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The synthesis of novel highly phosphorescent N^C^N tridentate platinum(II)-complex-peptide nucleic acid (PNA) bioconjugates was accomplished through the solid-phase approach. Melting temperature measurements and circular dichroism spectroscopy studies demonstrated that these conjugates maintain the PNA ability to recognize complementary ssDNA and ssRNA, though the length of the spacer between the metal center and the PNA sequence affects their hybridization properties. Noteworthy, the conjugation of PNA to this family of Pt(II) complexes significantly enhanced the luminescent features of the organometallic moiety, leading to increased quantum yields (82.8%, 10–5 M), even in the presence of oxygen (48.6%, 10–5 M). An in vitro cytotoxicity study of Pt(II)-PNA conjugates on HeLa cells showed no significative effect on cell growth in the dark (1 μM for 72 h). [ABSTRACT FROM AUTHOR]

Active cancer targeting consists of the selective recognition of overexpressed biomarkers on cancer cell surfaces or within the tumor microenvironment, enabled by ligands conjugated to drug carriers. Nanoparticle (NP)-based systems are highly relevant for such an approach due to their large surface area which is amenable to a variety of chemical modifications. Over the past decades, several studies have debated the efficiency of passive targeting, highlighting active targeting as a more specific and selective approach. The choice of conjugation chemistry for attaching ligands to nanocarriers is critical to ensure a stable and robust system. Among the panel of cancer biomarkers, the epidermal growth factor receptor (EGFR) stands as one of the most frequently overexpressed receptors in different cancer types. The design and development of nanocarriers with surface-bound anti-EGFR ligands are vital for targeted therapy, relying on their facilitated capture by EGFR-overexpressing tumor cells and enabling receptor-mediated endocytosis to improve drug accumulation within the tumor microenvironment. In this review, we examine several examples of the most recent and significant anti-EGFR nanocarriers and explore the various conjugation strategies for NP functionalization with anti-EGFR biomolecules and small molecular ligands. In addition, we also describe some of the most common characterization techniques to confirm and analyze the conjugation patterns. [ABSTRACT FROM AUTHOR]

The search for new biologically active compounds with prospective pharmaceutical applications has motivated the investigation of alternative synthesis pathways. One such approach involves the development of compounds with established biological activity as lead compounds. The focus on compounds of natural origin is gaining prominence, with steroids and alkaloids representing notable examples. Our research aimed to synthesize novel steroid–alkaloid bioconjugates with potential biological activity. The structure of all new compounds was determined using spectroscopic methods. The final heats of formation (HOF) for all bioconjugates were also calculated. In silico methods demonstrated that most obtained compounds, especially caffeine derivatives, exhibited potential biological activity. These compounds act as cholesterol antagonists, analeptics, antihypercholesterolemic, and respiratory analeptic compounds. The molecular docking results for the 1HWK and 6RZ4 protein domains indicate that the selected bioconjugates exhibit affinities comparable to or lower than those of atorvastatin (−9.6 kcal/mol), the reference ligand in cholesterol-lowering. Conversely, the affinities of the selected bioconjugates are higher than those of caffeine (−6.2 kcal/mol), which is used as the reference ligand for analeptic drugs. [ABSTRACT FROM AUTHOR]

Cellulose tosylate (MCC-Tos) is a key derivative for surface modification and a crucial precursor for cellulose compatibilization in click reactions, enabling its functionalization for advanced applications. Replacing tosyl groups with alkyne groups broadens cellulose's potential in biocompatible reactions, such as thiol-yne click chemistry and protein/enzyme immobilization. To achieve this, we optimized the heterogeneous synthesis of MCC-Tos using a Doehlert matrix statistical design, evaluating the influence and interaction of the reaction conditions. The optimized conditions—144 h reaction time, 10:1 molar ratio, and 30 °C—yielded a degree of substitution for tosyl groups (DStos) of 1.80, determined via elemental analysis and FTIR-ATR spectroscopy. The reaction kinetics followed a first-order model. A subsequent reaction with propargylamine produced aminopropargyl cellulose (MCC-PNH), reducing DStos by 65%, which was confirmed via FTIR, and improving thermal stability by a margin of 30 °C (TGA/DTG). 13C CP/MAS NMR confirmed the alkyne group attachment, further validated via coupling an azide-functionalized coumarin through copper(I)-catalyzed alkyne-azide cycloaddition (CuAAC). Fluorescence microscopy and UV spectroscopy were used to estimate a substitution degree of 0.21. This study establishes a feasible route for synthesizing alkyne-functionalized cellulose, paving the way for eco-friendly materials, including protein/enzyme bioconjugates, composites, and advanced materials via thiol-yne and CuAAC reactions. [ABSTRACT FROM AUTHOR]

During crime scene investigations, numerous traces are secured and may be used as evidence for the evaluation of source and/or activity level propositions. The rapid chemical analysis of a biological trace enables the identification of body fluids and can provide significant donor profiling information, including age, sex, drug abuse, and lifestyle. Such information can be used to provide new leads, exclude from, or restrict the list of possible suspects during the investigative phase. This paper reviews the state-of-the-art labelling techniques to identify the most suitable visual enhancer to be implemented in a lateral flow immunoassay setup for the purpose of trace identification and/or donor profiling. Upon comparison, and with reference to the strengths and limitations of each label, the simplistic one-step analysis of noncompetitive lateral flow immunoassay (LFA) together with the implementation of carbon nanoparticles (CNPs) as visual enhancers is proposed for a sensitive, accurate, and reproducible in situ trace analysis. This approach is versatile and stable over different environmental conditions and external stimuli. The findings of the present comparative analysis may have important implications for future forensic practice. The selection of an appropriate enhancer is crucial for a well-designed LFA that can be implemented at the crime scene for a time- and cost-efficient investigation. [ABSTRACT FROM AUTHOR]

Urease enzyme activity can be related to various issues, both in biological systems and in industrial processes. A significant breakthrough in addressing the challenge to maintain enzymatic activity involves the development of enzyme-polymer based bioconjugates. This entails the utilization of dual responsive polymer matrix involving pH-responsive polymer (2-acrylamido-2-methylpropane sulfonic acid (AMPS)), neutral polymer (N, N-dimethylacrylamide (DMA)), temperature-responsive polymer (N-isopropylacrylamide (NIPAM)), and urease derivative (UD) collectively known as [PAMPS-co-PDMA-co-PUD]-b-PNIPAM (ADUN) centered on the RAFT-Grafting through polymerization process. Kinetic assessments, including a comparative study between the free enzyme and urease linked in ADUN, were conducted using berthelot reagents at different pH and temperature levels. The Lineweaver–Burk plot was used to ascertain the Michaelis–Menten kinetic constant (Km) values, which unveiled strikingly similar Km values for the free enzyme and ADUN. This indicates that urease remains active for over a month when stored at room temperature and up to around 70 °C in the case of ADUN, while it becomes dormant in the case of free urease. The microenvironment of the bioconjugates remains intact for urease in acidic pH conditions and at temperatures ranging from mild to high, when the concentration exceeds the critical micelle concentration (CMC). In general, this approach highlights the prolonged durability and commercial adaptability of ADUN for wider uses. [ABSTRACT FROM AUTHOR]

Chemical protein synthesis enables access to proteins that would otherwise be difficult or impossible to obtain with traditional means such as recombinant expression. Chemoselective ligations provide the ability to join peptide segments prepared by solid‐phase peptide synthesis. While native chemical ligation (NCL) is widely used, it is limited by the need for C‐terminal thioesters with suitable reaction kinetics, properly placed native Cys or thiolated derivatives, and peptide segment solubility at low mM concentrations. Moreover, repetitive purifications to isolate ligated products are often yield‐sapping, hampering efficiency and progress. In this work, we demonstrate the use of Controlled Activation of Peptides for Templated NCL (CAPTN). This traceless multi‐segment templated NCL approach permits the one‐pot synthesis of proteins by harnessing selective thioester activation and orthogonal conjugation chemistries to favor formation of the full‐length ligated product while minimizing side reactions. Importantly, CAPTN provides kinetic enhancements allowing ligations at sterically hindered junctions and low peptide concentrations. Additionally, this one‐pot approach removes the need for intermediate purification. We report the synthesis of two E. coli ribosomal subunits S16 and S17 enabled by the chemical tools described herein. We anticipate that CAPTN will expedite the synthesis of valuable proteins and expand on templated approaches for chemical protein synthesis. [ABSTRACT FROM AUTHOR]

Promoting the generation of triplet states is essential for developing efficient photocatalytic systems. This research presents a novel approach of host‐stabilized through‐space conjugation via the combination of covalent and non‐covalent methods. The designed building block, 4,4′‐(1,4(1,4)‐dibenzene cyclohexaphane‐1,4‐diyl)bis(1‐phenylpyridinium) chloride, features inherently stable through‐space conjugation. When this block forms a 1 : 1 host–guest complex with cucurbit[8]uril, the through‐space conjugation is further stabilized within the confined cavity. Both the generation and lifetime of triplet state are significantly increased, resulting from the host‐stabilized through‐space conjugation. Additionally, the ultrahigh binding constant of 6.58×1014 M−1 ensures the persistence of host‐stabilization effect. As a result, the host–guest complex acts as a highly efficient catalyst in the photocatalytic oxidation of thioether and aromatic alcohol. In the photodegradation of lignin, a complex natural product, the host–guest complex also exhibits high efficiency, demonstrating its robustness. This line of research is anticipated to enrich the toolbox of supramolecular photochemistry and provide a strategy for fabricating efficient supramolecular photocatalysts. [ABSTRACT FROM AUTHOR]

Introduction: Antisense oligonucleotide (ASO) was established as a viable therapeutic option for genetic disorders. ASOs can target RNAs implicated in various diseases, including upregulated mRNA and pre-mRNA undergoing abnormal alternative splicing events. Therapeutic applications of ASOs have been proven with the Food and Drug Administration approval of several drugs in recent years. Earlier enzymatic stability and delivery remains a big challenge for ASOs. Introducing new chemical modifications and new formulations resolving the issues related to the nuclease stability and delivery of the ASOs. Excitingly, ASOs-based bioconjugates that target the hepatocyte have gained much attraction. Efforts are ongoing to increase the therapeutic application of the ASOs to the extrahepatic tissue as well. Area covered: We have briefly discussed the mechanism of ASOs, the development of new chemistries, and delivery strategies for ASO-based drug discovery and development. The discussion focuses more on the already approved ASOs and those in the clinical development stage. Expert opinion: To expand the clinical application of ASOs, continuous effort is required to develop precise delivery strategies for targeting extrahepatic tissue to minimize the off-target effects. [ABSTRACT FROM AUTHOR]

Abstract Gold nanoparticles (AuNPs) and their biocompatible conjugates find wide use as transducers in (bio)sensors and as Nano-pharmaceutics. The study of the interaction between AuNPs and proteins in representative application media helps to better understand their intrinsic behaviors. A multi-environment, multi-parameter screening strategy is proposed based on asymmetric flow field flow fractionation (AF4)-multidetector. Citrate-coated AuNPs ( AuCIT, 25.1 ± 0.2 nm) and PEG-coated AuNPs (AuPEG, 38.3 ± 0.8 nm) were employed with albumin as a model system. Attention was put in investigating the influence of Au/BSA mass ratios, that allowed to identify the yield-maximizing (1:1) and product-maximizing (2.5:1) conditions for the generation of AuNPs-protein conjugates. Furthermore, bioconjugate properties were thoroughly assessed across various saline media with different pH and ionic strengths. While AuNPs with PEG coating exhibit greater stability at high salinities, such as 30 mM, their conjugates are less stable over time. In contrast, although bare AuNPs are significantly affected by pH and salt concentration, once conjugates are formed, their stability surpasses that of the conjugates formed with AuPEG. The developed methodology can fill the vacancy of standard reference quality control (QC) procedures for bioconjugate synthesis and application in (bio)sensors and Nano-pharmaceutics, screening in a short time many combinations, easily scaling up to the semi-preparative scale or translating to different bioconjugates.

Gold nanoparticles (AuNPs) and their biocompatible conjugates find wide use as transducers in (bio)sensors and as Nano-pharmaceutics. The study of the interaction between AuNPs and proteins in representative application media helps to better understand their intrinsic behaviors. A multi-environment, multi-parameter screening strategy is proposed based on asymmetric flow field flow fractionation (AF4)-multidetector. Citrate-coated AuNPs (AuCIT, 25.1 ± 0.2 nm) and PEG-coated AuNPs (AuPEG, 38.3 ± 0.8 nm) were employed with albumin as a model system. Attention was put in investigating the influence of Au/BSA mass ratios, that allowed to identify the yield-maximizing (1:1) and product-maximizing (2.5:1) conditions for the generation of AuNPs-protein conjugates. Furthermore, bioconjugate properties were thoroughly assessed across various saline media with different pH and ionic strengths. While AuNPs with PEG coating exhibit greater stability at high salinities, such as 30 mM, their conjugates are less stable over time. In contrast, although bare AuNPs are significantly affected by pH and salt concentration, once conjugates are formed, their stability surpasses that of the conjugates formed with AuPEG. The developed methodology can fill the vacancy of standard reference quality control (QC) procedures for bioconjugate synthesis and application in (bio)sensors and Nano-pharmaceutics, screening in a short time many combinations, easily scaling up to the semi-preparative scale or translating to different bioconjugates. [ABSTRACT FROM AUTHOR]

Proquinazid is a new-generation fungicide authorized in the EU for combating powdery mildew infections in high-value crops. Due to the perishable nature of fruits, alternative analytical methods are necessary to protect consumer's health from pesticide residues. Currently, immunoassays are a well-established approach for rapidly monitoring chemical contaminants. However, the production of high-quality immunoreagents, such as antibodies and bioconjugates, is essential. This study presents a newly designed hapten that maintains the characteristic moieties of proquinazid unmodified. The linear aliphatic substituents of this molecule were used to introduce the spacer arm. A three-step synthesis strategy was optimized to prepare a hapten that displays the entire 6-iodoquinazolin-4(3H)-one moiety with excellent yields. The N-hydroxysuccimidyl ester of the hapten was activated and purified to prepare a protein conjugate with high hapten density, which was used as an immunogen. Antibodies were raised and competitive enzyme-linked immunosorbent assays were developed. To enhance the assay's sensitivity, two additional heterologous haptens were prepared by modifying the halogenated substituent at C-6. The optimized assays demonstrated low limits of detection in buffer, approximately 0.05 μg/L. When applied to the analysis of proquinazid in QuEChERS extracts of strawberry samples, the immunoassays produced precise and accurate results, particularly in the 10–1000 μg/kg range. [ABSTRACT FROM AUTHOR]

ABSTRACT Antimicrobial peptides (AMPs), derived from numerous life forms, is being recognised as favourable contenders in today's era to overcome the multi drug resistance of cancer cells. Despite of their diverse structural orientations (β‐sheet, α‐helical, loop and extended peptides), they are immensely involved in immune defences and potentially involved for combating cancers and other types of infections via cellular membrane depolarization. Buforins (Bf), the histone H2A derived AMPs along with their analogues (like Bf‐I, Bf‐II and Bf‐IIb) demonstrated substantial anticancer efficacy despite of numerous challenges. They are quite effective in controlling the apoptosis in various cancer cell lines like breast, HeLa, ovarian, lung, liver and prostate cancers. Bf conjugated with bioconjugates were explored for enhancing the bioavailability, drug resistance and stability of these super giants in cancer therapy. The functionalized nanoparticles could possibly help to overwhelm the shortcomings of AMPs, towards cancer therapy. However, the success of in vivo approach may lead to the clinical translation of such therapeutics. In this review, we emphasized on the characteristic features, mechanisms of action, numerous anticancer approaches of Bf. Further discussion was continued with the challenges and their overcoming, advancement and future directions of Bf towards their success in chemotherapy. [ABSTRACT FROM AUTHOR]

Attaching DNA oligonucleotides to gold nanoparticles (AuNPs) to prepare spherical nucleic acids (SNAs) has offered tremendous insights into surface chemistry with resulting bioconjugates serving as critical reagents in biosensors and nanotechnology. While thiolated DNA is generally required to achieve stable conjugates, we herein communicate that using a thermal drying method, a high DNA density and excellent SNA stability was achieved using nonthiolated DNA, rivaling the performance of thiolated DNA such as surviving 1 M NaCl, 2 month stability in 0.3 M NaCl and working in 50 % serum. A poly‐adenine block with as few as two consecutive terminal adenine bases is sufficient for anchoring on AuNPs. By side‐by‐side comparison with the salt‐aging method, the conjugation mechanism was attributed to competitive adenine adsorption at high temperature along with an extremely high DNA concentration upon drying. Bioanalytical applications of nonthiolated SNAs were validated in both solution and paper‐based sensor platforms, facilitating cost‐effective applications for SNAs. [ABSTRACT FROM AUTHOR]

A novel approach to developing lateral flow assays (LFAs) for the detection of CYFRA 21-1 (cytokeratin 19 fragment, a molecular biomarker for epithelial-origin cancers) is proposed. Magnetic bioconjugates (MBCs) were employed in combination with advanced optical and magnetic tools to optimize assay conditions. The approach integrates such techniques as label-free spectral-phase interferometry, colorimetric detection, and ultrasensitive magnetometry using the magnetic particle quantification (MPQ) technique. For the first time in LFA applications, the MPQ-based and colorimetry-based detection methods were compared side by side, and superior analytical performance was demonstrated. The limit of detection (LOD) of 0.9 pg/mL was achieved using MPQ, and 2.9 pg/mL with optical detection. This study has demonstrated that MPQ provides elimination of signal saturation, higher sensitivity (slope of the calibration curve), and a 19-fold wider dynamic range of detected signals. Both optical and magnetic detection results are comparable to the best laboratory-based tests with the added benefits of a 20-min assay duration and the LFA format convenience. The assay effectiveness was validated in human serum and artificial saliva, and high recovery rates were observed. The proposed approach offers rapid and reliable detection of molecular biomarkers and holds significant potential for point-of-care diagnostics, particularly in resource-limited settings. [ABSTRACT FROM AUTHOR]

This review discusses the development and applications of bioconjugates derived from natural hydroxycinnamic acids (HCA), such as coumaric, sinapic, ferulic, and caffeic acids, combined with various biomaterials. These bioconjugates offer a range of benefits including antioxidant properties, UV protection, customized hydrophilic–lipophilic balance, improved safety, solubility, emolliency, biocompatibility, biodegradability, and targeted delivery for biomedical, cosmetic, and food applications. The increasing demand for natural products in the biomedical, cosmetic, and food industries has led to the exploration of these hydroxycinnamic acids and their derivatives. We discuss the synthesis and modification of hydroxycinnamic acids with biomaterials such as ω-hydroxy fatty acids, castor and lesquerella oils, glycerol, isosorbides, and synthetic polyethylene glycol to form functional phenolipids for biomedical, sunscreen, and skincare applications. Encapsulation techniques with β-cyclodextrins and modification of polymeric supports like polysaccharides and starch are discussed for enhancing bioavailability and solubility and targeted delivery. The fine-tuned development of bioconjugates from hydroxycinnamic acids using glycerol to modify the hydrophilic–lipophilic balance, substitution by water-soluble carboxylic acid groups, vegetable oil-based phenolipids, polysaccharides, and PEGylation provide enhanced dual functionalities and offer a promising avenue for creating effective products across various applications. [ABSTRACT FROM AUTHOR]

The combination of natural and synthetic polymers for nanomedicine development had many advantages, including less toxicity, biocompatibility, prolonged circulation, higher stability, and ease of surface modification. Here, a novel folic acid-conjugated Camptothecin-loaded-poly (lactic-co-glycolic) acid-glutenin nanoparticles (FA-CPT-PLGA-Glu NPs) was fabricated to treat breast cancer. FA-CPT-PLGA-Glu NPs target breast cancer cells via upregulated folate receptors and delivered their toxic payloads without disrupting healthy cells. First, CPT-loaded PLGA NPs were created using a modified emulsification/evaporation technique. Second, Glu-based CPT-PLGA NPs were synthesized using a layer-by-layer assembly, and their physiochemical properties were validated. CPT encapsulation efficiency and loading capacity into PLGA-Glu NPs were 74.95 ± 1.34% and 4.78 ± 1.08%, respectively. CPT-PLGA-Glu NPs exhibited sustained and controlled release of loaded-CPT from NPs, and the highest content was released in an acidic environment (pH 5.3), which will be advantageous for cancer treatment. Later, FA-CPT-PLGA-Glu NPs were synthesized by simple conjugation chemistry. The fabricated FA-CPT-PLGA-Glu NPs were around 100 nm in size, with a spherical form and crystalline nature. FA-CPT-PLGA-Glu NPs show strong cytotoxicity activity, and its IC50 value was 16.33 µg × mL− 1 against breast cancer cell line (MCF-7). This folate-receptor-targeted NPs are more effectively internalized into MCF-7 cells, causing ROS generation, cell growth inhibition, and apoptosis. The activity of caspase-3 and − 9 causes MCF-7 cells apoptosis by internalized CPT. Further, internalized CPT induces potential loss of mitochondrial transmembrane and damages the nuclear integrity of the cancer cells. These results showed that the FA-CPT-PLGA-Glu NPs target upregulated folate receptors on the surface of MCF-7 cells.. [ABSTRACT FROM AUTHOR]

Background: Targeted radionuclide therapy with 177Lu-labelled small conjugates is expanding rapidly, and its success is linked to appropriate patient selection. Companion diagnostic conjugates are usually labelled with 68Ga, offering good imaging up to ≈2 h post-injection. However, the optimal tumor-to-background ratio is often reached later. This study examined promising positron-emitting radiometals with half-lives between 3 h and 24 h and β+ intensity (Iβ+) ≥ 15% and compared them to 68Ga. The radiometals included: 43Sc, 44Sc, 45Ti, 55Co, 61Cu, 64Cu, 66Ga, 85mY, 86Y, 90Nb, 132La, 150Tb and 152Tb. 133La (7.2% Iβ+) was also examined because it was recently discussed, in combination with 132La, as a possible diagnostic match for 225Ac. Methods: Total electron and photon doses per decay and per positron; possibly interfering γ-ray emissions; typical activities to be injected for same-day imaging; positron range; and available production routes were examined. Results: For each annihilation process useful for PET imaging, the total energy released (MeV) is: 45Ti (1.5), 43Sc (1.6), 61Cu and 64Cu (1.8), 68Ga (1.9), 44Sc and 133La (2.9), 55Co (3.2), 85mY (3.3), 132La (4.8), 152Tb (6.5), 150Tb (7.1), 90Nb (8.6), and 86Y (13.6). Significant amounts (≥ 10%) of ≈0.5 MeV photons that may fall into the acceptance window of PET scanners are emitted by 55Co, 66Ga, 85mY, 86Y, 132La, and 152Tb. Compton background from more energetic photons would be expected for 44Sc, 55Co, 66Ga, 86Y, 90Nb, 132La,150Tb, and 152Tb. The mean positron ranges (mm) of 64Cu (0.6), 85mY (1.0), 45Ti (1.5), 133La (1.6), 43Sc and 61Cu (1.7), 55Co (2.1), 44Sc and 86Y (2.5), and 90Nb (2.6) were lower than that of 68Ga (3.6). DOTA chelation is applicable for most of the radiometals, though not ideal for 61Cu/64Cu. Recent data showed that chelation of 45Ti with DOTA is feasible. 90Nb requires different complexing agents (e.g., DFO). Finally, they could be economically produced by proton-induced reactions at medical cyclotrons. Conclusion: In particular, 43Sc, 45Ti, and 61Cu have overall excellent β+ decay-characteristics for theranostic applications complementing 177Lu-labelled small conjugates, and they could be sustainably produced. Like Lu, 43Sc, 45Ti and to a lesser extent 61Cu could be labelled with DOTA. [ABSTRACT FROM AUTHOR]

Recent advances in genetics and nucleic acid chemistry have created fundamentally new tools, both for practical applications in therapy and diagnostics and for fundamental genome editing tasks. Nucleic acid-based therapeutic agents offer a distinct advantage of selectively targeting the underlying cause of the disease. Nevertheless, despite the success achieved thus far, there remain unresolved issues regarding the improvement of the pharmacokinetic properties of therapeutic nucleic acids while preserving their biological activity. In order to address these challenges, there is a growing focus on the study of safe and effective delivery methods utilising modified nucleic acid analogues and their lipid bioconjugates. The present review article provides an overview of the current state of the art in the use of chemically modified nucleic acid derivatives for therapeutic applications, with a particular focus on oligonucleotides conjugated to lipid moieties. A systematic analysis has been conducted to investigate the ability of amphiphilic oligonucleotides to self-assemble into micelle-like structures, as well as the influence of non-covalent interactions of such derivatives with serum albumin on their biodistribution and therapeutic effects. [ABSTRACT FROM AUTHOR]

Cyclen‐peptide bioconjugates are usually prepared in multiple steps that require individual preparation and purification of the cyclic peptide and hydrophilic cyclen derivatives. An efficient strategy is discovered for peptide cyclization and functionalization toward lanthanide probe via three components intermolecular crosslinking on solid‐phase peptide synthesis with high conversion yield. Multifunctionality can be conferred by introducing different modular parts or/and metal ions on the cyclen‐embedded cyclopeptide. As a proof‐of‐concept, a luminescent Eu3+ complex and a Gd3+‐based contrasting agent for in vitro optical imaging and in vivo magnetic resonance imaging, respectively, are demonstrated through utilizing this preparation of cyclen‐embedded cyclic arginylglycylaspartic acid (RGD) peptide. [ABSTRACT FROM AUTHOR]

Charge variants are one of the most important quality attributes for protein therapeutics, including antibody drug conjugates (ADCs). ADCs are conjugation products between monoclonal antibodies (mAbs) and highly potent payloads. After attaching a payload, the charge profile of a mAb can be modified due to the change in net charge or surface charge. In this study, we present a unique challenge of charge assay development that arises from a desirable engineering of ADCs that incorporates the hydrolysis-prone succinimide-thioether conjugation chemistry. This engineered hydrolysis at conjugation sites is usually not complete during conjugation process and continuously progressing during mild stress. This hydrolysis also creates a carboxylic functional group, which manifests as acidic peaks in the ADC charge profiles. As a result, ion exchange chromatograms become sensitive measurements of this hydrolysis, which often masks the charge profile change due to other important post-translational modifications. In this study, two approaches were explored to address this unique challenge: to remove the hydrolysis heterogeneity by incubating ADCs under high pH conditions to drive complete hydrolysis; and to analyze charge variants at the subunit level after IdeS digestion. Acceptable charge profiles and quantitative integration results were successfully obtained by both approaches. [Display omitted] • Succinimide-thioether linkage can hydrolyze in ADCs and complicates charge assay. • Hydrolysis kinetics studied; using high pH incubation to induce complete hydrolysis. • An affinity-IEC tandem column separation for ADC charge analysis at subunit level. [ABSTRACT FROM AUTHOR]

A series of indole-1,4-disubstituted-1,2,3-triazole conjugates were synthesised by click chemistry. The haemolytic properties and cytoprotective activity of all the newly synthesised indole-triazole conjugates were tested in vitro. In addition, molecular docking was performed in silico for the selected conjugates to determine their antibacterial and antifungal properties. The results indicate that indole-triazole derivatives effectively protect human erythrocytes against free radical-induced haemolysis in a structure-dependent manner and that bis-indole-bis-triazole derivatives with alkyl linkers are excellent cytoprotective agents against oxidative haemolysis. The tested series of indole-1,4-disubstituted-1,2,3-triazole conjugates may have an affinity for the active sites of specific protein domains (PDB IDs: 2Q85 and 5V5Z) according to molecular docking studies. [ABSTRACT FROM AUTHOR]

Glycoconjugate vaccines have been effective in preventing numerous bacterial infectious diseases and have shown recent potential to treat cancers through active immunotherapy. Soluble polysaccharides elicit short-lasting immune responses and are usually covalently linked to immunogenic carrier proteins to enhance the antigen-specific immune response by stimulating T-cell-dependent mechanisms. Nonetheless, the conjugation of purified polysaccharides to carrier proteins complexifies vaccine production, and immunization with protein glycoconjugates can lead to the undesirable immunogenic interference of the carrier. Recently, the use of nanoparticles and nanoassemblies for the delivery of antigenic saccharides has gathered attention from the scientific community. Nanoparticles can be easily functionalized with a diversity of functionalities, including T-cell epitope, immunomodulator and synthetic saccharides, allowing for the modulation and polarization of the glycoantigen-specific immune response. Notably, the conjugation of glycan to nanoparticles protects the antigens from degradation and enhances their uptake by immune cells. Different types of nanoparticles, such as liposomes assembled from lipids, inorganic nanoparticles, virus-like particles and dendrimers, have been explored for glycovaccine design. The versatility of nanoparticles and their ability to induce robust immune responses make them attractive delivery platforms for antigenic saccharides. The present review aims at summarizing recent advancements in the use of nano-scaled systems for the delivery of synthetic glycoantigens. After briefly presenting the immunological mechanisms required to promote a robust immune response against antigenic saccharides, this review will offer an overview of the current trends in the nanoparticle-based delivery of glycoantigens. [ABSTRACT FROM AUTHOR]

Multivalent meningococcal conjugate vaccines are a significant focus for the scientific community in light of the WHO's mission to defeat meningitidis by 2030. Well-known meningococcal vaccines such as MenAfriVac, Nimenrix, Menveo, and MenQuadfi are licensed in various parts of the world and have been successful. Recently, the World Health Organization (WHO) qualified MenFive (meningococcal A, C, Y, W, and X) conjugate vaccine, further enhancing the battery of vaccines against meningitis. The antigenic nature of the current and new serogroups, the selection of carrier proteins, and the optimal formulation of these biomolecules are pivotal parameters for determining whether a biological preparation qualifies as a vaccine candidate. Creating appropriate quality control analytical tools for a complex biological formulation is challenging. A scoping review aims to identify the main challenges and gaps in analyzing multivalent vaccines, especially in the case of novel serogroups, such as X, as the limited literature addresses these analytical challenges. In summary, the similarities in polysaccharide backbones between meningococcal serogroups (C, Y, W sharing a sialic acid backbone and A, X sharing a phosphorous backbone) along with various conjugation chemistries (such as CNBr activation, reductive amination, CDAP, CPIP, thioether bond formation, N-hydroxy succinimide activation, and carbodiimide-mediated coupling) resulting into a wide variety of polysaccharide -protein conjugates. The challenge in analyzing carrier proteins used in conjugation (such as diphtheria toxoid, tetanus toxoid, CRM diphtheria protein, and recombinant CRM) is assessing their purity (whether they are monomeric or polymeric in nature as well as their polydispersity). Additional analytical challenges include the impact of excipients, potential interference from serogroups, selection and establishment of standards, age-dependent behavior of biomolecules indicated by molecular size distributions, and process-driven variations. This article explains the analytical insights gained (polysaccharide content, free saccharide, free proteins, MSD) during the development of the MenFive vaccine and highlights the crucial gaps and challenges in testing. [ABSTRACT FROM AUTHOR]

The development of peptide drugs has made tremendous progress in the past few decades because of the advancements in modification chemistry and analytical technologies. The novel‐designed peptide drugs have been modified through various biochemical methods with improved diagnostic, therapeutic, and drug‐delivery strategies. Researchers found it a helping hand to overcome the inherent limitations of peptides and bring continued advancements in their applications. Furthermore, the emergence of peptide‐drug conjugates (PDCs)—utilizes target‐oriented peptide moieties as a vehicle for cytotoxic payloads via conjugation with cleavable chemical agents, resulting in the key foundation of the new era of targeted peptide drugs. This review summarizes the various classifications of peptide drugs, suitable chemical modification strategies to improve the ADME (adsorption, distribution, metabolism, and excretion) features of peptide drugs, and recent (2015–early 2024) progress/achievements in peptide‐based drug delivery systems as well as their fruitful implication in preclinical and clinical studies. Furthermore, we also summarized the brief description of other types of PDCs, including peptide‐MOF conjugates and peptide‐UCNP conjugates. The principal aim is to provide scattered and diversified knowledge in one place and to help researchers understand the pinching knots in the science of PDC development and progress toward a bright future of novel peptide drugs. [ABSTRACT FROM AUTHOR]

Nanostructured materials represent promising substrates for biocatalyst immobilization and activation. Cellulose nanocrystals (CNCs), accessible from waste and/or renewable sources, are sustainable and biodegradable, show high specific surface area for anchoring a high number of enzymatic units, and high thermal and mechanical stability. In this work, we present a holistic enzyme‐based approach to functional antibacterial materials by bioconjugation between the lysozyme from chicken egg white and enzymatic cellulose nanocrystals. The neutral CNCs were prepared by endoglucanase hydrolysis from Avicel. We explore the covalent immobilization of lysozyme on enzymatic CNCs and on their TEMPO oxidized derivatives (TO‐CNCs), comparing immobilization yields, material properties, and enzymatic activities. The materials were characterized by X‐ray diffractometry (XRD), attenuated total reflectance Fourier Transform infrared spectroscopy (ATR‐FTIR), bicinchoninic acid (BCA) assay, field‐emission scanning electron microscopy (FE‐SEM) and dynamic light scattering (DLS). We demonstrate the higher overall efficiency of the immobilization process carried out on TO‐CNCs, based on the success of covalent bonding and on the stability of the isolated bioconjugates. [ABSTRACT FROM AUTHOR]

Artificial metallo‐nucleases (AMNs) are small molecule DNA cleavage agents, also known as DNA molecular scissors, and represent an important class of chemotherapeutic with high clinical potential. This review provides a primary level of exploration on the concepts key to this area including an introduction to DNA structure, function, recognition, along with damage and repair mechanisms. Building on this foundation, we describe hybrid molecules where AMNs are covalently attached to directing groups that provide molecular scissors with enhanced or sequence specific DNA damaging capabilities. As this research field continues to evolve, understanding the applications of AMNs along with synthetic conjugation strategies can provide the basis for future innovations, particularly for designing new artificial gene editing systems. [ABSTRACT FROM AUTHOR]

Driven by their exceptional optical characteristics, robust chemical stability, and facile bioconjugation, gold nanoparticles (AuNPs) have emerged as a preferred material for detection and biosensing applications in scientific research. This study involves the development of a simple, rapid, and cost-effective colorimetric immuno-sensing probe to detect aflatoxin B1 and zearalenone using AuNP antibody (AuNP-mAb) conjugates. Anti-toxin antibodies were attached to the AuNPs by using the physical adsorption method. The colorimetric immunosensor developed operates on the principle that the optical properties of the AuNP are very sensitive to aggregation, which can be induced by a critical high salt concentration. Although the presence of antibodies on the AuNP surface inhibits the aggregation, these antibodies bind to the toxin with higher affinity, which leads to exposure of the surface of AuNPs and aggregation in a salt environment. The aggregation triggers a noticeable but variable alteration in color from red to purple and blueish gray, as a result of a red shift in the surface plasmon resonance band of the AuNPs. The extent of the shift is dependent on the toxin exposure dose and can be quantified using a calibration curve through UV–Visible–NIR spectroscopy. The limit of detection using this assay was determined to be as low as 0.15 ng/mL for both zearalenone and aflatoxin B1. The specificity of the prepared immunoprobe was analyzed for a particular mycotoxin in the presence of other mycotoxins. The developed immunoprobe was evaluated for real-world applicability using artificially spiked samples. This colorimetric immunoprobe based on localized surface plasmon resonance (LSPR) has a reduced detection limit compared to other immunoassays, a rapid readout, low cost, and facile fabrication. [ABSTRACT FROM AUTHOR]

RATIONALE: Conjugation sites are a quality attribute of conjugate vaccines. Proteolysis of bioconjugates synthesized by maleimide–thiol chemistry generates type 2 peptides with a hydrolyzed thiosuccinimide linker containing information on the conjugation sites. A mass spectrometry (MS)‐cleavable linker could make the identification of conjugation sites by MS more reliable. METHODS: Four synthetic type 2 peptides with a hydrolyzed thiosuccinimide linker were analyzed by matrix‐assisted laser desorption ionization (MALDI) MS/MS with and without collision gas. These peptides were also partially labeled with 18O in the linker to confirm the proposed fragmentation mechanism. A conjugate vaccine with the hydrolyzed thiosuccinimide linker was reduced and S‐alkylated, digested with trypsin and analyzed by liquid chromatography–MS/MS using collision‐induced dissociation (CID) and higher‐energy collisional dissociation (HCD) fragmentation methods at a normalized collision energy of 30. RESULTS: A metastable fragmentation preferentially cleaves the newly formed pseudopeptide bond within the hydrolyzed thiosuccinimide linker of type 2 peptides to yield P + 71 and C + 98 ions. These ions make the assignment of conjugation sites more reliable. Partial 18O‐labeling and MS/MS analysis confirmed the proposed structures. CID produces these ions as the two most intense signals more favorably than HCD. The latter also yields these ions, guarantees better sequence coverage and promotes other fragmentations in the linker. CONCLUSIONS: Hydrolyzed thiosuccinimide linker is cleavable in MALDI and electrospray ionization MS/MS analysis by a gas‐phase metastable fragmentation. The resulting fragment ions (P + 71 and C + 98) make the identification of conjugation sites more reliable. These results could be extended to self‐hydrolyzing maleimides, which efficiently stabilize the thiosuccinimide linker upon hydrolysis, in antibody–drug conjugates. [ABSTRACT FROM AUTHOR]

This study reports the development of an innovative electrochemical bioplatform for the indirect determination of acrylamide (AM) based on its inhibitory effect on the hybridization of nucleic acids through the formation of AM‐guanine single‐stranded (ssDNA) adducts. The method combines the advantages of the formation and selective immunorecognition of DNA/RNA heteroduplex, with the use of MBs and amperometric transduction using disposable SPCEs. Increasing concentrations of acrylamide prevented the efficient hybridization of a synthetic biotinylated DNA probe immobilized on streptavidin‐coated magnetic beads with its complementary RNA target and the as‐formed heteroduplex recognition with a specific antibody and a secondary HRP‐labeled antibody used for electrochemical readout. The resulting magnetic bioconjugates were magnetically trapped on the surface of the working electrode of a SPCE and the amperometric transduction was carried out in the presence of the H2O2/HQ system, recording a cathodic response inversely proportional to the AM concentration. The developed bioplatform is more competitive with other reported methods in terms of simplicity and assay time, allowing the selective detection of 1 nM AM in 60 min. In addition, the bioplatform was applied to the analysis of potato crisp water extracts. [ABSTRACT FROM AUTHOR]

With the versatile utility of bio-conjugated peptides and proteins in the fields of agriculture, food, cosmetics and pharmaceutical industry, the design of smart protocols to conjugate and modulate biomolecules becomes highly desirable. During this process, the most important consideration for biochemists is the retention of configurational integrity of the biomolecules. Moreover, this type of bioconjugation of peptide and protein becomes frivolous if the reaction is not performed with precise amino acid residues. Hence, chemo-selective, as well as site-selective reactions, that are biocompatible and possess an appropriate level of reactivity are necessary. Based on click chemistry, there are so many tyrosine (Y) conjugation strategies, such as sulfur-fluoride exchange (SuFEx), sulfur-triazole exchange (SuTEx), coupling with π-allyl palladium complexes, diazonium salts, diazodicarboxyamide-based reagents etc. Among these techniques, diazodicarboxyamide-based Y-conjugation, which is commonly known as the "tyrosine-click (Y-click) reaction", has met the expectations of synthetic and biochemists for the tyrosine-specific functionalization of biomolecules. Over the past one and a half decades, significant progress has been made in the classical organic synthesis approach, as well as its biochemical, photochemical, and electrochemical variants. Despite such progress and increasing importance, the Y-click reaction has not been reviewed to document variations in its methodology, applications, and broad utility. The present article aims to provide a summary of the approaches for the modulation of biomolecules at the hotspot of tyrosine residue by employing the Y-click reaction. The article also highlights its application for the mapping of proteins, imaging of living cells, and in the fields of analytical and medicinal chemistry. [ABSTRACT FROM AUTHOR]

Photon-upconversion nanoparticles (UCNP) have already been established as labels for affinity assays in analog and digital formats. Here, advanced, or smart, systems based on UCNPs coated with active shells, fluorescent dyes, and metal and semiconductor nanoparticles participating in energy transfer reactions are reviewed. In addition, switching elements can be embedded in such assemblies and provide temporal and spatial control of action, which is important for intracellular imaging and monitoring activities. Demonstration and critical comments on representative approaches demonstrating the progress in the use of such UCNPs in bioanalytical assays, imaging, and monitoring of target molecules in cells are reported, including particular examples in the field of cancer theranostics. [ABSTRACT FROM AUTHOR]

Advancement in nanotechnology paves new way in nanomedicine via synthesis of various scaffolds bearing multifunctionalities having characteristics of both as diagnostic and therapeutic applications. Quantum dots (QDs) with exceptional luminescent properties exhibited some unbeatable characteristics of small size with ease in surface modifications and thus make them suitable candidates to be used in drug delivery and diagnosis of diseases. Meanwhile, various type of peptides are known in literature for their excellent therapeutic properties and thus make them promising candidates to be used as conjugate partner with the QDs. Highly luminescent properties and surface modifications flexibility of QDs, further urge to use them as promising conjugate partner with many small molecules including peptides and thus found applications in the field of nanomedicines. Thus Peptides ‐QDs conjugates exhibited the properties of both QDs and bioactivity, biocompatibility features of peptides and hence leave behind the limitation of both the individual partners (i. e. QDs and peptides). Thus the combined Peptides‐QDs conjugates acts as theranostic agent for various diseases where these acts as both imaging and drug delivery agent. Hence the present review focused on the theranostic applications of the Peptides‐QDs conjugates in various area of the nanomedicines. Also we focus on the area of improvement and advancement in nanomedicines for the diagnosis and treatment of the diseases in the early stages. [ABSTRACT FROM AUTHOR]

Nitrile‐aminothiol conjugation (NATC) stands out as a promising biocompatible ligation technique due to its high chemo‐selectivity. Herein we investigated the reactivity and substrate scope of NAT conjugation chemistry, thus developing a novel pH dependent orthogonal NATC as a valuable tool for chemical biology. The study of reaction kinetics elucidated that the combination of heteroaromatic nitrile and aminothiol groups led to the formation of an optimal bioorthogonal pairing, which is pH dependent. This pairing system was effectively utilized for sequential and dual conjugation. Subsequently, these rapid (≈1 h) and high yield (>90 %) conjugation strategies were successfully applied to a broad range of complex biomolecules, including oligonucleotides, chelates, small molecules and peptides. The effectiveness of this conjugation chemistry was demonstrated by synthesizing a fluorescently labelled antimicrobial peptide‐oligonucleotide complex as a dual conjugate to imaging in live cells. This first‐of‐its‐kind sequential NATC approach unveils unprecedented opportunities in modern chemical biology, showcasing exceptional adaptability in rapidly creating structurally complex bioconjugates. Furthermore, the results highlight its potential for versatile applications across fundamental and translational biomedical research. [ABSTRACT FROM AUTHOR]

Late-stage specific and selective diversifications of peptides and proteins performed at target residues under ambient conditions are recognized to be the most facile route to various and abundant conjugates. Herein, we report an orthogonal modification of cysteine residues using alkyl thianthreium salts, which proceeds with excellent chemoselectivity and compatibility under mild conditions, introducing a diverse array of functional structures. Crucially, multifaceted bioconjugation is achieved through clickable handles to incorporate structurally diverse functional molecules. This "two steps, one pot" bioconjugation method is successfully applied to label bovine serum albumin. Therefore, our technique is a versatile and powerful tool for late-stage orthogonal bioconjugation. Late-stage specific and selective modifications of peptides and proteins at target residues under ambient conditions are the most facile routes to various bioconjugates. Here, the authors report an orthogonal modification of cysteine residues using alkyl thianthreium salts, which proceeds with excellent chemoselectivity and compatibility under mild conditions, introducing a diverse array of functional structures. [ABSTRACT FROM AUTHOR]

Recently, for the enhancement of the therapeutic qualities of peptides, proteins, small molecules, antibodies, or oligonucleotides, the conjugation technique has immerged as a new domain in drug delivery systems. Conjugated medications have a longer half-life, greater stability, fewer side effects, enhanced solubility, reduced immunogenicity, and a specific targeting approach. The present review discusses various conjugation methods, comparing cleavable and non-cleavable linkers and current advancements in linker technology. It focuses on the molecular characteristics of each component of drug conjugations and linker chemistry techniques. The correlation between in vitro and in vivo studies of these conjugated systems has also been discussed. [ABSTRACT FROM AUTHOR]

Purpose: Multiple myeloma (MM) affects over 35,000 patients each year in the US. There remains a need for versatile Positron Emission Tomography (PET) tracers for the detection, accurate staging, and monitoring of treatment response of MM that have optimal specificity and translational attributes. CD38 is uniformly overexpressed in MM and thus represents an ideal target to develop CD38-targeted small molecule PET radiopharmaceuticals to address these challenges. Procedures: Using phage display peptide libraries and pioneering algorithms, we identified novel CD38 specific peptides. Imaging bioconjugates were synthesized using solid phase peptide chemistry, and systematically analyzed in vitro and in vivo in relevant MM systems. Results: The CD38-targeted bioconjugates were radiolabeled with copper-64 (64Cu) with100% radiochemical purity and an average specific activity of 3.3 – 6.6 MBq/nmol. The analog NODAGA-PEG4-SL022-GGS (SL022: Thr-His-Tyr-Pro-Ile-Val-Ile) had a Kd of 7.55 ± 0.291 nM and was chosen as the lead candidate. 64Cu-NODAGA-PEG4-SL022-GGS demonstrated high binding affinity to CD38 expressing human myeloma MM.1S-CBR-GFP-WT cells, which was blocked by the non-radiolabeled version of the peptide analog and anti-CD38 clinical antibodies, daratumumab and isatuximab, by 58%, 73%, and 78%, respectively. The CD38 positive MM.1S-CBR-GFP-WT cells had > 68% enhanced cellular binding when compared to MM.1S-CBR-GFP-KO cells devoid of CD38. Furthermore, our new CD38-targeted radiopharmaceutical allowed visualization of tumors located in marrow rich bones, remaining there for up to 4 h. Clearance from non-target organs occurred within 60 min. Quantitative PET data from a murine disseminated tumor model showed significantly higher accumulation in the bones of tumor-bearing animals compared to tumor-naïve animals (SUVmax 2.06 ± 0.4 versus 1.24 ± 0.4, P = 0.02). Independently, tumor uptake of the target compound was significantly higher (P = 0.003) compared to the scrambled peptide, 64Cu-NODAGA-PEG4-SL041-GGS (SL041: Thr-Tyr-His-Ile-Pro-Ile-Val). The subcutaneous MM model demonstrated significantly higher accumulation in tumors compared to muscle at 1 and 4 h after tracer administration (SUVmax 0.8 ± 0.2 and 0.14 ± 0.04, P = 0.04 at 1 h; SUVmax 0.89 ± 0.01 and 0.09 ± 0.01, P = 0.0002 at 4 h). Conclusions: The novel CD38-targeted, radiolabeled bioconjugates were specific and allowed visualization of MM, providing a starting point for the clinical translation of such tracers for the detection of MM. [ABSTRACT FROM AUTHOR]

Comprehensive Summary: Protein‐protein interactions (PPIs) play a crucial role in drug discovery and disease treatment. However, the development of effective drugs targeting PPIs remains challenging due to limited methodologies for probing their spatiotemporal anisotropy. Here, we propose a single‐molecule approach using a unique force circuit to investigate PPI dynamics and anisotropy under mechanical forces. Unlike conventional techniques, this approach enables the manipulation and real‐time monitoring of individual proteins at specific amino acids with defined geometry, offering insights into molecular mechanisms at the single‐molecule level. The DNA force circuit was constructed using click chemistry conjugation methods and genetic code expansion techniques, facilitating orthogonal conjugation between proteins and nucleic acids. The SET domain of the MLL1 protein and the tail of histone H3 were used as a model system to demonstrate the application of the DNA force circuit. With the use of atomic force microscopy and magnetic tweezers, optimized assembly procedures were developed. The DNA force circuit provides an exceptional platform for studying the anisotropy of PPIs and holds promise for advancing drug discovery research targeted at PPIs. [ABSTRACT FROM AUTHOR]

The field of coordination chemistry has evolved to intersect with organic chemistry and biochemistry, giving rise to the disciplines of organometallic and bioinorganic chemistry. Nuclear magnetic resonance (NMR) spectroscopy can be applied for characterizing transition metal complexes, spanning both diamagnetic and paramagnetic complexes prevalent in organometallic compounds and metalloproteins. This review offers a comprehensive overview of a wide variety of characterization techniques, ranging from basic 1H and 13C NMR spectroscopy to advanced methods such as heteronuclear experiments, polarization transfer techniques, relaxometry, and multidimensional NMR spectroscopy. The diverse array of NMR spectroscopic methods outlined here promises to enhance our comprehension of transition metal complexes, facilitating the development of innovative catalysts and therapeutics. [ABSTRACT FROM AUTHOR]

This article describes the design, synthesis, and folding behaviors of β‐1,2‐linked glycopolymers as a bioconjugate‐based foldamer. The synthesis of alkyne‐containing β‐glycosyl azides has been achieved via anomeric substitution using tetrabutylammonium azide (Bu4NN3) to 1,2‐anhydro sugar and subsequent esterification with 5‐hexynoic acid. Copper‐catalyzed alkyne‐azide 1,3‐dipolar cycloaddition of the glycosyl azide leads to the formation of β‐1,2‐linked glycopolymer alternating 4‐triazolyl butanoate and tribenzylated d‐galactose (PolyGal) or d‐glucose unit (PolyGlu). The folding and chiroptical properties of the polymers are discussed through UV–vis and CD spectroscopy and anisotropic factors of CD (gCD) values. The results suggest that PolyGal would adopt a helically folded structure in ClCH2CH2Cl (DCE) or THF. Thermal stability of the folded structure of PolyGal in less polar DCE appears to increase compared with that in THF. It is found that the folded structure of PolyGal in THF denatures to a random coil above 40°C. It is indicated that PolyGlu can possibly form the helically folded structure in DCE and adopt a random coil structure at high temperature region. In polar THF, PolyGlu does not fold the conformation at all the observed temperature region. Such phenomena suggest a special role of stereochemistry at the 4 position on the folding behaviors. [ABSTRACT FROM AUTHOR]

Comprehensive Summary: Protein‐protein interactions (PPIs) play a crucial role in drug discovery and disease treatment. However, the development of effective drugs targeting PPIs remains challenging due to limited methodologies for probing their spatiotemporal anisotropy. Here, we propose a single‐molecule approach using a unique force circuit to investigate PPI dynamics and anisotropy under mechanical forces. Unlike conventional techniques, this approach enables the manipulation and real‐time monitoring of individual proteins at specific amino acids with defined geometry, offering insights into molecular mechanisms at the single‐molecule level. The DNA force circuit was constructed using click chemistry conjugation methods and genetic code expansion techniques, facilitating orthogonal conjugation between proteins and nucleic acids. The SET domain of the MLL1 protein and the tail of histone H3 were used as a model system to demonstrate the application of the DNA force circuit. With the use of atomic force microscopy and magnetic tweezers, optimized assembly procedures were developed. The DNA force circuit provides an exceptional platform for studying the anisotropy of PPIs and holds promise for advancing drug discovery research targeted at PPIs. [ABSTRACT FROM AUTHOR]

The search for new biologically active compounds with prospective pharmaceutical applications has motivated the investigation of alternative synthesis pathways. One such approach involves the development of compounds with established biological activity as lead compounds. The focus on compounds of natural origin is gaining prominence, with steroids and alkaloids representing notable examples. Our research aimed to synthesize novel steroid–alkaloid bioconjugates with potential biological activity. The structure of all new compounds was determined using spectroscopic methods. The final heats of formation (HOF) for all bioconjugates were also calculated. In silico methods demonstrated that most obtained compounds, especially caffeine derivatives, exhibited potential biological activity. These compounds act as cholesterol antagonists, analeptics, antihypercholesterolemic, and respiratory analeptic compounds. The molecular docking results for the 1HWK and 6RZ4 protein domains indicate that the selected bioconjugates exhibit affinities comparable to or lower than those of atorvastatin (−9.6 kcal/mol), the reference ligand in cholesterol-lowering. Conversely, the affinities of the selected bioconjugates are higher than those of caffeine (−6.2 kcal/mol), which is used as the reference ligand for analeptic drugs.

Upconversion nanoparticles (UCNPs) are known to possess unique characteristics, which allow them to overcome a number of issues that plague traditional fluorescence probes. UCNPs have been employed in a variety of applications, but it is arguably in the realm of optical sensors where they have shown the most promise. Biomolecule conjugated UCNPs-based fluorescence probes have been developed to detect and quantify a wide range of analytes, from metal ions to biomolecules, with great specificity and sensitivity. In this review, we have given much emphasis on the recent trends and progress in the preparation strategies of bioconjugated UCNPs and their potential application as fluorescence sensors in the trace level detection of food industry-based toxicants and adulterants. The paper discusses the preparation and functionalisation strategies of commonly used biomolecules over the surface of UCNPs. The use of different sensing strategies namely heterogenous and homogenous assays, underlying fluorescence mechanisms in the detection process of food adulterants are summarized in detail. This review might set a precedent for future multidisciplinary research including the development of novel biomolecules conjugated UCNPs for potential applications in food science and technology. [ABSTRACT FROM AUTHOR]

Peptide and protein postmodification have gained significant attention due to their extensive impact on biomolecule engineering and drug discovery, of which cysteine‐specific modification strategies are prominent due to their inherent nucleophilicity and low abundance. Herein, the study introduces a novel approach utilizing multifunctional 5‐substituted 1,2,3‐triazine derivatives to achieve multifaceted bioconjugation targeting cysteine‐containing peptides and proteins. On the one hand, this represents an inaugural instance of employing 1,2,3‐triazine in biomolecular‐specific modification within a physiological solution. On the other hand, as a powerful combination of precision modification and biorthogonality, this strategy allows for the one‐pot dual‐orthogonal functionalization of biomolecules utilizing the aldehyde group generated simultaneously. 1,2,3‐Triazine derivatives with diverse functional groups allow conjugation to peptides or proteins, while bi‐triazines enable peptide cyclization and dimerization. The examination of the stability of bi‐triazines revealed their potential for reversible peptide modification. This work establishes a comprehensive platform for identifying cysteine‐selective modifications, providing new avenues for peptide‐based drug development, protein bioconjugation, and chemical biology research. [ABSTRACT FROM AUTHOR]

The 3,4-dihydroxyphenyl-l-alanine (DOPA) is responsible for the outstanding adhesiveness of foot proteins secreted by mussels. Thanks to DOPA these invertebrates could adhere to various surfaces under wet conditions. Inspired by this natural phenomenon, the DOPA could be bioconjugate with a natural polymer to approach the properties of mussel foot proteins. These bioconjugates can be used to create patches and injectable bioadhesives to stop bleeding in medical emergencies. In this work, the DOPA was bioconjugate with gelatin employing N-hydroxysuccinimide (NHS) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrocloride (EDC) coupling agents. Up to 67% of bioconjugation degree was achieved when the EDC:NHS ratio was set to 1:1. However since the gelatin is highly soluble in water, the bioconjugate is also soluble. To address this, the GD bioconjugate was manufactured into films and crosslinked with either genipin (GP) or (3-glycidyloxypropyl)trimethoxysilane (GPTMS). Fourier transform infrared spectroscopy showed the C–N and Si–O–Si bonds formed after the modification with GP and GPTMS, respectively. After crosslinking, the GD films modified with GP retained around half of the amino groups, in contrast to those modified with GPTMS which severely lost this functional group. the swelling degree was also higher in GD films treated with GP compared to GPTMS. These findings suggest that the GD films crosslinked with GP have potential use as a tissue adhesive. [ABSTRACT FROM AUTHOR]

This publication describes oxazoline–amino acid bioconjugates 1 capable of supramolecular interactions. The bioconjugates contain three main building blocks: an oxazoline ring, a central aromatic unit and an amino acid substituent. Benzene, naphthalene or anthracene with several substitution motifs was used as the central aromatic unit. Two synthetic pathways to β-amino alcohol precursors 3–5 are presented; one-pot synthesis with various coupling reagents is compared to linearly sequenced synthesis using protecting groups. In the final step, a cyclization of precursors 3–5 to oxazolines 1 is described. Single crystal X-ray diffraction of seven oxazoline bioconjugates is reported (1p, 1m6, 1n2, 1n4, 1n5, 1a and 1t4), with an emphasis on supramolecular interactions in the solid state. The capability of bioconjugates 1 to participate in supramolecular interactions in solution was screened by NMR and CD spectroscopy, varying concentrations, temperatures and solvents. The results obtained by crystallography and spectroscopy were further corroborated by computational results for most interesting bioconjugate 1t1. Computational analysis was performed using a CREST/CENSO protocol. In particular, the free energy of formation, ΔG, as well as mean absolute error (MAE) values and correlations of experimental and GIAO calculated NMR parameters have been compared for 1t1 DFT models of monomers and dimers. These results revealed that for 1t1, the supramolecular dimer ensembles are more stable than monomer ensembles. [ABSTRACT FROM AUTHOR]

Asparaginyl ligases have been extensively utilized as valuable tools for site-specific bioconjugation or surface-modification. However, the application is hindered by the laborious and poorly reproducible preparation processes, unstable activity and ambiguous substrate requirements. To address these limitations, this study employed a structure-based rational approach to obtain a high-yield and high-activity protein ligase called OaAEP1-C247A-aa55-351. It was observed that OaAEP1-C247A-aa55-351 exhibits appreciable catalytic activities across a wide pH range, and the addition of the Fe3+ metal ion effectively enhances the catalytic power. Importantly, this study provides insight into the recognition and nucleophile peptide profiles of OaAEP1-C247A-aa55-351. The ligase demonstrates a higher recognition ability for the "Asn-Ala-Leu" motif and an N-terminus "Arg-Leu" as nucleophiles, which significantly increases the reaction yield. Consequently, the catalytic activity of OaAEP1-C247A-aa55-351 with highly efficient recognition and nucleophile motif, "Asn-Ala-Leu" and "Arg-Leu" under the buffer containing Fe3+ is 70-fold and 2-fold higher than previously reported OaAEP1-C247A and the most efficient butelase-1, respectively. Thus, the designed OaAEP1-C247A-aa55-351, with its highly efficient recognition and alternative nucleophile options, holds promising potential for applications in protein engineering, chemo-enzymatic modification, and the development of drugs. Asparaginyl ligases have been utilized as valuable tools for protein engineering, such as through site-specific bioconjugation or surface modification, however, their application is limited due to time-consuming preparation processes and unstable activities. Here, the authors develop a truncated protein ligase OaAEP1-C247A-aa55-351 from an OaAEP1-C247A mutant, which simplifies the preparation steps, tolerates a wider pH range, and enhances the catalytic activities by using efficient recognition and nucleophile motifs. [ABSTRACT FROM AUTHOR]

Click chemistry is a powerful molecular assembly strategy for rapid functional discovery. The development of click reactions with new connecting linkage is of great importance for expanding the click chemistry toolbox. We report the first selenium‐nitrogen exchange (SeNEx) click reaction between benzoselenazolones and terminal alkynes (Se−N to Se−C), which is inspired by the biochemical SeNEx between Ebselen and cysteine (Cys) residue (Se−N to Se−S). The formed selenoalkyne connection is readily elaborated, thus endowing this chemistry with multidimensional molecular diversity. Besides, this reaction is modular, predictable, and high‐yielding, features fast kinetics (k2≥14.43 M−1 s−1), excellent functional group compatibility, and works well at miniaturization (nanomole‐scale), opening up many interesting opportunities for organo‐Se synthesis and bioconjugation, as exemplified by sequential click chemistry (coupled with ruthenium‐catalyzed azide‐alkyne cycloaddition (RuAAC) and sulfur‐fluoride exchange (SuFEx)), selenomacrocycle synthesis, nanomole‐scale synthesis of Se‐containing natural product library and DNA‐encoded library (DEL), late‐stage peptide modification and ligation, and multiple functionalization of proteins. These results indicated that SeNEx is a useful strategy for new click chemistry developments, and the established SeNEx chemistry will serve as a transformative platform in multidisciplinary fields such as synthetic chemistry, material science, chemical biology, medical chemistry, and drug discovery. [ABSTRACT FROM AUTHOR]