| Autor: |
Sabharwal P; Department of Biochemistry, Indian Institute of Science, Biological Sciences Building, Bangalore, 560012, India., Sushmitha C; Department of Biochemistry, Indian Institute of Science, Biological Sciences Building, Bangalore, 560012, India., Amritha CK; Department of Biochemistry, Indian Institute of Science, Biological Sciences Building, Bangalore, 560012, India., Natraj U; Department of Biochemistry, Indian Institute of Science, Biological Sciences Building, Bangalore, 560012, India., Murthy MRN; Molecular Biophysics Unit, Indian Institute of Science, Bangalore, 560012, India., Savithri HS; Department of Biochemistry, Indian Institute of Science, Biological Sciences Building, Bangalore, 560012, India. hss@alumni.iisc.ac.in. |
| Abstrakt: |
Monoclonal antibodies have attracted wide attention in therapeutics owing to their high efficacy, low toxicity, and specific targeting. However, antibodies cannot cross the cell membrane barrier. Therefore, their therapeutic potential is limited to surface-exposed antigens or secreted proteins. In the present investigation, we have developed a chimeric virus-like particle (VLP) of pepper vein banding virus (PVBV) and explored the possibility of using it as a delivery vehicle for antibodies against intracellular antigens as well as for future applications in immunodiagnostics. The chimeric PVBV particles were generated by genetically engineering the B domain of Staphylococcus aureus protein A (SpA) at the N-terminus of the PVBV coat protein (CP). The chimeric VLPs purified by sucrose density gradient centrifugation had ~440-fold higher affinity towards IgG antibody when compared to SpA. Interestingly, the unassembled chimeric CP with the B-domain at the N-terminus (BCP) purified by Ni-NTA chromatography was a monomer, and it had ~45-fold higher affinity towards antibodies compared to SpA. Additionally, the chimeric particles were able to bind and deliver antibodies against both intracellular (α-tubulin) and surface-exposed antigens (CD 20). However, the BCP monomer failed to enter mammalian cells. Thus, for the first time, we have demonstrated that the assembled VLPs are essential for internalization. These results demonstrate the potential of the use of chimeric PVBV VLPs in diagnostics and, more importantly, as nanocarriers for intracellular delivery of antibodies. |
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