Design, synthesis, and functional testing of recombinant cell penetrating peptides

The Use of Cell-penetrating Peptide for Delivery of Recombinant Transcription Factor DNA into Primary Human Fibroblast

“…This is in accordance with a previous study demonstrates that ALMR works more effectively on non-dividing cells and requires lipofectamine to increase transfection efficiency. 10 Fibroblasts are functionally and anatomically heterogeneous, determined by their function in different parts of the body. Fibroblasts from papillary cells are pro-regenerative, since they have the general characteristics of actively dividing cells.…”

“…Meanwhile, ALMR is a recombinant peptide that has been equipped with the NLS sequence (Rev), hence it can be recognized by cellular importers and facilitates the delivering system of extracellular DNA complexes into the nucleus via nuclear pore complex (NPC). 10 Rev is one of the HIV accessory proteins that has the ability to enter and leave the nucleus and bind to HIV mRNA (Figure 6). This is the reason why the ALMR system is more appropriate to be used in non-dividing cells (BRED) compared to PAL and CHO-K1 cells.…”

“…Then, the DNA will bind to histones and form a nucleoprotein structure resembling chromatin. 10 Based on ratio optimization, HNF4A showed an important role in the transfection of ALMR. HNF4A as a nuclear receptor contains a NLS, hence it may assist the process of recognizing nuclear signals.…”

“…Design, synthesis, and functional testing of ALMR was performed as described in previous studies. 10 ALMR peptide was obtained from culture stock of pQE80L-ALMR recombinant plasmid in E. coli DH5α (stored at -80°C). The culture stock were cultured with LB medium overnight (16-19 hours) and the protein were expressed in an optimized condition for isopropyl-β-Dthiogalactopyranoside (IPTG) induction.…”

“…It has been reported that ALMR can overcome cellular barriers, such as cell membranes, endosome compartments, and nuclear membranes, hence expression of transgene protein could occur even in non-dividing cells. 10 The aim of this study was to design the recombinant vector for the transfection of transcription factors and analyze the effectiveness of ALMR peptide for reprogramming human primary fibroblast to hepatocyte-like cells. ALMR peptide has the potential to substitute virus delivery systems in cell reprogramming, hence the safety of cell therapy can be more enforced.…”