Peptide–glycosaminoglycan cluster formation involving cell penetrating peptides

Abstract

Glycosaminoglycans (GAGs) affect the efficiency of cellular uptake of a wide range of cell penetrating peptides (CPPs). GAGs have been proposed to cluster with CPPs at the cell surface before uptake but little is known about the formation or stability of CPP-GAG clusters. Here we apply a combination of heparin affinity chromatography, dynamic light scattering, and fluorescence spectroscopy to characterize the formation, stability, and size of the clusters formed between CPPs and heparin. Under conditions similar to those used in cell uptake experiments the CPP, penetratin (Antp), was observed to form significantly more stable clusters with heparin than the CPP TAT, despite TAT showing a comparable affinity for heparin. This difference in cluster stability may explain the origins of the preferred cell uptake pathways followed by Antp and TAT, and may be an important parameter for optimizing the efficiency of designed CPP delivery vectors.

Tryptophan within basic peptide sequences triggers glycosaminoglycan‐dependent endocytosis

“…It should be noted that on addition of the peptides to HI, a fine precipitate formed in the solution. This might be due to the aggregation phenomena that take place with CPP‐HI binding, through formation of β‐sheet aggregates (13, 16, 17). Light scattering due to the particulate nature of the solution might have interfered with the CD experiment, but, nonetheless, the increase in ordered structure content came at the expense of other secondary structural features.…”

“…These peptide helices do not form stable and large aggregates as do Trp‐rich peptides. However, with a minimum number of 9 Arg residues, a level of internalization similar to that of a peptide containing 6 Arg and 3 Trp residues was recovered, suggesting that α‐helical peptides with a higher density of positive charges could also transiently but efficiently aggregate GAGs and internalize (17). Thus, Trp‐rich basic peptides are more prone than other CPPs to form stable clusters of GAGs, when GAG clustering is a key step for subsequent endocytosis (10, 51, 52).…”

A Fibronectin Peptide Redirects PDGF-BB/PDGFR Complexes to Macropinocytosis-Like Internalization and Augments PDGF-BB Survival Signals

“…To further characterize the P12 internalization pathway, P12-Alexa488 was incubated with cells at low temperature, with ATP depletion or chondroitin sulfate removal since internalization of highly cationic peptides by macropinocytosis requires active metabolism and interaction with cell surface GAGs (Rullo et al, 2011). Alexa488-P12 uptake was almost completely inhibited at 4°C or when cells were pre-incubated with sodium azide and deoxy-glucose to deplete cellular ATP, and greatly inhibited after digestion with chondrointinase ABC (Figure 2c and d).…”

Secondary conformational conversion is involved in glycosaminoglycans‐mediated cellular uptake of the cationic cell‐penetrating peptide PACAP

“…Cell surface GAGs were shown to interact with relatively high affinity with polycationic peptides [33][34][35][36] and to contribute significantly to the cellular uptake of CPPs [12][13][14][15]. It has been originally hypothesized that the contribution of GAGs in uptake is primarily related to the adsorption of CPPs to the outer leaflet of the plasma membrane through electrostatic interactions [37].…”