Functionalized Peptide Fibrils as a Scaffold for Active Substances in Wound Healing

Abstract

Technological developments in the field of biologically active peptide applications in medicine have increased the need for new methods for peptide delivery. The disadvantage of peptides as drugs is their low biological stability. Recently, great attention has been paid to self-assembling peptides that can form fibrils. Such a formulation makes bioactive peptides more resistant to enzymatic degradation and druggable. Peptide fibrils can be carriers for peptides with interesting biological activities. These features open up prospects for using the peptide fibrils as long-acting drugs and are a valid alternative to conventional peptidic therapies. In our study, we designed new peptide scaffolds that are a hybrid of three interconnected amino acid sequences and are: pro-regenerative, cleavable by neutrophilic elastase, and fibril-forming. We intended to obtain peptides that are stable in the wound environment and that, when applied, would release a biologically active sequence. Our studies showed that the designed hybrid peptides show a high tendency toward regular fibril formation and are able to release the pro-regenerative sequence. Cytotoxicity studies showed that all the designed peptides were safe, did not cause cytotoxic effects and revealed a pro-regenerative potential in human fibroblast and keratinocyte cell lines. In vivo experiments in a dorsal skin injury model in mice indicated that two tested peptides moderately promote tissue repair in their free form. Our research proves that peptide fibrils can be a druggable form and a scaffold for active peptides.

Prediction of Aggregation of Biologically-Active Peptides with the UNRES Coarse-Grained Model

“…We used the experimental aggregation data of our previous work [ 27 , 31 , 32 ] of 13 peptides, while seven peptides were synthesized and investigated in this work. In all but one case (the 7–13 fragment of the Sup35 prion protein, with sequence H-GNNQQNY-NH ), UNRES correctly predicted the octamer dissociation temperatures to be above the experimental incubation temperature (310 K).…”

“…As shown in Table 1 , the data of 13 of the peptides studies were taken from our previous work [ 27 , 31 , 32 , 34 ]. Three peptides (Tau R2/wt, Tau R3/-P312 and Sup35(7-13) were studied by other researchers [ 28 , 29 , 33 ]; however, we synthesized them and performed measurements in this work.…”

“…The pure products were analyzed using analytical RP-HPLC and mass spectrometry (BIFLEX III MALDI-TOF, Bruker, Germany and LC-MS ESI-IT-TOF, Shimadzu, Japan). The detailed procedure of the synthesis steps, the concentrations of the coupling and deprotection reagents we used, as well as the purification conditions were described in our earlier work [ 31 ].…”

“…In this study, we used the physics-based UNRES coarse-grained model [ 23 , 24 , 25 , 26 ] developed in our laboratory to investigate the association of 16 peptides, whose aggregation has already been studied experimentally [ 27 , 28 , 29 , 30 , 31 , 32 , 33 , 34 ], as well as of four new peptides. These peptides have sizes ranging from 6 to 32 amino-acid residues and varying contents of hydrophobic, polar and charged residues.…”

“…The other ones contain potentially aggregating sequences merged with charged sequences with the intent of making functionalized gels or films or are signaling peptides [ 31 , 32 ]. Even though the aggressive coarse-graining has been implemented in UNRES (only two sites per residue), it is capable of ab initio modeling the structure, dynamics, thermodynamics and free-energy landscapes of small and medium-size proteins and bioninformatics- and data-assisted modeling of large proteins, including oligomeric proteins [ 23 , 26 ].…”