Synthesis of N-acyl derivatives of Pro-Gly-Pro-Leu peptide: Proteolytic stability in vitro and effects on mouse macrophage cells RAW264.7

Abstract

Acetyl, oleoyl, arachidonoyl, and docosahexaenoyl derivatives of the Pro-Gly-Pro-Leu peptide with a chemical purity of 99.8% were synthesized. The degradation kinetics of the Pro-Gly-Pro-Leu derivatives under the action of leucine aminopeptidase, nasal mucus, and microsomal fraction of the brain and blood of rats was studied. It was shown that the N-acyl derivatives of Pro-Gly-Pro-Leu proved to be more resistant to the action of leucine aminopeptidase and other enzyme systems. The study of the cytotoxic and anti-inflammatory activity of preparations on the mouse macrophage cell line RAW264.7 showed that acylation with oleic and arachidonic acid makes the peptide cytotoxic with LC50 in the range of 70-15 μM and gives it anti-inflammatory properties with EC50 of 32 and 36 μM, respectively.

Short peptide sequences: current knowledge and future prospects

“…Studies of the tetrapeptide Lys-Glu-Аsр-Ala showed that this compound was not hydrolyzed even by the peptidases of the small intestine and under the conditions in vitro it was hydrolyzed only for 50% [23]. As mentioned above, different glyprolines were reported to modulate the activity of gastrointestinal enzymes along with high stability to enzymatic hydrolysis [2,21,[24][25][26]. The pentadecapeptide BPC 157 (Gly-Glu-Pro-Pro-Pro-Gly-Lys-Pro-Ala-Asp-Asp-Ala-Gly-Leu-Val) was also reported to be especially stable and in vitro studies showed that this compound was not broken down by gastric juice during a 24 hours period [27].…”

“…In vitro studies of the hydrolysis of 2 peptides of the structure Lys-Glu-Asp-Ala (livagen) and epithalon (Ala-Glu-Asp-Gly) in different parts of gastrointestinal tract showed that Lys-Glu-Asp-Ala was unsignificantly hydrolysed in stomach and both Lys-Glu-Asp-Ala and Ala-Glu-Asp-Gly were resistant to proteases of the proximal part of the small intestine and jejunum [22]. N-acyl derivatives of Pro-Gly-Pro-Leu proved to be resistant to the action of leucine aminopeptidase and other enzyme systems [25].…”

“…Pinocytosis was presumed to be the main mechanism how short sequences enter the cell [25]. For a long time it was unclear whether short peptides penetrate the cytoplasmic and nuclear membrane and then bind to DNA or they bind to receptors (intracellular or on cell membranes) and activate the intracel-lular signal cascades, which results in modification of the gene expression [3,14].…”

“…[21] Mechanism of cell penetration Pinocytosis was presumed to be the main mechanism how short sequences enter the cell [25] Biological activity of some short peptide sequences is mediated by their specific ligand-receptor interaction on the plasma membranes [21] Mechanism of action Short peptides can penetrate both the cytoplasm and nucleus, bind to complementary sequences in the gene promoters, affecting the processes of transcription and translation…”