence of a prominent maintenance protein band in the expected region despite the inability to identify the protein production by SDS PAGE analysis.
URI: http://hdl.handle.net/10356/54302
Fulltext Permission: restricted
Fulltext Availability: With Fulltext
Appears in Collections:SCBE Theses

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