Poly Amino Acid Research Articles

Electrochemical investigation of a graphite-polyurethane composite electrode modified with electrodeposited gold nanoparticles in the voltammetric determination of tryptophan

Gold nanoparticles (AuNP) were electrodeposited onto the surface of a graphite-polyurethane composite electrode (EGPU), followed by electrochemical treatment in NaOH medium using cyclic voltammetry. After characterization and optimization of the experimental conditions, the resulting device (EGPU-tAuNP) was used for voltammetric determination of tryptophan (TRP). Analyses using scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDX) revealed a homogeneous covering of the electrode surface, with no alteration of the morphology due to the electrochemical treatment in NaOH. Electrochemical Impedance Spectroscopy (EIS) measurements showed that modification with AuNP and electrochemical treatment in basic media reduces the electron-transfer resistance and improves the electron-transfer rate. Chronocoulometric data and cyclic voltammograms obtained in the presence of 100.0μmolL−1 TRP revealed an increase in the peak current and a displacement of the peak potential to more negative values after the pretreatment in basic medium. A mechanism involving the Au0/Au3+ reaction on the electrode surface and TRP oxidation was proposed. Under optimized conditions, using differential pulse voltammetry (DPV), the EGPU-tAuNP presented a detection limit of 5.3×10−8molL−1 for the analysis of TRP. The device was used for the determination of TRP in synthetic urine and commercial poly-amino acids supplement. Tests of interference were carried out with biologically relevant compounds. The standard additions procedure could be used to overcome interferences in the analysis of synthetic urine containing creatinine.

Folding of poly-amino acids and intrinsically disordered proteins in overcrowded milieu induced by pH change

pH-induced structural changes of the synthetic homopolypeptides poly-E, poly-K, poly-R, and intrinsically disordered proteins (IDPs) prothymosin α (ProTα) and linker histone H1, in concentrated PEG solutions simulating macromolecular crowding conditions within the membrane-less organelles, were characterized. The conformational transitions of the studied poly-amino acids in the concentrated PEG solutions depend on the polymerization degree of these homopolypeptides, the size of their side chains, the charge distribution of the side chains, and the crowding agent concentration. The results obtained for poly-amino acids are valid for IDPs having a significant total charge. The overcrowded conditions promote a significant increase in the cooperativity of the pH-induced coil–α-helix transition of ProTα and provoke histone H1 aggregation. The most favorable conditions for the pH-induced structural transitions in concentrated PEG solutions are realized when the charged residues are grouped in blocks, and when the distance between the end of the side group carrying charge and the backbone is small. Therefore, the block-wise distribution of charged residues within the IDPs not only plays an important role in the liquid–liquid phase transitions, but may also define the expressivity of structural transitions of these proteins in the overcrowded conditions of the membrane-less organelles.

22 Vitrification of bovine embryo using antifreeze polyamino acid

Carboxylated poly-l-lysine (CPLL) is an ampholytic polymer compound and a polyamino acid with a known functional resemblance to antifreeze proteins. We previously reported that CPLL is an effective cryoprotectant for bovine cells, sperm, and slow-frozen embryos. In this study, we investigated CPLL as a cryoprotectant for vitrified bovine embryos. We developed bovine embryos in vitro and vitrified them at the blastocyst stage. Embryos were equilibrated (3min) and vitrified (1min). Vitrified embryos were cryopreserved in LN (Cryotop® device; Kitazato Corp., Tokyo, Japan) for at least 1 week, thawed with a 0.3M sucrose warming solution, and then cultured in a basal medium (Gibco® medium 199, Grand Island, NY, USA; supplemented with 100µM 2-mercaptoethanol, 10% fetal bovine serum, and antibiotics) at 38.5°C in a humidified atmosphere (5% CO2, 5% O2, 90% N2). We evaluated the embryos morphologically for survival and hatched rate at 0, 24, 48, and 72h post-thawing. In control, the equilibration solution (ES) consisted of 7.5% (vol/vol) dimethyl sulfoxide (DMSO) and 7.5% (vol/vol) ethylene glycol, and the vitrification solution (VS) consisted of 16.5% (vol/vol) DMSO and 16.5% (vol/vol) ethylene glycol and 0.5M sucrose. In this study, CPLL was added to ES and VS at various concentrations instead of DMSO. The CPLL was added at 16.5, 11.0, 5.5, and 2.2% (wt/vol) to VS; respectively, these solutions were named P16.5, P11.0, P5.5, and P2.2. The ES was used 45% CPLL of VS each. Embryos underwent the above procedure concurrently, with testing replicated at least 3 times. We evaluated 88, 34, 38, 44, and 28 embryos with each solution (control, P16.5, P11.0, P5.5, and P2.2, respectively). Results were analysed statistically with a chi-square test and residual analysis, regarding P<0.05 as significant. Survival rates were significantly greater in P11.0 at 24h post-thawing (55.7% v. 89.5%; P<0.05) and in P11.0 and P5.5 at 48h post-thawing (47.7% v. 78.9% and 47.7% v. 79.5%, respectively; P<0.05) relative to controls but showed no significant differences at 0h post-thawing. Hatched rates were significantly greater in P11.0 and P5.5 through 72h post-thawing relative to controls (44.7% v. 22.7% and 52.3% v. 22.7%, respectively; P<0.05). The CPLL improved post-thawing embryo survival and hatched rates when applied during vitrification, thus demonstrating cryoprotective effectiveness. We conclude that CPLL acts as a low-toxicity cryoprotectant for vitrified bovine embryos, and our results are consistent with previous reports of protective CPLL effects for cells and cell membranes.

SPIONs embedded in polyamino acid nanogels to synergistically treat tumor microenvironment and breast cancer cells

The extremely complex tumor microenvironment (TME) in humans is the major responsible for the therapeutic failure in cancer nanomedicine. A new concept of disease-driven nanomedicine, henceforth named “Theranomics”, which attempts to target cancer cells and TME on the whole, represents an attractive alternative. Herein, a nanomedicine able to co-deliver doxorubicin and a tumor suppressive proteolytic protein such as collagenase-2 was developed. We successfully obtained superparamagnetic nanogels (SPIONs/Doco@Col) via the intermolecular azide-alkyne Huisgen cycloaddition. We demonstrated that a local ECM degradation and remodeling in solid tumors by means of collagenase-2 could enhance tumor penetration of nanomedicines and the in situ sustained release of the drug payload throughout 3-D tumor spheroids up to the core (parenchyma), thus enabling a synergistic and efficient anticancer effect toward highly invasive breast tumors. We illustrate that SPIONs/Doxo@Col is also capable of reducing the invasivity of cancer cells.

Synthesis of multivalent polymer-aptamer conjugates with enhanced inhibitory potency.

PurposeWe are interested in designing a modular strategy for creating potent multivalent ligands, which frequently can be used as effective inhibitors of undesired biomolecular interactions. For example, such inhibitors might prevent the self-assembly of bacterial toxins or the attachment of a virus to its host cell receptors.MethodsWe used a biocompatible polyamino acid polymer as a scaffold for grafting multiple copies of an oligonucleotide aptamer (OA). Specifically, the carboxylates on the side chains of polyglutamic acid (PGA) were modified with a thiol-reactive linker, N-aminoethyl maleimide (AEM), and thiol-functionalized OAs were attached to the maleimide moieties. The resulting conjugates were tested for their ability to compete with and inhibit the binding of unconjugated monovalent OAs to the target cell receptor.ResultsMultivalent PGA–OA conjugates with low, medium, and high valency were successfully prepared. The varying valency and successful purification to remove unconjugated OAs were confirmed by polyacrylamide gel electrophoresis. The resulting purified conjugates inhibited the binding of unconjugated monovalent OAs, and the measured half maximal inhibitory concentration (IC50) values corresponded to a 38–88-fold enhancement of potency on a per-aptamer basis, relative to OA alone.ConclusionMultivalent conjugation of OA ligands has potential as a generally useful way to improve the potency of the interaction between the ligand and its target receptor. We have demonstrated this principle with a known OA as a proof of concept as well a synthetic strategy that can be used to synthesize multivalent conjugates of other OAs.

Measurements of Atmospheric Proteinaceous Aerosol in the Arctic Using a Selective UHPLC/ESI-MS/MS Strategy

In this article, an analytical methodology to investigate the proteinaceous content in atmospheric size-resolved aerosols collected at the Zeppelin observatory (79 °N, 12 °E) at Ny Ålesund, Svalbard, from September to December 2015, is proposed. Quantitative determination was performed after acidic hydrolysis using ultrahigh-performance liquid chromatography in reversed-phase mode coupled to electrospray ionization tandem mass spectrometry. Chromatographic separation, as well as specificity in the identification, was achieved by derivatization of the amino acids with N-butyl nicotinic acid N-hydroxysuccinimide ester prior to the analysis. The chromatographic run was performed within 11 min and instrumental levels of detection (LODs) were between 0.2 and 8.1 pg injected on the column, except for arginine which exhibited an LOD of 37 pg. Corresponding method LODs were between 0.01 and 1.9 fmol/m3, based on the average air sampling volume of 57 m3. The sum of free amino acids and hydrolyzed polyamino acids was shown to vary within 6–2914 and 0.02–1417 pmol/m3 for particles in sizes < 2 and 2–10 μm in equivalent aerodynamic diameter, respectively. Leucine, alanine, and valine were the most abundant among the amino acids in both aerosol size fractions. In an attempt to elucidate source areas of the collected aerosols, 5- to 10-day 3D backward trajectories reaching the sampling station were calculated. Overall, the method described here provides a first time estimate of the proteinaceous content, that is, the sum of free and polyamino acids, in size-resolved aerosols collected in the Arctic.Graphical ᅟ

Polyamino Acid Layer-by-Layer (LbL) Constructed Silica-Supported Mesoporous Titania Nanocarriers for Stimuli-Responsive Delivery of microRNA 708 and Paclitaxel for Combined Chemotherapy.

Cellular Fas-associated protein with death domain-like interleukin-1β-converting enzyme-inhibitory protein (c-FLIP), often strongly expressed in numerous cancers, plays a pivotal role in thwarting apoptosis and inducing chemotherapy resistance in cancer. An integrated approach combining chemotherapy with suppression of c-FLIP levels could prove paramount in the treatment of cancers with c-FLIP overexpression. In this study, we utilized a polymeric layer-by-layer (LbL) assembly of silica-supported mesoporous titania nanoparticles (MTNst) to co-deliver paclitaxel (PTX) and microRNA 708 (miR708) for simultaneous chemotherapy and c-FLIP suppression in colorectal carcinoma. The resulting LbL miR708/PTX-MTNst showed dose-dependent cytotoxicity in HCT-116 and DLD-1 colorectal carcinoma cell lines, which was remarkably superior to that of free PTX or LbL PTX-MTNst. LbL miR708/PTX-MTNst strongly inhibited c-FLIP expression and resulted in increased expression of proapoptotic proteins. In DLD-1 xenograft tumor-bearing mice, the nanoparticles accumulated in the tumor, resulting in remarkable tumor regression, with the PTX and miR708-loaded nanoparticles showing significantly greater inhibitory effects than the free PTX or PTX-loaded nanoparticles. Immunohistochemical analyses of the tumors further confirmed the remarkable apoptotic and antiproliferative effects of the nanoparticles, whereas organ histology reinforced the biocompatibility of the system. Therefore, the LbL miR708/PTX-MTNst system, owing to its ability to deliver both chemotherapeutic drug and inhibitory miRNA to the tumor site, shows great potential to treat colorectal carcinoma in clinical settings.

Polyamino acid interlayer facilitates electron extraction in narrow band gap fullerene-free organic solar cells with an outstanding short-circuit current

In this communication, we present a study to investigate interfacial characteristics modulated by electron extraction layers (EEL) comprising of a polyamino acid α-poly-L-lysine (α-PLL) in fullerene-free organic solar cells (OSC) based on narrow-band-gap PTB7-Th:IEICO-4 heterojunctions. Benefitting from effective interfacial modifications with the α-PLL EEL, PTB7-Th:IEICO-4F solar cells receive a considerable boost in the short-circuit current (26.6 mA/cm2) and ultimate power conversion efficiencies exceeding 12%, which is a result of promoted charge extraction. Aided by combinatorial analyses using electron spin resonance spectroscopy, angle-resolved X-ray photoelectron spectroscopy together with charge transport measurements, we found that the interfacial doping occurs at the α-PLL/IEICO-4F interface, possibly due to the low steric hindrance of the bared primary amines and non-conjugated backbones in α-PLL. The interface strategy with α-PLL displays a universal applicability, enabling to produce a series of high efficiency PCBM or non-fullerene OSCs with different device architectures. This work enriches fundamental insights into key interfacial processes and provides a promising pathway utilizing wide-accessible biomaterials to attain high performance OSCs with emerging fullerene-free photovoltaic materials.

One-step synthesis of europium complexes containing polyamino acids through ring-opening polymerization and their potential for biological imaging applications

Lanthanide-doped nanoprobes have received significant attentions for utilization in biological sensing and imaging due to their unique optical properties. However, only few works were focused on fabrication of europium complexes based fluorescence polymeric nanoparticles (FPNs) with excellent biocompatibility and biodegradability. In this study, we fabricated the FPNs (named as Eu(TTA)3Phen-GluEG FPNs) with encapsulated europium complexes which show low cytotoxicity, high sensitivity and deep penetration. Free amine group present on europium complexes initiated the ring-opening polymerization (ROP) of side-chain modified glutamic acid NCAs, offering a simple and effective method to prepare europium core FPNs with a uniform size distribution. Europium (III) chelates were furnished with a functional polyamino acid shell to fabricate biocompatible and biodegradable FPNs. Biological evaluation results demonstrate that such fabricated FPNs process excellent biocompatibility and dyeing performance. Therefore, we can expect that the fabrication approach will attract much research interest and effort on the preparation of biodegradable probes for various biological applications.

Electrochemical monitoring of malondialdehyde biomarker in biological samples via electropolymerized amino acid/chitosan nanocomposite.

This study reports on the electropolymerization of a low toxic and biocompatible nanopolymer with entitle poly arginine-graphene quantum dots-chitosan (PARG-GQDs-CS) as a novel strategy for surface modification of glassy carbon surface and preparation of a new interface for measurement of malondialdehyde (MDA) in exhaled breath condensate. Electrochemical deposition, as a well-controlled synthesis procedure, has been used for subsequently layer-by-layer preparation of GQDs-CS nanostructures on a PARG prepolymerized on the surface of glassy carbon electrode using cyclic voltammetry techniques in the regime of -1.5 to 2V. The modified electrode appeared as an effective electroactivity for detection of MDA by using cyclic voltammetry, linear sweep voltammetry, and differential pulse voltammetry. The prepared modified electrode demonstrated a noticeably good activity for electrooxidation of MDA than PARG. Enhancement of peak currents is ascribed to the fast heterogeneous electron transfer kinetics that arise from the synergistic coupling between the excellent properties of PARG and semiconducting polymer, GQDs as high density of edge plane sites and subtle electronic characteristics and unique properties of CS such as excellent film-forming ability, high permeability, good adhesion, nontoxicity, cheapness, and a susceptibility to chemical modification. The prepared sensor showed 1 oxidation processes for MDA at potentials about 1V with a low limit of quantification 5.94nM. Finally, application of new sensor for determination of MDA in exhaled breath condensate was suited. In general, the simultaneous attachment of GQDs and CS to structure of poly amino acids provides new opportunities within the personal healthcare.

Incorporation of gelatin microparticles on the formation of adipose-derived stem cell spheroids

The three-dimensional (3D) cell culture method provides a physiological environment that is similar to the in vivo conditions. The spheroid culture method is a 3D cell-culture approach that facilitates aggregation and interaction of cells with other adjacent cells to establish a microenvironment in vitro. In this study, gelatin microparticles (GMPs) were introduced into spheroids to enhance stem cell proliferation and viability. GMPs with different sizes and numbers were prepared to investigate their corresponding effects on cell proliferation. To investigate the effect of surface charge on the properties of spheroids, positively and negatively charged GMPs were also prepared by conjugating poly-amino acids on the surfaces of the GMPs. GMPs were successfully incorporated into adipose-derived stem cells (ASCs) to form spheroids. The results revealed no cytotoxicity in all treatment groups. Spheroids combined with GMPs showed higher proliferation potential compared to cell spheroids alone, suggesting that GMP/cell spheroids can be used for efficient delivery of stem cells to defect sites without the need for surgery, thereby highlighting the potential use of GMP/cell spheroids for stem cell therapy.

Calcite Crystallization on Polarized Single Calcite Crystal Substrates in the Presence of Poly-Lysine

We demonstrated the crystallization of calcite on nonpolarized and polarized (10.4)-oriented calcite single crystal substrates in the presence of poly-lysine (PLys) as a poly amino acid. The PLys and the electric field due to polarized calcite substrates showed a significant effect on the morphologies of calcite precipitates. At a small amount of PLys, rhombohedral calcite crystals, with notches in the edges of the rhombohedra, formed through homoepitaxy and constructed aggregates with an island structure. At a large amount of PLys, the calcite precipitates had overgrown calcite crystals, which were formed by epitaxial growth on each of the rhombohedral surfaces of already formed calcite rhombohedra, and the calcite aggregates became elongated in the direction perpendicular to the substrate. As the amount of PLys increased, the overgrown calcite crystals tapered toward their ends, and some of the overgrown calcite crystals exhibited a cone-shaped appearance. These cone-shaped crystals consisted of aggrega...

RNA Related Pathology in Huntington's Disease.

This chapter summarises research investigating the expression of huntingtin sense and anti-sense transcripts, the effect of the mutation on huntingtin processing as well as the more global effect of the mutation on the coding and non-coding transcriptomes. The huntingtin gene is ubiquitously expressed, although expression levels vary between tissues and cell types. A SNP that affects NF-ĸB binding in the huntingtin promoter modulates the expression level of huntingtin transcripts and is associated with the age of disease onset. Incomplete splicing between exon 1 and exon 2 has been shown to result in the expression of a small polyadenylated mRNA that encodes the highly pathogenic exon 1 huntingtin protein. This occurs in a CAG-repeat length dependent manner in all full-length mouse models of HD as well as HD patient post-mortem brains and fibroblasts. An antisense transcript to huntingtin is generated that contains a CUG repeat that is expanded in HD patients. In myotonic dystrophy, expanded CUG repeats form RNA foci in cell nuclei that bind specific proteins (e.g. MBL1). Short, pure CAG RNAs of approximately 21 nucleotides that have been processed by DICER can inhibit the translation of other CAG repeat containing mRNAs. The HD mutation affects the transcriptome at the level of mRNA expression, splicing and the expression of non-coding RNAs. Finally, expanded repetitive stretched of nucleotides can lead to RAN translation, in which the ribosome translates from the expanded repeat in all possible reading frames, producing proteins with various poly-amino acid tracts. The extent to which these events contribute to HD pathogenesis is largely unknown.

Self-enhanced PEI-Ru(II) complex with polyamino acid as booster to construct ultrasensitive electrochemiluminescence immunosensor for carcinoembryonic antigen detection

In this work, the luminophor tris (4,4′-dicarboxylicacid-2,2′-bipyridyl) ruthenium (II) dichloride (Ru(dcbpy)32+) was linked to coreactant polyethylenimine (PEI) with effectively shorted distance, forming a novel self-enhanced PEI-Ru(II) complex to fabricate ultrasensitive electrochemiluminescence immunosensor for carcinoembryonic antigen (CEA) detection by using polyamino acid l-cysteine (pL-Cys) as booster for further signal amplification. The light-emitting species (PEI-Ru(II)*) based on intramolecular electron transfer between PEI• and Ru(III) could efficiently amplify the ECL signal of Ru(dcbpy)32+ owing to its benefit for electron transmission. Secondly, the pL-Cys film was fabricated on the glassy carbon electrode by cyclic voltammogram. After that, the pL-Cys acted as a strongly reducing radical could react with PEI-Ru(III) to generate the excited state PEI-Ru(II)*, which further amplify the ECL signal. With the signal amplification factors, the prepared ECL immunosensor exhibited great advantages in simplification, rapidity, stability and selectivity. Furthermore, the linear range for the determination of CEA was from 0.10 pg/mL to 80 ng/mL with a correlation coefficient of 0.9942 and a detection limit of 0.045 pg/mL (S/N = 3).

Electrochemical quantification of some water soluble vitamins in commercial multi-vitamin using poly-amino acid caped by graphene quantum dots nanocomposite as dual signal amplification elements

Rapid analyses of some water soluble vitamins (Vitamin B2, B9, and C) in commercial multi vitamins could be routinely performed in analytical laboratories. This study reports on the electropolymerization of a low toxic and biocompatible polymer “poly aspartic acid-graphene quantum dots” as a novel strategy for surface modification of glassy carbon electrode and preparation a new interface for measurement of selected vitamins in commercial multi vitamins. Electrochemical deposition, as a well-controlled synthesis procedure, has been used for subsequently layer-by-layer preparation of graphene quantum dots nanostructures on a poly aspartic acid using cyclic voltammetry techniques in the regime of −1.5 to 2 V. The field emission scanning electron microscopy indicated immobilization of graphene quantum dots onto poly aspartic acid film. The modified electrode possessed as an effective electroactivity for detection of water soluble vitamins by using cyclic voltammetry, chronoamperometry and differential pulse voltammetry. Enhancement of peak currents is ascribed to the fast heterogeneous electron transfer kinetics that arise from the synergistic coupling between the excellent properties of poly aspartic acid as semiconducting polymer, graphene quantum dots as high density of edge plane sites and chemical modification. Under the optimized analysis conditions, the prepared sensor for detection of VB2, VB9, and VC showed a low limit of quantification 0.22, 0.1, 0.1 μM, respectively.

Turbidimetric studies of colloidal silica/aqueous solution system stability

The effects of solid content, temperature, supporting electrolyte (sodium chloride (NaCl)) concentration, solution pH, molecular weight of synthetic non-ionic polymer, natural polymer addition and synthetic copolymer presence on the silica suspension stability were examined. The natural proteins – human serum albumin and lysozyme (LSZ) – as well as the exopolysaccharide synthesized by soil bacterium Sinorhizobium meliloti 1021 were used. Moreover, synthetic homopolymers – non-ionic poly(vinylpyrrolidone) and copolymers of polyamino acids with poly(ethylene glycol) – were applied. The turbidimetric method was used to monitor the colloidal silica aqueous suspension stability as a function of time. LSZ and copolymers containing polylysine at the basic values of solution pH have the greatest effect on system destabilization. The negative surface charge neutralization by the positively charged groups of the polymeric chains is mainly responsible for such behavior of the systems under examination. On the other hand, the synthetic polyvinylpyrrolidone causes considerable improvement of silica suspension stability. Its adsorption on the solid particles surface causes efficient stabilization due to steric repulsion occurrence.

His6-OPH and Its Stabilized Forms Combating Quorum Sensing Molecules of Gram-Negative Bacteria in Combination with Antibiotics

Background: In most species of gram-negative pathogenic and conditionally pathogenic bacteria, quorum sensing-dependent systems have been discovered, in which various N-acyl-L-homoserine lactones function as signaling molecules inducing the development of resistance towards the effects of antibiotics. Objectives: The purpose of this study was to investigate the possibility of stabilizing lactonase activity of His6-OPH by forming enzyme polyelectrolyte non-covalent complexes (EPNCs) containing both this enzyme and polyanionic polymer. It also targeted at studying the efficacy of these EPNCs in combination with a variety of antibiotics (ampicillin, gentamicin, kanamycin, and rifampicin) against gram-negative bacteria. Cheap non-toxic biodegradable polyamino acids (poly-L-glutamic acid and poly-L-aspartic acid) were selected as polyanionic polymers to produce EPNCs. Methods: Recombinant Escherichia coli strain SG13009 [pREP4], transformed by plasmid encoding His6-OPH, was used for His6-OPH production. The His6-OPH lactonase activity was determined according to known methods, using a pH-sensitive indicator (cresol red) and a colorimetric method. The minimum inhibitory concentration (MIC) of antibiotics was determined with the suspension of bacteria cells, with antibiotic added in a concentration range from 0 to 500 mg/L at 37°C during 16 hours. Results: All the tested enzyme preparations were equally efficient in catalyzing the hydrolysis of N-acyl-homoserine lactones of various structures containing and lacking a 3-oxo group in the acyl radical. It has been established that the presence of enzyme preparations facilitates the reduction of MIC of antibiotics (ampicillin, gentamicin sulfate, kanamycin sulfate, and rifampicin) on the growth of highly concentrated (106 cells/mL) cell populations of Pseudomonas aeruginosa B-6643 and Escherichia coli B-6645. Polyelectrolyte complexes of this enzyme have an increased efficiency at a lower pH (6.5) of the medium in comparison to its highly purified form. It is possible to reduce the minimum inhibitory concentration of antibiotics by 10% to 850% in combined use of antibiotics and His6-OPH enzyme preparations. Conclusions: The discussed approaches for the use of original biological products based on His6-OPH opens up opportunities for creating new ways of effectively controlling bacterial diseases in animals.

Environment-Sensitive Turn-On Fluorescent Polyamino Acid: Fingerprinting Protein Populations with Post-Translational Modifications.

The identification of post-translational modifications (PTMs) in proteins has been of particular interest in the elucidation of human diseases and the improvement of therapeutic proteins. Herein, we report a novel strategy toward the construction of fingerprint-based PTM-sensing systems as an alternative to conventional specific recognition tools. Our strategy is based on poly-l-lysine (PLL) derivatives with two distinct properties suitable to fingerprint-based protein-sensing: (i) a turn-on fluorescent signal upon binding to proteins and (ii) condition-dependent cross-reactivity toward proteins and PTMs. One type of PLL derivative under varying solution properties (pH value and ionic strength) was sufficient to construct a sensing array that produces unique fluorescence fingerprints for structurally similar mammalian albumins with/without a wide variety of chemical modifications corresponding to PTMs. This approach was also applicable for the recognition of deviations in physicochemical properties of proteins as a result of realistic glycation and phosphorylation events. Multivariate analyses of the thus obtained fingerprints successfully identified analytes with 100% accuracy (qualitatively and quantitatively) in all cases. This study thus demonstrates for the first time a fingerprint-based sensing of proteins with/without PTMs using a single, highly accessible, and tunable synthetic polymer, and accordingly offers a powerful platform for simple high-throughput sensing of PTMs in proteins.

Removal studies of Cr2O3 colloidal particles using cationic poly(l-lysine) and its block copolymers with poly(ethylene glycol)

Cr2O3 dispersions are considered to be dangerous for aqueous ecosystems and for this reason it is necessary to develop methods allowing solid particles separation.Structure of the adsorption layer formed at the solid surface determines the colloidal suspension stability. Moreover, in the case of ionic polymers the solution pH considerably affects the chains arrangement. Hence, the influence of cationic poly(l-lysine) and its copolymers adsorption on the chromium(III) oxide (Cr2O3) particles stability was investigated. Although the analyzed substances are characterized by the similar average molecular weight, they exhibit various macromolecular structures resulting from the different PEG and polyamino acid length ratios. The sample's stability was examined by the use of the turbidimetric method enabling the Turbiscan Stability Index (TSI) parameter calculation.As one can see, solid suspensions without the tested polymers are relatively unstable. At pH 4, introduction of the cationic macromolecules causes the increase in the sample stability. This can be explained by the electrostatic repulsion between the totally ionized polymeric chains adsorbed on the adjacent Cr2O3 particles. At pHpzc, changes in the TSI values are related to the different polymer chain arrangements.

PH and redox dual-responsive multifunctional gene delivery with enhanced capability of transporting DNA into the nucleus

Stimuli-responsive gene delivery vectors based on physiologically triggered structure changing have been recently recognized as a new therapeutic agent for their excellent performance in vivo. Herein, we present an intelligent gene delivery system based on the octa-arginine peptides (R8)-conjugated polyamino acid derivatives noted as PPCRC (PVIm-(PAsp-Cystamine-R8)-Cholesteryl), which processed pH responsive, surface charge-switching, intracellular redox-responsive and enhanced nucleus import of gene together. Due to the imidazole group in the PPCRC backbone, the DNA/PPCRC polyplexes not only exhibited the enhanced buffering capacity in the endosome after endocytosis, but also displayed the reversible surface charge from negative to positive with decreasing the pH value form pH 7.4 to pH 6.5–6.8, which would promote the cell membrane binding and cellular uptake. The disulfide bond for R8 peptides conjugation in the polymer side chain could be rapidly cleaved under reductive conditions, facilitating DNA release in the cytoplasm. Subsequently, the DNA would be still associated with the R8 peptides, which would promote the intracellular nucleus import of DNA. The luciferase gene expression level of COS-7 cells transfected by DNA/PPCRC polyplexes was almost 2000 folds higher than cells transfected by DNA/PPCC polyplexes (without R8 peptides modification) in growth-arrested cell model. Nearly 10 folds enhanced gene transfection efficiency was found on human bone mesenchymal stem cells (hBMSCs) using the same strategy, which revealed that this intelligent vector can be also utilized in transfection of non-dividing cells. Intravenous injection of the DNA/PPCRC polyplexes also achieved the effective transfection in subcutaneous tumor model. Taken together, PPCRC vector has great potential for both dividing and non-dividing cells transfection and in vivo gene delivery application.