Interaction of Ionenes with Lipid Membrane: Unusual Impact of Charge Density.

Synthetic water-soluble polymers are increasingly used for gene delivery, stabilization, and delivery of proteins, and as prospective antimicrobial and antiviral agents. Therefore, study of their interaction with lipid membranes is of special importance. Herein, we studied interaction of aliphatic cationic ionenes (recently tested for gene delivery efficiency) differed in the length of spacer between charged groups (and therefore in charge density) with anionic lipid membrane.

Tightly bound polyelectrolytes enhance enzyme proteolysis and destroy amyloid aggregates.

The use of polyelectrolytes is a prospective approach to form nanocomplexes to transport different compounds including proteins. In many cases, the bound protein should be digested after delivery to the target. In the present work, we studied proteolysis of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in the complexes with polyelectrolytes.

Chaperone-like activity of synthetic polyanions can be higher than the activity of natural chaperones at elevated temperature.

Polyelectrolytes are a prospective tool for protection of proteins against aggregation. We compared synthetic polyanion, poly(styrene sulfonate), and natural chaperones of different types, namely, GroEL-like chaperonin from Pseudomonas aeruginosa phage EL and human small heat shock protein HspB5 (αB-crystallin), in their ability to prevent aggregation of client proteins. At 45 °C, all three agents efficiently suppressed thermal aggregation of phage endolysin.

Cationic nanogels as Trojan carriers for disruption of endosomes.

The comparison study of interaction of linear poly(2-dimethyl amino)ethyl methacrylate and its cationic nanogels of various cross-linking with both DNA and sodium poly(styrene sulfonate) has been performed. Although all amino groups of the nanogels proved to be susceptible for protonation, their accessibility for ion pairing with the polyanions was controlled and impaired with the cross-linking. The investigation of nanogels complexes with cells in culture that was accomplished by using of calcein pH-sensitive probe revealed a successive increase in the cytoplasmic fluorescence upon the growth in the cross-linking due to calceine leakage from acidic compartments to cytosol.

Sulfated and sulfonated polymers are able to solubilize efficiently the protein aggregates of different nature.

The search for new ways to suppress unwanted protein aggregation represents an important problem in modern biochemistry, bioengineering, and even medicine. Recently we succeeded in preventing the aggregation using synthetic polyelectrolytes. The present work describes a new approach to solubilizing pre-formed protein aggregates with sulfated or sulfonated polymers (polysulfoanions).

Water-soluble polyelectrolyte complexes of Astramol poly(propyleneimine) dendrimers with poly(methacrylate) anion.

Water-soluble complexes formed by pyrenyl-tagged poly(methacrylate) anion with cationic DAB-dendr-(NH2)x of five generations, x = 4, 8, 16, 32, and 64 were prepared and studied. The ability of the dendrimers to quench the pyrenyl fluorescence was used to monitor formation/dissociation of the complexes by fluorescence quenching technique. In salt-free solutions, dissociation of the complexes occurred in highly acidic and highly alkaline media independently on the dendrimer generation, whereas stability of the complexes against destruction by added salt (NaCl) enhanced markedly with x increase.

Interaction of Astramol poly(propyleneimine) dendrimers with DNA and poly(methacrylate) anion in water and water-salt solutions.

Interaction of poly(propyleneimine) dendrimers DAB-dendr-(NH2)x of five generations (x = 4, 8, 16, 32, and 64) with either calf thymus DNA or tagged by pyrenyl groups poly(methacrylate) anion (PMA*) as well as destruction of formed polyelectrolyte complexes by the added sodium chloride were studied by fluorescence quenching techniques. DNA-containing complexes (dendriplexes) were investigated by ethidium bromide assay, whereas formation of PMA* complexes was estimated by fluorescence of the pyrenyl groups that remained free of contact with the dendrimers-quenchers. The ion pairing with DNA phosphate groups was pH-sensitive and accompanied by inaccessibility of a part of the dendrimer amino groups even in slightly acidic media.

Effect of poly(phosphate) anions on glyceraldehyde-3-phosphate dehydrogenase structure and thermal aggregation: comparison with influence of poly(sulfoanions).

Background: It is well documented that poly(sulfate) and poly(sulfonate) anions suppress protein thermal aggregation much more efficiently than poly(carboxylic) anions, but as a rule, they denature protein molecules. In this work, a polymer of different nature, i.e.

Interaction of polyelectrolytes with proteins, 3. Influence of complexing polycations on the thermoaggregation of oligomeric enzymes.

The ability of quaternized polyamines (poly-N-alkyl-4-vinylpyridinium bromides possessing a number, m, of methylene groups in the N-alkyl substituent or a degree of alkylation, beta, and n,n-ionene bromides) to suppress the thermoaggregation of glyceraldehyde-3-phosphate dehydrogenase increased in the order m = 1 < 3 < 5, beta = 95 < 85 < 70 << 45 < 35 < 20 and n = 3 < 6 < 10, which agrees well with the increase, in the same order, in the hydrophobicity of the chains. Complexing suppressed thermoaggregation, but not thermodenaturation of the enzyme, which was even encouraged by the polycations and occurred at room temperature when the most efficient suppressor (with beta = 20) was used. The adverse effect was reduced by the addition of sodium chloride which destroyed the complex and resulted in a noticeable reactivation.

Quaternized poly(4-vinylpyridine)s as model gene delivery polycations: structure-function study by modification of side chain hydrophobicity and degree of alkylation.

To optimize polycation-based gene delivery agents, the influence of molecular characteristics of the polycations on physicochemical properties of polycation/DNA complexes and their relationships to cellular gene transfer need to be understood. With this aim, we have synthesized a series of model polymers based on quaternized poly(4-vinylpyridine)s (CnPVP-beta) with the same DP of 1600 but differing by the number n of methylene groups in N-alkyl ester substituents from 1 to 6 and/or by degree of alkylation beta from 25% to 95%. For polycations CnPVP-95, the efficiency of transfection of a plasmid vector expressing a secreted form of alkaline phosphatase started to be detectable at n = 4, noticeable at n = 5, and again undetectable at n = 6.

Interpolyelectrolyte reactions in solutions of polycarboxybetaines, 2: influence of alkyl spacer in the betaine moieties on complexing with polyanions.

Series of polycarboxybetaines (PCB-n) of pyridiniocarboxylate structure with the same degree of polymerization but differing in the number, n, of methylene groups in the alkyl spacer between charges in the betaine moieties, n = 1, 2, 3, 4, 5, and 8, were synthesized. The utility of PCB-n as positively charged components of polyelectrolyte complexes was elucidated by potentiometry, turbidimetry, and fluorescence spectroscopy. Affinity of PCB-n to the pyrenyl-tagged poly(methacrylic) acid (PMAA) or DNA was judged from the stability of the corresponding polyelectrolyte complexes in water-salt solutions at different pH values as monitored by fluorescence quenching techniques.

Direct capture of plasmid DNA from non-clarified bacterial lysate using polycation-grafted monoliths.

Monolith columns from macroporous polyacrylamide gel were grafted with polycations, poly(N,N-dimethylaminoethyl methacrylate) (polyDMAEMA), (2-(methacryloyloxy)ethyl)-trimethyl ammonium chloride (polyMETA) and partially quaternized polyDMAEMA prepared via treating polyDMAEMA-grafted columns with propylbromide. The polymer grafting degrees varied between 34 and 110%. The polycation-grafted monolithic columns are able to capture plasmid DNA directly from alkaline lysate of Escherichia coli cells.

Interaction of polyanions with basic proteins, 2(a) : influence of complexing polyanions on the thermo-aggregation of oligomeric enzymes.

The ability of synthetic polyanions to suppress thermo-aggregation of the oligomeric enzymes (glyceraldehyde-3-phosphate dehydrogenase, lactate dehydrogenase, and aspartate aminotransferase) has been established. The ability of the polyanions to reduce the thermo-aggregation increased in the order poly(methacrylic acid) < poly(acrylic acid) < sodium poly(styrene sulphonate), which agreed well with the increase, in the same order, of the charge density of the chains. The lengthening of the chains, as well as the rise in their relative content, resulted in an increase of the ability to reduce thermo-aggregation, mentioned above.

Recognition and selective binding of DNA by ionenes of different charge density.

The ability of aliphatic ionenes to recognize and bind DNA or poly(methacrylic acid) (PMA) in the equimolar mixture of these polyanions was studied by fluorescence quenching technique. Within a particular system, the selectivity of competitive interactions was shown to be determined by a component with the lowest degree of polymerization (DP). Ionene polycations with lowest DP values did not exhibit pronounced selectivity in binding DNA or PMA with higher values of DP.

Multilayers of a globular protein and a weak polyacid: role of polyacid ionization in growth and decomposition in salt solutions.

Thin films obtained from a layer-by-layer deposition of a weak polycarboxylic acid and a positively charged globular protein were studied by in situ ATR-FTIR. The system was chicken egg lysozyme (Lys), bovine pancrease ribonuclease A (RNase), or bovine gamma-globulin (IgG) self-assembled with polycarboxylic acids. When the pH value was lowered below a critical point, the growth of films and their tolerance to decomposition by added sodium chloride improved dramatically.

Polyelectrolyte complexes as a tool for purification of plasmid DNA. Background and development.

The demand for highly purified plasmids in gene therapy and plasmid-based vaccines requires large-scale production of pharmaceutical-grade plasmid. Plasmid DNA was selectively precipitated from a clarified alkaline lysate using the polycation poly(N,N'-dimethyldiallylammonium) chloride which formed insoluble polyelectrolyte complex (PEC) with the plasmid DNA. Soluble PECs of DNA with polycations have earlier been used for cell transformation, but now the focus has been on insoluble PECs.

Rapid polyelectrolyte-based immunofiltration technique for testosterone detection in serum samples.

A new immunofiltration assay for testosterone is proposed. During the first step of the assay, testosterone molecules in serum samples compete in solution with the testosterone-peroxidase conjugate for interaction with anti-testosterone antibodies pre-bound to the conjugate between staphylococcal protein A and polymethacrylate polyanion. The reaction mixture is then filtered through a membrane charged with immobilized poly(N-ethyl-4-vinylpyridinium) polycation.

Competitive reactions in solutions of poly-L-histidine, calf thymus DNA, and synthetic polyanions: determining the binding constants of polyelectrolytes.

The physicochemical characteristics of a nonviral gene delivery system will govern its functional bioactivity; however, empiricism dominates the literature in this field, and a significant deficiency of quantitative investigation and evaluation of nonviral gene delivery vehicles remains. Herein, we derive a physical model and experimental method to quantitatively determine the binding constants between a model polycationic nonviral gene delivery vehicle poly-L-histidine (PLH) and calf thymus DNA. The approach has utility to a variety of systems and is not limited to the described polymer model.

Polyhistidine-PEG:DNA nanocomposites for gene delivery.

Complexation of plasmid DNA with polycations is a popular method by which to transfer therapeutic nucleic acid sequences to cells. One caveat of the approach is that the positive zeta potential of the complexes facilitates interaction with blood constituents, leading to serum protein adsorption and complement activation. As a countermeasure, investigators have developed polycations combined with polyethylene glycol (PEG) to create complexes with reduced protein adsorption potential.

Aliphatic ionenes as gene delivery agents: elucidation of structure-function relationship through modification of charge density and polymer length.

Polycation-based gene delivery agents are generally polydisperse populations whose properties are averaged among the different molecular weight species. Therefore, to understand the physicochemical properties of polycations and their relationships to cellular gene transfer, one needs to control the molecular weight of the polymer as well as its cationic charge density. To investigate the structure-function correlation of polycations with respect to the degree of polymerization (DP) and charge density, a series of model materials based on aliphatic ionenes was synthesized and fractionated into distinct molecular weight fractions with DP range from 14 to 32.