Cell-Modulating Effect of Poly(Aspartic Acid) and Its Complex with Cationic Polyaspartamide

© 2020, Springer Science+Business Media, LLC, part of Springer Nature. In this communication, poly(aspartic acid) (pAsp), its fluorescently labeled conjugate with lucifer yellow ethylenediamine (pAsp–LY), and 2,2-dimethyl-1,3-propanediamine-derived cationic polyaspartamide (pDmpa) were synthesized by liquid-phase method. Octamer of L-aspartic acid (L-Asp8) was also obtained by solid-phase synthesis. pDmpa interacted with both pAsp and model plasmid DNA to form compact nanosized interpolymer complexes, which showed different colloidal properties. Effective cellular uptake of pAsp–LY by NIH 3T3 fibroblasts was detected by confocal microscopy. Pre-complexation of pAsp–LY with pDmpa did not noticeably increase intracellular accumulation of the conjugate. Both pAsp and L-Asp8 were found to increase viability of murine 3T3 cells and human skin fibroblasts at μg/mL concentrations according to the MTT assay (24 h). This effect was observed along with moderate prooxidant activity of the peptides in the cells according to the DCFDA fluorescence assay. The results suggest that aspartic acid-based peptides per se are capable of penetrating mammalian cells and affecting their metabolic activity. The peptides can be complexed with their cationic derivative, i.e., pDmpa polyaspartamide, to develop nanosized formulations of pAsp and its conjugates

Inhibition of nonspecific polymerase activity using Poly(Aspartic) acid as a model anionic polyelectrolyte

DNA polymerases with strand-displacement activity allow to amplify nucleic acids under isothermal conditions but often lead to undesirable by-products. Here, we report the increase of specificity of isothermal amplification in the presence of poly (aspartic) acids (pAsp). We hypothesized that side reactions occur due to the binding of the phosphate backbone of synthesized DNA strands with surface amino groups of the polymerase, and weakly acidic polyelectrolytes could shield polymerase molecules from DNA and thereby inhibit nonspecific amplification. Suppression of nonspecific polymerase activity by pAsp was studied on multimerization as a model side reaction. It was found that a low concentration of pAsp (0.01%) provides successful amplification of specific DNA targets. The inhibitory effect of pAsp is due to its polymeric structure since aspartic acid did affect neither specific nor nonspecific amplification. Strongly acidic polyelectrolyte heparin does not possess the same selectivity since it suppresses any DNA synthesis. The applicability of pAsp to prevent nonspecific reactions and reliable detection of the specific target has been demonstrated on the genetic material of SARS-CoV-2 coronavirus using Loop-mediated isothermal amplification

Fast dissolving nanofibrous matrices prepared by electrospinning of polyaspartamides

© 2020 Elsevier Ltd Owing to their versatile chemistry, polyaspartamides have recently attracted increased interest in various biomedical uses such as drug delivery systems and scaffolding materials. Solubility of these polymers in organic solvents and in water at certain values of pH can be fine-tuned by their chemical composition, which was exploited here to fabricate fast-dissolving electrospun matrices in ethanol with no additives. Various side groups were tested to control the solubility of the polymers as well as the morphology and moisture uptake of the matrices produced. Tertiary amine groups were immobilized to ensure high solubility around neutral pH, while modification with alkyl side groups limited moisture uptake. Finally, 3-(diethylamino)propyl and n-butyl side groups were used in equal amounts. The effect of viscosity, surface tension and specific conductivity of polymer solutions on the fiber morphology was determined in order to optimize the conditions for preparing fibers with a narrow size distribution and large specific surface area. The polymer withstood the electrospinning process without any chemical degradation, as determined by IR, and was thermally stable. Furthermore, the matrices exhibited a glass transition temperature above room temperature. The complete release of vitamin B12 was observed within one minute due to the fast dissolution of the matrices in simulated salivary fluid at pH = 6.8, supporting the potential of the developed materials in oral drug delivery

Dithiophosphate-induced redox conversions of reduced and oxidized glutathione

Phosphorus species are potent modulators of physicochemical and bioactive properties of peptide compounds. O,O-diorganyl dithiophoshoric acids (DTP) form bioactive salts with nitrogen-containing biomolecules; however, their potential as a peptide modifier is poorly known. We synthesized amphiphilic ammonium salts of O,O-dimenthyl DTP with glutathione, a vital tripeptide with antioxidant, protective and regulatory functions. DTP moiety imparted radical scavenging activity to oxidized glutathione (GSSG), modulated the activity of reduced glutathione (GSH) and profoundly improved adsorption and electrooxidation of both glutathione salts on graphene oxide modified electrode. According to NMR spectroscopy and GC–MS, the dithiophosphates persisted against immediate dissociation in an aqueous solution accompanied by hydrolysis of DTP moiety into phosphoric acid, menthol and hydrogen sulfide as well as in situ thiol-disulfide conversions in peptide moieties due to the oxidation of GSH and reduction of GSSG. The thiol content available in dissolved GSH dithiophosphate was more stable during air oxidation compared with free GSH. GSH and the dithiophosphates, unlike DTP, caused a thiol-dependent reduction of MTS tetrazolium salt. The results for the first time suggest O,O-dimenthyl DTP as a redox modifier for glutathione, which releases hydrogen sulfide and induces biorelevant redox conversions of thiol/disulfide groups

Polymer–colloid complexes based on cationic imidazolium amphiphile, polyacrylic acid and dna decamer

The solution behavior and physicochemical characteristics of polymer–colloid complexes based on cationic imidazolium amphiphile with a dodecyl tail (IA-12) and polyacrylic acid (PAA) or DNA decamer (oligonucleotide) were evaluated using tensiometry, conductometry, dynamic and electrophoretic light scattering and fluorescent spectroscopy and microscopy. It has been established that PAA addition to the surfactant system resulted in a ca. 200-fold decrease in the aggregation threshold of IA-12, with the hydrodynamic diameter of complexes ranging within 100–150 nm. Electrostatic forces are assumed to be the main driving force in the formation of IA-12/PAA complexes. Factors influencing the efficacy of the complexation of IA-12 with oligonucleotide were determined. The nonconventional mode of binding with the involvement of hydrophobic interactions and the intercalation mechanism is probably responsible for the IA-12/oligonucleotide complexation, and a minor contribution of electrostatic forces occurred. The latter was supported by zeta potential measurements and the gel electrophoresis technique, which demonstrated the low degree of charge neutralization of the complexes. Importantly, cellular uptake of the IA-12/oligonucleotide complex was confirmed by fluorescence microscopy and flow cytometry data on the example of M-HeLa cells. While single IA-12 samples exhibit roughly similar cytotoxicity, IA-12–oligonucleotide complexes show a selective effect toward M-HeLa cells (IC50 1.1 µM) compared to Chang liver cells (IC50 23.1 µM)