Browse
Submit Data
Databases
API
Help

Dataset Information

10 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Facile Method of Protein PEGylation by a Mono-Ion Complex.

ABSTRACT: Diethylaminoethyl end-modified poly(ethylene glycol) (DEAE-PEG) has been synthesized for the noncovalent PEGylation of proteins. The resulting DEAE-PEG and catalase formed an ion complex, that is, a protein mono-ion complex (MIC). The formation of the protein MIC was confirmed by native poly(acrylamide) gel electrophoresis and gel-filtration chromatography. The resulting catalase MIC preserved the catalase activity, confirmed by monitoring the O2 concentration with a Clark-type oxygen electrode, in spite of MIC formation. The catalase activity of the protein MIC was protected in the presence of a protease, trypsin, or 10% fetal bovine serum. Furthermore, less change in the circular dichroism measurements of the catalase MIC was observed as compared to those of a catalase-PEG conjugate (covalent PEGylation), suggesting less influence of the protein conformation. Consequently, the formation of the MIC is considered to be a facile method of protein PEGylation.

SUBMITTER: Asayama S

PROVIDER: S-EPMC6641148 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Facile Method of Protein PEGylation by a Mono-Ion Complex.

Asayama Shoichiro S Nagashima Kana K Kawakami Hiroyoshi H

ACS omega 20170531 5

Diethylaminoethyl end-modified poly(ethylene glycol) (DEAE-PEG) has been synthesized for the noncovalent PEGylation of proteins. The resulting DEAE-PEG and catalase formed an ion complex, that is, a protein mono-ion complex (MIC). The formation of the protein MIC was confirmed by native poly(acrylamide) gel electrophoresis and gel-filtration chromatography. The resulting catalase MIC preserved the catalase activity, confirmed by monitoring the O<sub>2</sub> concentration with a Clark-type oxygen ...[more]

PMID: 31457587

Similar Datasets

Plasmid DNA Mono-Ion Complex for in Vivo Sustainable Gene Expression.

Project description:To cleave biocompatible poly(ethylene glycol) (PEG) from the mono-ion complex (MIC) for sustainable cellular uptake in vivo, ω-amide-pentylimidazolium end-modified PEG with an ester bond, that is, APe-Im-E-PEG, has been synthesized. The hydrolysis of the resulting APe-Im-E-PEG proceeded during the incubation for 2 weeks under physiological conditions, which was confirmed by gel filtration chromatography. APe-Im-E-PEG formed the MIC with plasmid DNA (pDNA), assessed by agarose gel retardation assay. Furthermore, dynamic light scattering measurement and transmission electron microscopy observations have estimated that the particle size of the resulting MIC was approximately 30 nm, with a rather flexible structure. The APe-Im-E-PEG/pDNA MIC incubated for 2 weeks exhibited hemolytic activity at endosomal pH, presumably because the pH-sensitive carboxyl groups revealed after the hydrolysis of an ester bond of APe-Im-E-PEG. The APe-Im-E-PEG/pDNA MIC enhanced the gene expression 2 weeks after transfection in vivo by intramuscular administration in mice. Consequently, in vivo sustainable gene expression has been achieved by the molecular design of APe-Im-E-PEG for cellular uptake and endosomal escape proceeded by temporal hydrolysis of the ester bond.

| S-EPMC6681980 | biostudies-literature

Mono-PEGylation of a Thermostable Arginine-Depleting Enzyme for the Treatment of Lung Cancer.

Project description:L-arginine (L-Arg) depletion induced by randomly PEGylated arginine deiminase (ADI-PEG20) can treat arginosuccinate synthase (ASS)-negative cancers, and ADI-PEG20 is undergoing phase III clinical trials. Unfortunately, ASS-positive cancers are resistant to ADI-PEG20. Moreover, the yield of ADI production is low because of the formation of inclusion bodies. Here, we report a thermostable arginine-depleting enzyme, Bacillus caldovelox arginase mutant (BCA-M: Ser161->Cys161). An abundant amount of BCA-M was easily obtained via high cell-density fermentation and heat treatment purification. Subsequently, we prepared BCA-M-PEG20, by conjugating a single 20 kDa PEG monomer onto the Cys161 residue via thio-chemistry. Unlike ADI-PEG20, BCA-M-PEG20 significantly inhibited ASS-positive lung cancer cell growth. Pharmacodynamic studies showed that a single intraperitoneal injection (i.p). administration of 250 U/mouse of BCA-M-PEG20 induced low L-Arg level over 168 h. The mono-PEGylation of BCA-M prolonged its elimination half-life from 6.4 to 91.4 h (a 14-fold increase). In an A549 lung cancer xenograft model, a weekly administration of 250 U/mouse of BCA-M-PEG20 suppressed tumor growth significantly. We also observed that BCA-M-PEG20 did not cause any significant safety issue in mouse models. Overall, BCA-M-PEG20 showed excellent results in drug production, potency, and stability. Thereby, it has great potential to become a promising candidate for lung cancer therapy.

| S-EPMC7353006 | biostudies-literature

BuTT-Seq: a new method for facile profiling of transcription

Project description:A wide range of sequencing methods have been developed to assess nascent RNA transcription and resolve the single-nucleotide position of RNA polymerase genome-wide. These techniques are often burdened with high input material requirements and lengthy protocols. We leveraged the template-switching properties of thermostable group II intron reverse transcriptase (TGIRT) and developed BuTT-Seq (BUlk analysis of nascent Transcript Termini sequencing), which can produce libraries from purified nascent RNA in 6 hours and from as few as 10,000 cells – an improvement of at least 10-fold over existing techniques. BuTT-Seq shows that inhibition of the superelongation complex (SEC) causes promoter-proximal pausing to move upstream in a fashion correlated with subnucleosomal fragments. To address transcriptional regulation in a tissue, BuTT-Seq was used to measure the circadian regulation of transcription from fly heads. All the results indicate that BuTT-Seq is a simple and powerful technique to analyze transcription at a high level of resolution.

2023-03-30 | GSE228595 | GEO

A novel method for PEGylation of chitosan nanoparticles through photopolymerization.

Project description:An ultrafast and convenient method for PEGylation of chitosan nanoparticles has been established through a photopolymerization reaction between the acrylate groups of PEG and methacrylated-chitosan nanoparticles. The nanoparticle characteristics under physiological pH conditions were optimized through altered PEG chain length, concentration and duration of UV exposure. The method developed here has potential for clinical translation of chitosan nanoparticles. It also allows for the scalable and fast synthesis of nanoparticles with colloidal stability.

| S-EPMC9063996 | biostudies-literature

Mono-PEGylation of Alpha-MMC and MAP30 from Momordica charantia L.: Production, Identification and Anti-Tumor Activity.

Project description:PEGylation is a well-established and effective strategy to decrease immunogenicity, which can increase the stability and in vivo half-life time. However, the generation of multi-site modified products is inevitable due to the lysine chemistry, which will bring difficulties in subsequent research, such as purification and quantification. Site-specific modification by mPEG-succinimidyl carbonate (mPEG-SC) is a widely used method for N-terminal conjugation. In this study, we used it for site-directed modification on two ribosome-inactivating proteins (RIPs), alpha-momorcharin (?-MMC) and momordica anti-HIV protein (MAP30), from Momordica charantia L. According to the optimization of previous modification conditions, we compared Macro-Cap SP with SP-Sepharose FF chromatography for separating the final mPEGylated RIPs. Two kinds of methods both can obtain homogenous mPEGylated RIPs which were identified by sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE), isoelectric focusing electrophoresis (IEF), and matrix-assisted laser desorption ionization-time of flight/time of flight (MALDI-TOF/TOF) analysis. We also used iodine staining method to detect the amount of unmodified PEG. Furthermore, the inhibition activity of both mPEGylated and non-PEGylated RIPs against human lung adenocarcinoma epithelial A549 cells was detected. All of the results suggested that the mPEGylated ?-MMC/MAP30 might be potentially developed as new anti-tumor drugs.

| S-EPMC6274223 | biostudies-literature

Mix to validate: a facile, reversible PEGylation for fast screening of potential therapeutic proteins in vivo.

Project description: Not available

| S-EPMC3902477 | biostudies-literature

A facile synthesis of mono dispersed spherical silver doped bioactive glass nanoparticle.

Project description:Bioactive glasses have attracted enormous attention in the field of biomaterials for dental and medical applications. Incorporation of antibacterial ions within BGs has been proved to be a promising approach to fortify their bactericidal character. In this study, homogenous BGs containing silver (Ag) ions were synthesized by sol-gel method. Subsequently, the presence of the embedded ions were characterized by X-ray fluorescence (XRF) elemental analysis and energy dispersive X-ray (EDX) spectroscopy. Moreover, released ions were measured in simulated body fluid (SBF) and their antibacterial effectiveness was further verified using minimum bactericidal concentration (MBC) and minimum inhibitory concentration (MIC) tests. A crystalline hydroxyapatite layer was formed on the Ag-BG surfaces at day 5 approved by X-ray diffraction indicating the preserved bioactivity. The resultant uniform, mono-dispersed and dense nanoparticles show 19 great potential for a range of orthopedic and dental applications.

| S-EPMC7952368 | biostudies-literature

PEGylation to Improve Protein Stability During Melt Processing.

Project description:Biopharmaceuticals are some of the most effective drugs on the market, however, delivery remains a challenge. Melt processing is a viable protein encapsulation method because it is solvent free, is high throughput, and yields very high encapsulation efficiencies. Problematically, proteins can lose activity during melt processing due to high heat and shear forces. Covalent attachment of poly(ethylene glycol), or PEGylation, has been widely used to increase thermal stability and prevent aggregation in solution. This study explored the effect of PEGylation on protein stability during melt processing using lysozyme and PLGA. The results indicate that PEGylation increases the retained activity of lysozyme, increases dispersion in the melt, and reduces the biphasic release profile in melt processed systems.

| S-EPMC4615555 | biostudies-literature

Preferential Interactions and the Effect of Protein PEGylation.

Project description:BackgroundPEGylation is a strategy used by the pharmaceutical industry to prolong systemic circulation of protein drugs, whereas formulation excipients are used for stabilization of proteins during storage. Here we investigate the role of PEGylation in protein stabilization by formulation excipients that preferentially interact with the protein.Methodology/principal findingsThe model protein hen egg white lysozyme was doubly PEGylated on two lysines with 5 kDa linear PEGs (mPEG-succinimidyl valerate, MW 5000) and studied in the absence and presence of preferentially excluded sucrose and preferentially bound guanine hydrochloride. Structural characterization by far- and near-UV circular dichroism spectroscopy was supplemented by investigation of protein thermal stability with the use of differential scanning calorimetry, far and near-UV circular dichroism and fluorescence spectroscopy. It was found that PEGylated lysozyme was stabilized by the preferentially excluded excipient and destabilized by the preferentially bound excipient in a similar manner as lysozyme. However, compared to lysozyme in all cases the melting transition was lower by up to a few degrees and the calorimetric melting enthalpy was decreased to half the value for PEGylated lysozyme. The ratio between calorimetric and van't Hoff enthalpy suggests that our PEGylated lysozyme is a dimer.Conclusion/significanceThe PEGylated model protein displayed similar stability responses to the addition of preferentially active excipients. This suggests that formulation principles using preferentially interacting excipients are similar for PEGylated and non-PEGylated proteins.

| S-EPMC4521882 | biostudies-literature

Facile Method for Determining Lanthipeptide Stereochemistry.

Project description:Lanthipeptides make up a large group of natural products that belong to the ribosomally synthesized and post-translationally modified peptides (RiPPs). Lanthipeptides contain lanthionine and methyllanthionine bis-amino acids that have varying stereochemistry. The stereochemistry of new lanthipeptides is often not determined because current methods require equipment that is not standard in most laboratories. In this study, we developed a facile, efficient, and user-friendly method for detecting lanthipeptide stereochemistry, utilizing advanced Marfey's analysis with detection by liquid chromatography coupled with mass spectrometry (LC-MS). Under optimized conditions, 0.05 mg of peptide is sufficient to characterize the stereochemistry of five (methyl)lanthionines of different stereochemistry using a simple liquid chromatography setup, which is a much lower detection limit than current methods. In addition, we describe methods to readily access standards of the three different methyllanthionine stereoisomers and two different lanthionine stereoisomers that have been reported in known lanthipeptides. The developed workflow uses a commonly used nonchiral column system and offers a scalable platform to assist antimicrobial discovery. We illustrate its utility with an example of a lanthipeptide discovered by genome mining.

| S-EPMC10831782 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data