Background and Aim: The HIV - 1 regulatory proteins tat and nef are attractive as vaccine components since they are the HIV - 1 critical proteins and AIDS disease progression . For vaccine development, different types of lipid - and polymer - based vectors such as liposomes, microparticles and nanoparticles have been used for protein delivery, but with relatively poor success. Recently, a group of short, highly basic peptides known as cell - penetrating peptides (CPPs) were used to carry polypeptides and proteins into cells.
Methods: : In current study, expression and purification of Tat - Nef protein was performed in E. coli and transfected by the amphipathic CPPs including Pep - 1 and CADY - 2 into HEK - 293T cells. The size and morphology of the CPPs/ Tat - Nef complexes were evaluated by scanning electron microscopy (SEM) and SDS - PAGE. The efficiency of transfection of Tat - Nef/CPP nanoparticles was analyzed and compared with FITC - antibody protein as a control ,using western blotting
Results: SEM data confirmed the formation of discrete nanoparticles with a diameter of below 300 nm. Moreover, formation of the complexes was detected using SDS - PAGE as two individual bands indicating the non - covalent interaction. the Tat - Nef protein was expressed in prokaryotic expression system and efficiently transfected using CPP delivery systems at molar ratio of 20:1. Our results showed that the transfection efficiency of Pep - based nanoparticles was similar to CADY - based nanoparticles and comparable with Turbo Fect - protein complexes.
Conclusion: The in vitro efficient delivery of Tat - Nef fusion protein supports the potential of CPPs as potent carriers for HIV vaccine design and it will be considered as an advantage for therapeutic purposes in Future.