Peptide Moiety Research Articles

Intracellular Bacterial Targeting by a Thiazolyl Benzenesulfonamide and Octaarginine Peptide Complex.

A brominated thiazolyl benzenesulfonamide (BTB) derivative is conjugated with the cell-penetrating peptide octaarginine (R8) in an effort to construct innovative antibacterial products. The noncovalent complex of BTB and R8 is characterized by Fourier transform infrared (FTIR) spectroscopy, which indicates hydrogen bonding between the two constituents. Attachment of the peptide moiety renders aqueous solubility to the hydrophobic benzenesulfonamide drug and bestows bactericidal activity. Confocal imaging in conjunction with dye probes shows successful clearance of intracellular Staphylococcus aureus bacteria by the BTB-R8 complex. Scanning electron micrographs and studies with a set of fluorescent dyes suggest active disruption of the bacterial cell membrane by the BTB-R8 complex. In contrast, the complex of BTB with octalysine (K8) fails to cause membrane damage and displays a modest antibacterial effect. A complex of BTB with the water-soluble hydrophilic polymer poly(vinylpyrrolidone) (PVP) does not display any antibacterial effect, indicating the distinctive role of the cell-penetrating peptide (CPP) R8 in the cognate complex. The leakage of the encapsulated dye from giant unilamellar vesicles upon interaction with the BTB-R8 complex further highlights the membrane activity of the complex, which cannot be accomplished by bare sulfonamide alone. This work broadens the scope of use of CPPs with respect to eliciting antibacterial activity and potentially expands the limited arsenal of membrane-targeting antibiotics.

Prediction of Intact N-Glycopeptide Retention Time Windows in Hydrophilic Interaction Liquid Chromatography.

Analysis of protein glycosylation is challenging due to micro- and macro-heterogeneity of the attached glycans. Hydrophilic interaction liquid chromatography (HILIC) is a mode of choice for separation of intact glycopeptides, which are inadequately resolved by reversed phase chromatography. In this work, we propose an easy-to-use model to predict retention time windows of glycopeptides in HILIC. We constructed this model based on the parameters derived from chromatographic separation of six differently glycosylated peptides obtained from tryptic digests of three plasma proteins: haptoglobin, hemopexin, and sex hormone-binding globulin. We calculated relative retention times of different glycoforms attached to the same peptide to the bi-antennary form and showed that the character of the peptide moiety did not significantly change the relative retention time differences between the glycoforms. To challenge the model, we assessed chromatographic behavior of fetuin glycopeptides experimentally, and their retention times all fell within the calculated retention time windows, which suggests that the retention time window prediction model in HILIC is sufficiently accurate. Relative retention time windows provide complementary information to mass spectrometric data, and we consider them useful for reliable determination of protein glycosylation in a site-specific manner.

Design and synthesis of fluorescent peptide-based probes with aggregation-induced emission characteristic for detecting CH3Hg+ and Hg2+ in aqueous environment: Tuning fluorescent detection for CH3Hg+ by replacing peptide receptors

Even though methylmercury (CH3Hg+) is more toxic than Hg2+, it is more difficult to detect methylmercury than Hg2+ because of less charge and less thiophilicity. In the present study, we developed fluorescent probes for detecting CH3Hg+ and Hg2+ from the fluorescent peptide-based probe (1) for Hg2+ with aggregation induced emission (AIE) characteristic by replacing amino acids of the peptide moiety. The fluorescent probes (2–7) based on dipeptide receptors with AIE characteristics were synthesized for sensing CH3Hg+ and Hg2+ in aqueous environment. Probe 2, 4, 6, and 7 showed turn-on responses to CH3Hg+ and Hg2+ in aqueous solutions. Among them, probe 7 had the most desirable sensing features such as high solubility in water (<1% DMSO), sensitive Off-On response toward Hg2+ and CH3Hg+, high selectivity for Hg2+ and CH3Hg + among tested metal ions, reversibility, fast recognition (∼1 min), significant fluorescent signal changes, and low detection limits (1.4 ppb for Hg2+ and 152 ppb for CH3Hg+). Specially, the specific turn-on fluorescent signals to Hg2+ as well as CH3Hg+ were not significantly interference by coexisting Group I and II metal ions (1 mM) and most other heavy metal ions and the detection limit was lower than the maximum allowable levels of CH3Hg+ in food. The mechanism investigation using NMR, ESI mass spectrometer, and DLS measurements revealed that 7 formed nano-sized aggregates after complexation with Hg2+ and CH3Hg+, resulting in the enhanced fluorescent signals. The dipeptide receptor (AlaHis-OH) of 7 provided potent interactions with Hg2+ as well as CH3Hg+ in aqueous solutions. Probe 7 was successfully applied for the quantification of Hg2+ as well as CH3Hg+ in groundwater samples. We first developed fluorescent probes with AIE characteristics for reversibly detecting Hg2+ and CH3Hg+ in aqueous solution. The relationship between the peptide receptor structures and the specific interactions of Hg2+ and CH3Hg+ would provide useful information for further design of fluorescent detection systems for inorganic and organic mercury.

Nutrient composition of Chenopodium formosanum Koidz. bran: Fractionation and bioactivity of its soluble active polysaccharides.

BackgroundChenopodium formosanum Koidz. Amaranthaceae—also known as Djulis or red quinoa (RQ)—is a cereal plant indigenous to Taiwan, known for its high nutrient value. However, its bran is considered a waste product and the nutrient value has never been analyzed.MethodsIn this study, we examined the proximate composition of RQ bran, specifically its soluble polysaccharide fractions.ResultsRQ bran exhibited high contents of protein (16.56%), ash (7.10%), carbohydrate (60.45%), total polyphenolics (1.85%), betaxanthin (9.91 mg/100 g of RQ bran), and indicaxanthin (7.27 mg/100 g of RQ bran). Specifically, it was rich in polyunsaturated fatty acids (PUFAs; 39.24%)—with an n-6/n-3 and PUFA/saturated fatty acid (SFA) ratio of 18.137 and 0.743, respectively. Four soluble polysaccharide fractions were also obtained: CF-1, CF-2, CF-3, and CF-4, with yields of 3.90%, 6.74%, 22.28%, and 0.06%, respectively, and molecular weights of 32.54, 24.93, 72.39, and 55.45 kDa, respectively. CF-1, CF-2, CF-3, and CF-4 had respectively 15.67%, 42.41%, 5.44%, and 14.52% peptide moiety content and 38.92%, 50.70%, 93.76%, and 19.80% carbohydrate moiety. In CF-2, the glucose content was 95.86 mol% and that of leucine was 16.23%, implicating the presence of a typical leucinoglucan. All four polysaccharide fractions lacked glutamic acid and hydroxyproline. The IC50 of CF-1, CF-2, and CF-3 was respectively 12.05, 3.98, and 14.5 mg/mL for DPPH free radical–scavenging ability; 5.77, 4.10, and 7.03 mg/mL for hydrogen peroxide–scavenging capability; 0.26, 0.05, and 0.19 mg/mL for O2− free radical–scavenging capability; and 100.41, 28.12, and 29.73 mg/mL for Fe2+ chelation.ConclusionOur results indicated that RQ bran has a large amount of nutrient compounds, and a cost-efficient process for their extraction is needed. Their biomedical application as nutraceuticals also warrants further investigation.

Synthesis and biological study of new galanthamine-peptide derivatives designed for prevention and treatment of Alzheimer's disease.

The Alzheimer's disease leads to neurodegenerative processes and affecting negatively million people worldwide. The treatment of the disease is still difficult and incomplete in practice. Galanthamine is one of the most commonly used drugs against the illness. The main aim of this work is design and synthesis of new derivatives of galanthamine comprising peptide moiety as well as study of their β-secretase inhibitory activity and the anti-aggregating effect. All new derivatives of galanthamine containing analogues of Leu-Val-Phe-Phe (Aβ17-Aβ20) were synthesized in solution using fragment and consecutive condensation approaches. The new derivatives were characterized by melting points, NMR, and HPLC/MS. They were tested in vitro for β-secretase inhibition activity by means of fluorescent method and were investigated in vitro for anti-aggregation activity on sheep platelet-rich plasma. Although the new compounds do not contain a structural element responsible for the β-secretase inhibition, five of them show high or good β-secretase inhibitory activity between 19.98 and 51.19% with IC50 between 1.95 and 5.26nM. Four of the new molecules were able to inhibit platelet aggregation between 55.0 and 90.0% with IC50 between 0.69 and 1.36µM. Four of the compounds were able to inhibit platelet aggregation and two of them have high anti-aggregating effects.

Carboxymethylcellulose biofunctionalized ternary quantum dots for subcellular-targeted brain cancer nanotheranostics

Among the most lethal forms of cancer, malignant brain tumors persist as one of the greatest challenges faced by oncologists, where nanotechnology-driven theranostics can play a critical role in developing novel polymer-based supramolecular nanoarchitectures with multifunctional and multi-modal characteristics to fight cancer. However, it is virtually a consensus that, besides the complexity of active delivering anticancer drugs by the nanocarriers to the tumor site, the current evaluation methods primarily relying on in vitro assays and in vivo animal models have been accounted for the low translational effectiveness to clinical applications. In this view, the chick chorioallantoic membrane (CAM) assay has been increasingly recognized as one of the best preclinical models to study the effects of anticancer drugs on the tumor microenvironment (TME). Thus, in this study, we designed, characterized, and developed novel hybrid nanostructures encompassing chemically functionalized carboxymethylcellulose (CMC) with mitochondria-targeting pro-apoptotic peptide (KLA) and cell-penetrating moiety (cysteine, CYS) with fluorescent inorganic semiconductor (Ag-In-S, AIS) for simultaneously bioimaging and inducing glioblastoma cancer cell (U-87 MG, GBM) death. The results demonstrated that the CMC-peptide macromolecules produced supramolecular vesicle-like nanostructures with aqueous colloidal stability suitable as nanocarriers for passive and active targeting of cancer tumors. The optical properties and physicochemical features of the nanoconjugates confirmed their suitability as photoluminescent nanoprobes for cell bioimaging and intracellular tracking. Moreover, the results in vitro demonstrated a notable killing activity towards GBM cells of cysteine-bearing CMC conjugates coupled with pro-apoptotic KLA peptides. More importantly, compared to doxorubicin (DOX), a model anticancer drug in chemotherapy that is highly toxic, these innovative nanohybrids nanoconjugates displayed higher lethality against U-87 MG cancer cells. In vivo CAM assays validated these findings where the nanohybrids demonstrated a significant reduction of GBM tumor progression (41% area) and evidenced an antiangiogenic activity. These results pave the way for developing polymer-based hybrid nanoarchitectonics applied as targeted multifunctional theranostics for simultaneous imaging and therapy against glioblastoma while possibly reducing the systemic toxicity and side-effects of conventional anticancer chemotherapeutic agents.

Allosteric cooperation in β-lactam binding to a non-classical transpeptidase.

L,D-transpeptidase function predominates in atypical 3 → 3 transpeptide networking of peptidoglycan (PG) layer in Mycobacterium tuberculosis. Prior studies of L,D-transpeptidases have identified only the catalytic site that binds to peptide moiety of the PG substrate or β-lactam antibiotics. This insight was leveraged to develop mechanism of its activity and inhibition by β-lactams. Here, we report identification of an allosteric site at a distance of 21 Å from the catalytic site that binds the sugar moiety of PG substrates (hereafter referred to as the S-pocket). This site also binds a second β-lactam molecule and influences binding at the catalytic site. We provide evidence that two β-lactam molecules bind co-operatively to this enzyme, one non-covalently at the S-pocket and one covalently at the catalytic site. This dual β-lactam-binding phenomenon is previously unknown and is an observation that may offer novel approaches for the structure-based design of new drugs against M. tuberculosis.

Cyclic Peptide Protomer Detection in the Gas Phase: Impact on CCS Measurement and Fragmentation Patterns.

With the recent improvements in ion mobility resolution, it is now possible to separate small protomeric tautomers, called protomers. In larger molecules above 1000 Da such as peptides, a few studies suggest that protomers do exist as well and may contribute to their gas-phase conformational heterogeneity. In this work, we observed a CCS distribution that can be explained by the presence of protomers of surfactin, a small lipopeptide with no basic site. Following preliminary density functional theoretical calculations, several protonation sites in the gas phase were energetically favorable in positive ionization mode. Experimentally, at least three near-resolved IM peaks were observed in positive ionization mode, while only one was detected in negative ionization mode. These results were in good agreement with the DFT predictions. CID breakdown curve analysis after IM separation showed different inflection points (CE50) suggesting that different intramolecular interactions were implied in the stabilization of the structures of surfactin. The fragment ratio observed after collision-induced fragmentation was also different, suggesting different ring-opening localizations. All these observations support the presence of protomers on the cyclic peptide moieties of the surfactin. These data strongly suggest that protomeric tautomerism can still be observed on molecules above 1000 Da if the IM resolving power is sufficient. It also supports that the proton localization involves a change in the 3D structure that can affect the experimental CCS and the fragmentation channels of such peptides.

Diarylation of N- and O-nucleophiles through a metal-free cascade reaction

Diarylation of N- and O-nucleophiles through a metal-free cascade reaction

Ribosomally derived lipopeptides containing distinct fatty acyl moieties

Lipopeptides represent a large group of microbial natural products that include important antibacterial and antifungal drugs and some of the most-powerful known biosurfactants. The vast majority of lipopeptides comprise cyclic peptide backbones N-terminally equipped with various fatty acyl moieties. The known compounds of this type are biosynthesized by nonribosomal peptide synthetases, giant enzyme complexes that assemble their products in a non-gene-encoded manner. Here, we report the genome-guided discovery of ribosomally derived, fatty-acylated lipopeptides, termed selidamides. Heterologous reconstitution of three pathways, two from cyanobacteria and one from an arctic, ocean-derived alphaproteobacterium, allowed structural characterization of the probable natural products and suggest that selidamides are widespread over various bacterial phyla. The identified representatives feature cyclic peptide moieties and fatty acyl units attached to (hydroxy)ornithine or lysine side chains by maturases of the GCN5-related N-acetyltransferase superfamily. In contrast to nonribosomal lipopeptides that are usually produced as congener mixtures, the three selidamides are selectively fatty acylated with C10, C12, or C16 fatty acids, respectively. These results highlight the ability of ribosomal pathways to emulate products with diverse, nonribosomal-like features and add to the biocatalytic toolbox for peptide drug improvement and targeted discovery.

Analysis of Oligonucleotide Biodistribution and Metabolization in Experimental Animals.

We describe methods to follow the fate of oligonucleotides after their injection into experimental animals. The quantitation in various tissues, blood or bone marrow cells is possible by chemical ligation PCR. This method works independently of chemical modifications of the oligonucleotide and/or its conjugations to lipid or peptide moieties. Moreover, metabolization intermediates can be detected by mass spectrometry. Together with a readout assay for the biochemical or physiological effects, which will differ, depending on the particular purpose of the oligonucleotide, these methods allow for a comprehensive understanding of oligonucleotide behavior in a living organism.

Evolution of modern age drug discovery of lipopeptides and computer-aided drug discovery in India

The endeavor has been attempted to present a review on the evolution of modern age drug discovery in India. The contribution of next generation therapeutics options microbial metabolites and the computational drug discovery aspects to the global market from India have been represented. Microbial metabolites such as lipopeptides and peptide therapeutics are gaining worldwide importance due to their multiple applications as broad-spectrum antimicrobial, antiviral, anticancer properties etc. Due to the surge of microbial resistance, tumor resistance, and ongoing pandemic due to constantly mutating corona virus, there is a need to develop next-generation therapeutics options from natural origin, less toxic to the environment, and have higher specificity towards target. Small molecule therapeutics are certainly less specific towards cancer targets hence the cytotoxicity is a major issue in cancer treatment while drug resistance due to the mutations are coming as challenges every day for drug discovery researchers. Microbial lipopeptide reserves a sweet spot in between the small molecule inhibitors and peptide therapeutics because of their amphiphilic compounds consist of a fatty acid side chain and a cyclic peptide moiety of hydrophilic nature. The computational drug discovery approach accelerates the drug discovery process due to the advancement in supercomputer facilities provided by various funding agencies such as the Department of Biotechnology (DBT) and the Department of Science and Technology (DST) in India. The current review article is focusing light on the research contribution of Indian Scientists and Govt. of India in the field of lipopeptide-based research and applications of Computer-aided drug discovery.

Targeted β-catenin ubiquitination and degradation by multifunctional stapled peptides.

Aberrant activation of the Wnt signaling pathway has been identified in numerous types of cancer. One common feature of oncogenic Wnt regulation involves an increase in the cellular levels of β-catenin due to interference with its constitutive ubiquitin-dependent degradation. Targeting β-catenin has therefore emerged as an appealing approach for the treatment of Wnt-dependent cancers. Here, we report a strategy that employs multifunctional stapled peptides to recruit an E3 ubiquitin ligase to β-catenin, thereby rescuing β-catenin degradation by hijacking the endogenous ubiquitin-proteasome pathway. Specifically, we designed, synthesized, and evaluated a panel of multifunctional stapled peptides that have a β-catenin binding moiety (StAx-35) covalently linked to a second stapled peptide moiety (SAH-p53-8), which is capable to interact with the E3 ubiquitin ligase MDM2. We found that in vitro these multifunctional peptides can recruit the MDM2 protein to β-catenin and induce poly-ubiquitination on β-catenin. In cellulo, treatment of the human colorectal cancer cell line SW480 with the multifunctional stapled peptides showed dose-dependent degradation of endogenous β-catenin levels. In addition, a luciferase reporter assay showed that the multifunctional stapled peptides can suppress β-catenin-mediated gene expression via the Wnt signaling pathway. Therefore, these multifunctional stapled peptides provide a unique research tool for examining the Wnt signaling pathway by targeted knockdown of β-catenin at the protein level, and may serve as leads for potential drug candidates in the treatment of Wnt-dependent cancers.

Detection and Characterization of the Soybean Allergen Gly m 7 in Soybeans and Processed Soybean Foods

Gly m 7, a novel soybean allergen, was recently reported. In this study, we attempted to detect Gly m 7 in various soybeans and processed soybean foods using raised anti-Gly m 7 antibodies and enzyme-linked streptavidin, specifically binding to the biotin moiety of Gly m 7. There was a large difference in Gly m 7 levels in various soybean-processed foods. When Gly m 7 levels were determined, all cultivars contained this allergen almost completely, but the biotin moiety detected by streptavidin varied, suggesting that biotinylated levels of Gly m 7 might differ among cultivars. The thermal stability of Gly m 7 was determined by heating soybean extracts. During detection using anti-peptide antibodies, detectable intact Gly m 7 was gradually reduced by heating. Gly m 7 was not detected by peptide or biotin detection in worm-wounded soybeans. Soybeans were immersed in distilled water as a pretreatment step for germination, and Gly m 7 levels were compared by immersion time (4–96 h). Intact Gly m 7 was rapidly degraded in detection by both peptide and biotin moieties. This suggested that Gly m 7 was degraded by some protease(s) during germination. These results would be useful for understanding the properties or risk assessment of Gly m 7, a newly discovered soybean allergen.

Molecularly Engineered Aptamers Targeting Tumor Tissue and Cancer Cells for Efficient in Vivo Recognition and Enrichment

Molecularly Engineered Aptamers Targeting Tumor Tissue and Cancer Cells for Efficient in Vivo Recognition and Enrichment

Arginine‐glycine‐aspartate ( RGD ) peptide‐modified graphene as efficient support material for Pt electrocatalyst in proton exchange membrane fuel cells

Graphene with its two-dimensional structure and unique properties has immense potential in energy-related applications such as proton exchange membrane (PEM) fuel cells. Herein, we employ a well-known biomolecule arginine-glycine-aspartate (RGD) peptide–functionalized graphene-supported Pt nanoparticles as an electrocatalyst for PEM fuel cells for the first time. First, chemically reactive graphene oxide (GO) is used as a precursor to covalently functionalize it with RGD peptide through amide bond formation. The amino moieties of RGD peptide on graphene surface serve as ligands and active sites for the nucleation and controlled growth of Pt nanoparticles through polyol reduction method. The homogeneous distribution of ultra-small (about 3 nm) Pt nanoparticles supported on RGD functionalized graphene boosted the hydrogen oxidation reaction (HOR) and oxygen reduction reaction (ORR) electrocatalytic activities by 52% in electrochemical active surface area (ECSA), 112% in mass activity, and 39% in specific activity as compared to unmodified graphene surface. The strong interaction between the metal and the modified support's surface, assisted in evading serious agglomeration and dissolution during 1000 cycles of accelerated degradation tests (ADT), improving the long-term durability of the Pt electrocatalyst by showing about 21% higher ECSA retention than the unmodified support. Fuel cell performance of RGD functionalized graphene-supported Pt nanoparticles also depicted improved power output due to its better Pt utilization and electrocatalytic activity.

Insights into Detoxification of Tolaasins, the Toxins Behind Mushroom Bacterial Blotch, by Microbacterium foliorum NBRC 103072T

Tolaasins are lipodepsipeptides secreted by Pseudomonas tolaasii, the causal agent of brown blotch disease of mushroom, and are the toxins that cause the brown spots. We previously reported that Microbacterium foliorum NBRC 103072T is an effective tolaasin-detoxifying bacterium. In this study, we aimed to characterize the tolaasin-detoxification process of M. foliorum NBRC 103072T. The tolaasin detoxification by M. foliorum NBRC 103072T was carried out by hydrolyzation of tolaasins at two specific sites in the peptide moiety of tolaasins by its cells, and the resulting fragments were released from bacterial cells. The tolaasin-hydrolyzing activity can be extracted by a neutral detergent solution from M. foliorum NBRC 103072T cells. Moreover, tolaasin adsorption to the bacterial cells occurred prior to hydrolyzation of tolaasins, which might contribute to the effective tolaasin detoxification by M. foliorum NBRC 103072T. It is notable that the tolaasin-degradation process by M. foliorum NBRC 103072T is carried out by hydrolyzation at specific sites in the peptide moiety of lipopeptide by bacterial cells as a novel biological degradation process of cyclic lipopeptides. [Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Identification, Isolation, and Characterization of Medipeptins, Antimicrobial Peptides From Pseudomonas mediterranea EDOX.

The plant microbiome is a vastly underutilized resource for identifying new genes and bioactive compounds. Here, we used Pseudomonas sp. EDOX, isolated from the leaf endosphere of a tomato plant grown on a small farm in the Netherlands. To get more insight into its biosynthetic potential, the genome of Pseudomonas sp. EDOX was sequenced and subjected to bioinformatic analyses. The genome sequencing analysis identified strain EDOX as a member of the Pseudomonas mediterranea. In silico analysis for secondary metabolites identified a total of five non-ribosomally synthesized peptides synthetase (NRPS) gene clusters, related to the biosynthesis of syringomycin, syringopeptin, anikasin, crochelin A, and fragin. Subsequently, we purified and characterized several cyclic lipopeptides (CLPs) produced by NRPS, including some of the already known ones, which have biological activity against several plant and human pathogens. Most notably, mass spectrometric analysis led to the discovery of two yet unknown CLPs, designated medipeptins, consisting of a 22 amino acid peptide moiety with varying degrees of activity against Gram-positive and Gram-negative pathogens. Furthermore, we investigated the mode of action of medipeptin A. The results show that medipeptin A acts as a bactericidal antibiotic against Gram-positive pathogens, but as a bacteriostatic antibiotic against Gram-negative pathogens. Medipeptin A exerts its potent antimicrobial activity against Gram-positive bacteria via binding to both lipoteichoic acid (LTA) and lipid II as well as by forming pores in membranes. Collectively, our study provides important insights into the biosynthesis and mode of action of these novel medipeptins from P. mediterranea EDOX.

PD42-01 ENHANCED INTRACELLULAR DELIVERY OF BCG CELL WALL SKELETON INTO BLADDER CANCER CELLS USING LIPOSOMES FUNCTIONALIZED WITH FOLIC ACID AND PEP-1 PEPTIDE

You have accessJournal of UrologyBladder Cancer: Basic Research & Pathophysiology II (PD42)1 Sep 2021PD42-01 ENHANCED INTRACELLULAR DELIVERY OF BCG CELL WALL SKELETON INTO BLADDER CANCER CELLS USING LIPOSOMES FUNCTIONALIZED WITH FOLIC ACID AND PEP-1 PEPTIDE Jae Hun Shim, Joonhee Gook, Se Young Choi, Byung Hoon Chi, Jin Wook Kim, Tae-Hyoung Kim, Soon Chul Myung, and In Ho Chang Jae Hun ShimJae Hun Shim More articles by this author , Joonhee GookJoonhee Gook More articles by this author , Se Young ChoiSe Young Choi More articles by this author , Byung Hoon ChiByung Hoon Chi More articles by this author , Jin Wook KimJin Wook Kim More articles by this author , Tae-Hyoung KimTae-Hyoung Kim More articles by this author , Soon Chul MyungSoon Chul Myung More articles by this author , and In Ho ChangIn Ho Chang More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002056.01AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Although bacillus Calmette–Guérin cell wall skeleton (BCG-CWS) might function as a potential substitute for live BCG, its use in the treatment of bladder cancer remains limited owing to issues such as insolubility and micrometer-size following exposure to an aqueous environment. METHODS: To develop a novel nanoparticulate system for efficient BCG-CWS delivery, liposomal encapsulation was carried out using a modified emulsification-solvent evaporation method (targets: Size, <200 nm; encapsulation efficiency, ∼60%). Further, the liposomal surface was functionalized with specific ligands, folic acid (FA), and Pep-1 peptide (Pep1), as targeting and cell-penetrating moieties, respectively. RESULTS: Functionalized liposomes greatly increased the intracellular uptake of BCGCWS in the bladder cancer cell lines, 5637 and MBT2. The immunoactivity was verified through elevated cytokine production and a THP-1 migration assay. In vivo antitumor efficacy revealed that the BCG-CWS-loaded liposomes effectively inhibited tumor growth in mice bearing MBT2 tumors. Dual ligand-functionalized liposome was also superior to single ligand-functionalized liposomes. Immunohistochemistry supported the enhanced antitumor effect of BCG-CWS, with IL-6 production and CD4 infiltration. CONCLUSIONS: We conclude that FA- and Pep1-modified liposomes encapsulating BCG-CWS might be a good candidate for bladder cancer treatment with high target selectivity. Source of Funding: None © 2021 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 206Issue Supplement 3September 2021Page: e724-e724 Advertisement Copyright & Permissions© 2021 by American Urological Association Education and Research, Inc.MetricsAuthor Information Jae Hun Shim More articles by this author Joonhee Gook More articles by this author Se Young Choi More articles by this author Byung Hoon Chi More articles by this author Jin Wook Kim More articles by this author Tae-Hyoung Kim More articles by this author Soon Chul Myung More articles by this author In Ho Chang More articles by this author Expand All Advertisement Loading ...

Mechanism of LolCDE as a molecular extruder of bacterial triacylated lipoproteins

Lipoproteins are important for bacterial growth and antibiotic resistance. These proteins use lipid acyl chains attached to the N-terminal cysteine residue to anchor on the outer surface of cytoplasmic membrane. In Gram-negative bacteria, many lipoproteins are transported to the outer membrane (OM), a process dependent on the ATP-binding cassette (ABC) transporter LolCDE which extracts the OM-targeted lipoproteins from the cytoplasmic membrane. Lipid-anchored proteins pose a unique challenge for transport machinery as they have both hydrophobic lipid moieties and soluble protein component, and the underlying mechanism is poorly understood. Here we determined the cryo-EM structures of nanodisc-embedded LolCDE in the nucleotide-free and nucleotide-bound states at 3.8-Å and 3.5-Å resolution, respectively. The structural analyses, together with biochemical and mutagenesis studies, uncover how LolCDE recognizes its substrate by interacting with the lipid and N-terminal peptide moieties of the lipoprotein, and identify the amide-linked acyl chain as the key element for LolCDE interaction. Upon nucleotide binding, the transmembrane helices and the periplasmic domains of LolCDE undergo large-scale, asymmetric movements, resulting in extrusion of the captured lipoprotein. Comparison of LolCDE and MacB reveals the conserved mechanism of type VII ABC transporters and emphasizes the unique properties of LolCDE as a molecule extruder of triacylated lipoproteins.