Cell-penetrating peptide chains

Fig. 1. Different strategies of coupling cargoes to cell-penetrating peptides. (I) Non-covalent coupling by electrostatic interactions (A) (44,46), specific pairing of nucleotides (B) (39,48), biotin-avidin interaction (C) (37,66) or mixed-type interactions (D) (26). (II) Covalent coupling by connecting CPP and cargo molecule into a continuous chain (E) (36), forming a disulfide bond (F) (49,56) or chemical crosslinking (G) (37,57).

Figure 7.7-3. Cell penetrating peptides, CPPs. Suggested uptake mechanisms for CPPs and examples of delivered cargoes. (A) CPP and peptide in single amino acid chain. (B) Oligo-deoxynucleotides either in complex or covalently linked. (C) Plasmid in complex by electrostatic interaction. (D) Protein either as fusion protein or in complex with CPP. (E) siRNA, covalently linked or as complex. (With permission from Ref. 54.)...

Brase, Muhle-Goll and coworkers reported on functional peptoid helices [77]. In order to create cell-penetrating peptoids, butylamine residues were incorporated into peptoids. NMR spectroscopy and molecular modeling revealed an extended pseudo-helical structure with predominant c/s-conformation in the backbone. The electrostatic repulsion between the side chains leads to maximization of the spacing of the ammonium residues and thus dominates the conformation of the peptoid. With a pitch of 7.7 A, the charge distribution is somewhat less dense compared to a-helical peptides. Nevertheless, the peptoids were effective transporters, similar to other cell-penetrating peptides. 

Since its discovery, isolation, and purification in the early twentieth century, insulin has been administered to diabetic patients exclusively by injection until the recent introduction of inhaled insulin. Insulin possesses certain physiochemical properties that contribute to its limited absorption from the gastrointestinal tract, and requires subcutaneous injection to achieve clinically relevant bioavailability. With a molecular size of 5.7 kDa, insulin is a moderately sized polypeptide composed of two distinct peptide chains designated the A chain (21 amino acid residues) and the B chain (30 amino acid residues) and joined by two disulfide bonds. Like all polypeptides, insulin is a charged molecule that cannot easily penetrate the phospholipid membrane of the epithelial cells that line the nasal cavity. Furthermore, insulin monomers self-associate into hexameric units with a molecular mass greater than 30 kDa, which can further limit its passive absorption. Despite these constraints, successful delivery of insulin via the nasal route has been reported in humans and animals when an absorption enhancer was added to the formulation. 

The major difference between peptides and peptoids lies in the fact that the side chain substituents R -R in peptoids are attached to the amide nitrogens, thus avoiding the chiral a-centres of a peptide. There are no NH-bonds present in peptoids which has major implications on solubility, cell penetration properties and on the overall conformations which differ significantly from those of peptides. In addition, compared to peptides, peptoids show an increased protease stability due to the tertiary amide bond. 

The major stmctural feature of the HAz chain (blue in Figure 5.20) is a hairpin loop of two a helices packed together. The second a helix is 50 amino acids long and reaches back 76 A toward the membrane. At the bottom of the stem there is a i sheet of five antiparallel strands. The central i strand is from HAi, and this is flanked on both sides by hairpin loops from HAz. About 20 residues at the amino terminal end of HAz are associated with the activity by which the vims penetrates the host cell membrane to initiate infection. This region, which is quite hydrophobic, is called the fusion peptide. 

There are many examples of peptides that cannot form transmembrane channels on their own but can do so through aggregation. The gramicidin antibiotics, produced by bacteria as part of their chemical defence system, are only 1.6 nm or so in length [11], Specific placement of side chains, such as four tryptophan residues towards the C-terminus, ensures that the helix penetrates cell membranes to a particular depth but does not pass through the membrane. 

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