D‐SAP: A New, Noncytotoxic, and Fully Protease Resistant Cell‐Penetrating Peptide

Abstract

Protease resistant cell-penetrating peptides (CPPs) are promising carriers for drugs unable to cross the cell membrane. As these CPPs are stable in vivo for much longer periods of time compared to other classes of therapeutic peptides, noncytotoxicity is a property sine qua non for their pharmacological development. Described herein is a fully protease resistant CPP that is noncytotoxic at concentrations up to 1 mM. Proteolytic stability was obtained by chiral inversion of the residues of a known self-assembling CPP-from all L-amino acids to all D-amino acids-and then assessed against trypsin and human serum. Circular dichroism studies confirmed the enantiomeric structure of the analogue, and transmission electron microscopy (TEM) studies indicated that the new inverso analogue retains the ability of the original peptide to self-assemble. The results of uptake experiments indicate that the protease-stable (that is, D-amino acid) analogue of the peptide is internalised by cells to the same extent as the protease-susceptible (that is, L-amino acid) parent peptide. Also reported herein are the results of studies on the cellular internalisation mechanism of the all-D analogue, which reveal the steps followed by the peptide upon its entry into the cell.

Bactenecin 7 peptide fragment as a tool for intracellular delivery of a phosphorescent oxygen sensor

“…Furthermore, this localization pattern is similar to that of the FITC-bactenecin 7 conjugate, which displayed a diffuse cytoplasmic distribution [5]. A cytoplasmic localization has also been reported for the other proline-rich d-SAP-FITC conjugate [6].…”

“…The mechanisms of cellular uptake may be cell type specific and may also be dependent on peptide length. The dependence on lipid raft and clathrin-mediated endocytosis was similar to that of the d-SAP peptide [6]. Collectively, our data indicate that the PEPP0 conjugate employs at least two different mechanisms of cellular uptake: temperature-and ATP-independent direct translocation for PC12 cells, and clathrin and lipid raft-dependent endocytic pathways for PC12 and HCT116 cells.…”

“…This polypeptide is 59 amino acids in length and possesses repeated sequences which form regions with both antimicrobial and cell-penetrating properties [6]. Interestingly, the region (15-24) harbours a cell-penetrating ability in the absence of antimicrobial ability, and does not influence the function of mammalian cells (e.g.…”

Proteolysis of Peptide Dendrimers

“…MS calcd/found [b] Trypsin a-Chymotrypsin www.chembiochem.org ral amino acids, such as b-amino acids [23] or d-amino acids, [24] as is often done for linear peptides. For instance the prevalence of single cleavages of the dendrimers when the proteolytic site was placed directly at a branching point probably indicates the reduced reactivity of the side-chain amide group.…”

Dimeric Cationic Amphiphilic Polyproline Helices for Mitochondrial Targeting

“…Generally, CPPs are described as a class of short peptide sequences that are rich in basic amino acids and able to deliver a variety of bioactive molecules across membranes of different cell lines with high efficiency and minimal toxicity (1). Some CPPs also feature an alternating pattern of charged, polar amino acids and hydrophobic, non-polar amino acids, rendering an overall amphiphilic peptide (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12). CPPs have been found to deliver a range of cargo into cells, from proteins and oligonucleotides to magnetic nanoparticles (13)(14)(15)(16).…”