Pharmacokinetics of aspirin: evaluating shortcomings in the literature.

INTRODUCTION: Aspirin is known for its therapeutic benefits in preventing strokes and relieving pain. However, it is toxic to some individuals, and the biological mechanisms causing toxicity are unknown. Limited literature is available on the role of glycine conjugation as the principal pathway in aspirin detoxification. Previous studies have quantified this two-step enzyme reaction as a singular enzymatic process. Consequently, the individual contributions of these enzymes to the kinetics remain unclear. AREAS COVERED: This review summarized the available information on the pharmacokinetics and detoxification of aspirin by the glycine conjugation pathway. Literature searches were conducted using Google Scholar and the academic journal databases accessible through the North-West University Library. Furthermore, the factors affecting interindividual variation in aspirin metabolism and what is known regarding aspirin toxicity were discussed. EXPERT OPINION: The greatest drawback in understanding the pharmacokinetics of aspirin is the limited information available on the substrate preference of the xenobiotic ligase (ACSM) responsible for activating salicylate to salicyl-CoA. Furthermore, previous pharmacokinetic studies did not consider the contribution of other substrates from the diet or genetic variants, to the detoxification rate of glycine conjugation. Impaired glycine conjugation might contribute to adverse health effects seen in Reye's syndrome and cancer.

Small RNA GadY in Escherichia coli enhances conjugation system of IncP-1 by targeting SdiA.

Plasmid-mediated conjugation is a common mechanism for most bacteria to transfer antibiotic resistance genes (ARGs). The conjugative transfer of ARGs is emerging as a major threat to human beings. Although several transfer-related factors are known to regulate this process, small RNAs (sRNAs)-based regulatory roles remain to be clarified. Here, the Hfq-binding sRNA GadY in donor strain Escherichia coli (E. coli) SM10λπ was identified as a new regulator for bacterial conjugation. Two conjugation models established in our previous studies were used, which SM10λπ carrying a chromosomally integrated IncP-1α plasmid RP4 and a mobilizable plasmid pUCP24T served as donor cells, and P. aeruginosa PAO1 or E. coli EC600 as the recipients. GadY was found to promote SM10λπ-PAO1 conjugation by base-pairing with its target mRNA SdiA, an orphan LuxR-type receptor that responds to exogenous N-acylated homoserine lactones (AHLs). However, SM10λπ-EC600 conjugation was not affected due to EC600 lacking AHLs synthase. It indicates that the effects of GadY on conjugation depended on AHLs-SdiA signalling. Further study found GadY bound SdiA to negatively regulate the global RP4 repressors KorA and KorB. When under ciprofloxacin or levofloxacin treatment, GadY expression in donor strain was enhanced, and it positively regulated quinolone-induced SM10λπ-PAO1 conjugation. Thus, our study provides a novel role for sRNA GadY in regulating plasmid-mediated conjugation, which helps us better understand bacterial conjugation to counter antibiotic resistance.

Soluble hemp protein-xylose conjugates fabricated by high-pressure homogenization and pH-shifting treatments.

BACKGROUND: The process of Maillard conjugation occurs with plant proteins and sugars and can be influenced by several factors, such as processing time, pH, and shear force. By utilizing cavitation processes such as high-pressure homogenization (HPH) and pH-shifting, it is possible to regulate the degree of grafting, functional characteristics, and structural changes in the formation of conjugates. The present study aimed to improve the hemp protein concentrate (HPC) through two different conjugation techniques: HPH and pH-shifting-assisted processes. RESULTS: The best conjugation conditions for the conventional method were identified as a 1:2 HPC to xylose ratio, a pH of 10, and 3 h of treatment at 70 °C. The use of HPH and pH 12-shifting methods resulted in a remarkable 2.5-fold increase in grafting degree, requiring less processing time. Fourier transform infrared spectra confirmed the formation of conjugates. Conjugates produced through HPH with pH 12-shifting (MPHX) transformed into soluble glycoproteins with a particle size of 74 nm. MPHX solubility increased by 5.7-fold than HPC, reaching 85.7%, with a more negatively charged surface at -32.4 mV. Microimages showed cracked and sharp forms for conjugated proteins compared to untreated HPC. Additionally, MPHX conjugates demonstrated superior properties in emulsion stability, foaming capacity, and antioxidant activity compared to HPC and classical conjugates. CONCLUSION: The use of HPH and pH-shifting-assisted Maillard conjugation was highly effective in enhancing the functional attributes of hemp protein conjugates. © 2024 The Author(s). Journal of the Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Sequence and origin of the Streptomyces intergenetic-conjugation helper plasmid pUZ8002.

Conjugation of plasmids from Escherichia coli is essential for the genetic manipulation of Streptomyces spp. To facilitate intergeneric conjugation from E. coli to Streptomyces the conjugative machinery required for genetic transfer is usually provided by the non-transferable helper plasmid, pUZ8002. Here we present the complete nucleotide sequence of pUZ8002, describe the previously undocumented creation process, and provide details of the sequence relative to the parental pUZ8 plasmid and another previously published pUZ8002 sequence.

Kinetic models towards an enhanced understanding of diverse ADC conjugation reactions.

The conjugation reaction is the central step in the manufacturing process of antibody-drug conjugates (ADCs). This reaction generates a heterogeneous and complex mixture of differently conjugated sub-species depending on the chosen conjugation chemistry. The parametrization of the conjugation reaction through mechanistic kinetic models offers a chance to enhance valuable reaction knowledge and ensure process robustness. This study introduces a versatile modeling framework for the conjugation reaction of cysteine-conjugated ADC modalities-site-specific and interchain disulfide conjugation. Various conjugation kinetics involving different maleimide-functionalized payloads were performed, while controlled gradual payload feeding was employed to decelerate the conjugation, facilitating a more detailed investigation of the reaction mechanism. The kinetic data were analyzed with a reducing reversed phase (RP) chromatography method, that can readily be implemented for the accurate characterization of ADCs with diverse drug-to-antibody ratios, providing the conjugation trajectories of the single chains of the monoclonal antibody (mAb). Possible kinetic models for the conjugation mechanism were then developed and selected based on multiple criteria. When calibrating the established model to kinetics involving different payloads, conjugation rates were determined to be payload-specific. Further conclusions regarding the kinetic comparability across the two modalities could also be derived. One calibrated model was used for an exemplary in silico screening of the initial concentrations offering valuable insights for profound understanding of the conjugation process in ADC development.

Conjugation Linked PEDOT:PSS with Low Impedance and High Stretchability for Epidermal Electrophysiology.

Long-term epidermal recording of bioelectricity is of paramount importance for personal health monitoring. It requires stretchable and dry film electrodes that can be seamlessly integrated with skin. The simultaneous achievement of high conductivity and skin-like ductility of conducting materials is a prerequisite for reliable signal transduction at the dynamic interface, which is also the bottleneck of epidermal electrophysiology. Here, carbon nanotubes (CNTs) are introduced as "conjugation linkers" into a topologically plasticized conducting polymer (PEDOT:PSS). A thin-film electrode with high conductivity (≈3250 S cm-1) and high stretchability (crack-onset strain>100%) is obtained. In particular, the conjugation linker enables the high volumetric capacitance and the low film resistance, both of which synergically reduce the interfacial impedance. The capabilities of this electrode is further demonstrated in the precise recording of various electrophysiological signals.

Peptide-Carrier Conjugation.

To produce antibodies against synthetic peptides, it is necessary to couple them to a protein carrier. This chapter provides a nonspecialist overview of peptide-carrier conjugation. Furthermore, a protocol for coupling cysteine-containing peptides to bovine serum albumin is outlined.

Solid Phase Peptide Carrier Conjugation.

Conjugation to carrier proteins is necessary for peptides to be able to induce antibody formation when injected into animals together with a suitable adjuvant. This is usually performed by conjugation in solution followed by mixing with the adjuvant. Alternatively, the carrier may be adsorbed onto a solid support followed by activation and conjugation with the peptide by solid-phase chemistry. Different reagents can be used for conjugation through peptide functional groups (-SH, -NH2, -COOH), and various carrier proteins may be used depending on the peptides and the intended use of the antibodies. The solid phase may be an ion exchange matrix, from which the conjugate can subsequently be eluted and mixed with adjuvant. Alternatively, the adjuvant aluminum hydroxide may be used as the solid-phase matrix, whereupon the carrier is immobilized and conjugated with peptide. The resulting adjuvant-carrier-peptide complexes may then be used directly for immunization.

Gene Editing with Artificial DNA Scissors.

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.

Interfacial interaction mechanism between Mn doped highly conjugated biochar and berberine hydrochloride.

MnSO4-modified biochar (Mn-BC) was synthesized to remove berberine hydrochloride (BH) from wastewater by utilizing tea waste as raw material and MnSO4 as modifier. Brunel Emmett Taylor (BET) analysis reveals that the specific surface area (SSA) and average pore size (Dave) of Mn-BC are 1.4 and 7 times higher than those of pristine biochar apart, attributing to the dissociation effect can promote the dispersion of MnSO4 in the pores of the biochar. Meanwhile, the doping of Mn not only introduces additional oxygen-containing functional groups (OCFGs), but also modulates the π electron density. Furthermore, Response surface method (RSM) analysis reveals that Mn-BC dosage has the most significant effect on BH removal, followed by BH concentration and pH value. Kinetic and isothermal studies reveal that the BH adsorption process of Mn-BC was mainly dominated by chemical and monolayer adsorption. Meanwhile, density functional theory (DFT) calculations confirm the contribution of Mn doping to the conjugation effect in the adsorption system. Originally proposed Mn-BC is one potentially propitious material to eliminate BH from wastewater, meanwhile this also provides a newfangled conception over the sustainable utilization of tea waste resources.

Modeling 1-Cyano-4-Dimethylaminopyridine Tetrafluoroborate (CDAP) Chemistry to Design Glycoconjugate Vaccines with Desired Structural and Immunological Characteristics.

Glycoconjugation is a well-established technology for vaccine development: linkage of the polysaccharide (PS) antigen to an appropriate carrier protein overcomes the limitations of PS T-independent antigens, making them effective in infants and providing immunological memory. Glycoconjugate vaccines have been successful in reducing the burden of different diseases globally. However, many pathogens still require a vaccine, and many of them display a variety of glycans on their surface that have been proposed as key antigens for the development of high-valency glycoconjugate vaccines. CDAP chemistry represents a generic conjugation strategy that is easily applied to PS with different structures. This chemistry utilizes common groups to a large range of PS and proteins, e.g., hydroxyl groups on the PS and amino groups on the protein. Here, new fast analytical tools to study CDAP reaction have been developed, and reaction conditions for PS activation and conjugation have been extensively investigated. Mathematical models have been built to identify reaction conditions to generate conjugates with wanted characteristics and successfully applied to a large number of bacterial PSs from different pathogens, e.g., Klebsiella pneumoniae, Salmonella Paratyphi A, Salmonella Enteritidis, Salmonella Typhimurium, Shighella sonnei and Shigella flexneri. Furthermore, using Salmonella Paratyphi A O-antigen and CRM197 as models, a design of experiment approach has been used to study the impact of conjugation conditions and conjugate features on immunogenicity in rabbits. The approach used can be rapidly extended to other PSs and accelerate the development of high-valency glycoconjugate vaccines.

Structural and functional characterization of the bacterial Cap3 enzyme in deconjugation and regulation of the cyclic dinucleotide transferase CD-NTase.

The Cyclic GMP-AMP synthase (cGAS) and cGAS/DncV-like nucleotidyltransferase (CD-NTase) enzymes belong to the key components of the innate immune sensor system that generates cyclic dinucleotide molecules in response to danger signals. Recently, it was discovered that CD-NTase in bacteria can undergo conjugation to protein substrates via an E1/E2 enzyme-mediated process, resembling ubiquitin modification system. Subsequently, these CD-NTase conjugated molecules will be hydrolyzed by the Cap3 enzyme in the same gene cluster. However, the experimental structure of bacterial CD-NTase recognized by Cap3 is unknown. Here, we first determined the crystal structure of the Cap3 enzyme in complex with the C-terminal tail of CD-NTase. Our structural and enzymatic analysis revealed that the C-terminal tail of CD-NTase is both necessary and sufficient for the Cap3-mediated hydrolysis of CD-NTase from its substrates. Interestingly, we further observed that after the hydrolysis reaction, the terminal glycine residue of the CD-NTase C-terminal tail was sequentially removed by Cap3, indicating that Cap3 might play a role in quenching the CD-NTase conjugation reaction. Our work provides experimental evidence elucidating the interaction between Cap3 and CD-NTase, and suggests a potential role for Cap3 in the bacterial Cyclic-oligonucleotide-based anti-phage signaling system (CBASS).

Genomic Characterization of a Clinical NDM-1-Producing Klebsiella michiganensis from Brazil.

Public health faces daily challenges due to increasing reports of pathogenic microorganisms with new antimicrobial resistance. Klebsiella michiganensis, an emerging pathogen, poses difficulty in its identification using conventional techniques. This study presents the first documented case of NDM-1-producing K. michiganensis in Brazil, identified as the new ST418. Initially, the isolate from a tracheal secretion was misidentified as K. oxytoca. However, accurate identification was achieved through ANI analyses. Whole-genome sequencing was conducted to characterize the genetic context of the resistance genes, to identify virulence factors, and to construct a phylogenetic tree. The blaNDM-1 gene was found to be harbored on an IncFIB plasmid approximately 112 kb in length, which was transferable in conjugation assays. The detection of carbapenem resistance genes in this species highlights the importance of public health vigilance, as it may serve as a reservoir and disseminator of significant resistance genes.

Design and synthesis of novel site-specific antibody-drug conjugates that target TROP2.

The approval of Trodelvy® validates TROP2 as a druggable but challenging target for antibody-drug conjugates (ADCs) to treat metastatic triple-negative breast cancer (mTNBC). Here, based on the TROP2-targeted antibody sacituzumab, we designed and developed several site-specific ADC candidates, which employ MMAE (monomethyl auristatin E) as the toxin, via IgG glycoengineering or affinity-directed traceless conjugation. Systematic evaluation of these site-specific ADCs in homogeneity, hydrophilicity, stability, and antitumor efficiency was conducted. The results indicate that the site-specific ADCs gsADC 3b made from one-step glycoengineering exhibit good aggregation stability and in vivo efficacy, providing a new format of ADCs that target TROP2.

Cyanobacteria mediate the dissemination of bacterial antibiotic resistance through conjugal transfer.

Cyanobacterial blooms are expanding world-wide in freshwater and marine environments, and can cause serious ecological and environmental issues, which also contribute to the spread of antibiotic resistance genes (ARGs). However, the mechanistic understanding of cyanobacteria-mediated resistance dynamics is not fully elucidated yet. We selected Microcystis aeruginosa as a model cyanobacteria to illustrate how cyanobacteria mediate the evolution and transfer processes of bacterial antibiotic resistance. The results show that the presence of cyanobacteria significantly decreased the abundance of antibiotic resistant bacteria (ARB) and antibiotic resistant genes (ARGs) by 3% - 99% and 2%-18%, respectively. In addition, it clearly altered bacterial community structure, with the dominant genera evolving from Acinetobacter (27%) and Enterobacter (42%) to Porphyrobacter (59%). The abundance of ARGs positively correlated with Proteobacteria and Firmicutes, rather than Cyanobacteria, and Bacteroidetes. In the presence of cyanobacteria, the transfer events of bacterial resistance genes via conjugation were found to decrease by 10% - 89% (p < 0.05). Surprisingly, we found an extradentary high transfer frequency (about 0.1) for the ARGs via plasmid conjugation from the bacteria into M. aeruginosa population. It confirmed the role of cyanobacterial population as the competent hosts to facilitate ARGs spreading. Our findings provide valuable information on the risk evaluation of ARGs caused by cyanobacterial blooms in aquatic environments, key for the protection and assessment of aquatic environmental quality.

In vitro conjugation of IncB/O-plasmid: Minimum inhibitory concentration of ß-lactams increases 16-fold in Salmonella enterica compared with Escherichia coli.

The use of antimicrobials in poultry leaves residues in the litter, favoring the emergence of antimicrobial-resistant pathogens and making it a source of contamination. An in vitro 4 × 4 factorial trial was performed to investigate the influence of four treatments, consisting of antimicrobial sub-concentrations, on the transference of IncB/O-plasmid through conjugation in four groups. Each group was composed of one plasmid donor bacterium (Escherichia coli H2332) and a recipient bacterium (Escherichia coli J62 or Salmonella enterica serovars, Enteritidis, Typhimurium, or Heidelberg). Our results showed a little decrease in the conjugation frequency in almost all treatments between the two bacterial species, which varied according to each strain. The MIC test revealed an increase of up to 4096-fold in resistance to beta-lactams in Salmonella serovars after plasmid acquisition. This finding suggests that some genetic apparatus may be involved in increased antimicrobial resistance in Salmonella serovars after the acquisition of primary resistance determinants.

Addressing astringency of grape seed extract by covalent conjugation with lupin protein.

Astringency of phenolic-rich foods is a key tactile perception responsible for acceptability/rejection of plant extracts as ingredients in formulations. Covalent conjugation of phenolic extracts with plant proteins might be a promising strategy to control astringency, but suffers from a lack of mechanistic understanding from the lubrication point of view. To shed light on this, this ex vivo study evaluated the effect of conjugation of a phenolic grape seed extract (GSE) with legume protein (lupin, LP) on tribological and surface adsorption performance of GSE in the absence and presence of human saliva (ex vivo). Tribological results confirmed GSE had an inferior lubrication capacity as compared to LP. The lubrication performance of LP-GSE dispersions was comparable to their corresponding LP dispersion (p > 0.05) when covalently conjugated with LP (LP-GSE) with increasing LP:GSE ratio up to 1:0.04 w/w and at a specific degree of conjugation (DC: 2%). Tribological and surface adsorption measurements confirmed the tendency of GSE to interact with human saliva (ex vivo, n = 17 subjects), impairing the lubricity of salivary films. The covalent bonding of LP to GSE hindered GSE's interaction with human saliva, implying the potential influence of covalent conjugation on attenuating astringency. LP appeared to compete with human saliva for surface adsorption and governed the lubrication behaviour in LP-GSE dispersions. Findings from this study provide valuable knowledge to guide the rational design of sustainable, functional foods using conjugation of phenolics with plant proteins to incorporate larger proportions of health-promoting phenolics while controlling astringency, which needs validation by sensory trials.

Gold nanoparticles-conjugation of irisin enhances therapeutic effect by improving cardiac function and attenuating inflammation in sepsis.

Irisin is considered to be a promising therapeutic approach for cardiac depression and inflammatory disorders. The short half-life of irisin impeded its use and drug efficacy in the treatment. This study aimed to examine if pegylated gold nanoparticles-conjugated to irisin would improve therapeutic effects in cecal ligation and puncture (CLP)-induced sepsis in mice. Recombinant irisin were conjugated to a pegylated gold nanoparticle, which was given to mice exposed to CLP. The cecal ligation procedure and sham on mice were operated and assigned to one of following five groups: (I) CLP group: The mouse models underwent the CLP surgical procedure and received only vehicle saline treatment (n = 5); (II) CLP + soluble Irisin: The mouse underwent the CLP and received an intramuscular injection (i.m) (TA) injection of 1 ug of soluble irisin into each tibialis anterior (TA) leg (n = 5); (III) CLP + Gold nanoparticle-conjugated to Irisin: The mouse models underwent the CLP and received an i.m (TA) injection of 1 µg of Gold nanoparticle-irisin via intramuscular injection (TA) into each leg (n = 5); (IV) CLP + Gold nanoparticles- conjugated to IgG: The mouse underwent the CLP and received an i.m (TA) injection of gold nanoparticles conjugated to IgG (n = 5). (V) Sham: The mouse underwent the surgical operation without conducting the CLP (n = 10). The post-operated animals were observed for one week, and survival rates were estimated. Echocardiography was performed to measure cardiac function at 12 h following CLP. TUNEL was employed to detect apoptosis in both cardiac and skeletal muscles; histology was conducted to assess tissue injury in muscles. Enzyme linked immunosorbent assay (ELISA) was conducted to examine release of interleukin 6 (IL6) and the tumor necrosis factor (TNF) alpha. Compared to the CLP control, soluble irisin treatment improved cardiac function recovery, as indicated by the fractional shortening (FS) and ejection fraction (EF). Irisin treatment exhibited reduced IL6 and TNF-alpha release in association with less apoptosis, lower muscle injury index and improved survival post-CLP. However, compared to soluble irisin treatment, gold nanoparticles-conjugated to irisin showed a significant improvement in cardiac function, suppression of apoptosis, reduced IL6 and TNF-alpha releases, decreased muscle injury and an improved survival rate of post-CLP. This study reveals that gold nanoparticles-conjugated irisin can serve to improve irisin's therapeutic effects over a longer course of treatment.

Current trends in development and manufacturing of higher-valent pneumococcal polysaccharide conjugate vaccine and its challenges.

Pneumococcal conjugate vaccines (PCVs) have been developed to protect against pneumococcal diseases caused by the more than 100 serotypes of the bacterium Streptococcus pneumoniae. PCVs primarily prevent pneumococcal infections such as sepsis, bacteraemia, meningitis, otitis media, pneumonia, septicaemia, and sinusitis among infants, adults, elderly, and immunocompromised individuals. The current available PCVs only cover a limited number of serotypes, and there is an immense need for developing higher-valent PCVs that can protect against non-vaccine serotypes to overcome challenges like serotype replacement and antibiotic resistance. The main challenges for developing higher valent PCVs are the complexity of the manufacturing process comprising polysaccharide fermentation, purification, modification or sizing of multiple polysaccharides and conjugation between polysaccharides and carrier proteins, the stability of the conjugates, and the immunogenicity of the vaccine. Different manufacturing processes have been explored to produce higher valent PCVs using different serotypes of S. pneumoniae and conjugation with different carrier proteins. The global coverage of higher valent PCVs are still low, mainly due to the high cost and limited supply of the vaccine. This review focuses on the existing and emerging manufacturing processes and challenges associated with higher-valent pneumococcal PCV development.

Atomistic Molecular Dynamics Simulations of ABA-Type Polymer Peptide Conjugates: Insights into Supramolecular Structures and their Circular Dichroism Spectra.

A combination of atomistic molecular dynamics (aMD) simulations and circular dichroism (CD) analysis is used to explore supramolecular structures of amphiphilic ABA-type triblock polymer peptide conjugates (PPC). Using the example of a recently introduced PPC with pH- and temperature responsive self-assembling behavior [Otter et al., Macromolecular Rapid Communications 2018, 39, 1800459], this study shows how molecular dynamics simulations of simplified fragment molecules can add crucial information to CD data, which helps to correctly identify the self-assembled structures and monitor the folding/unfolding pathways of the molecules. The findings offer insights into the nature of structural transitions induced by external stimuli, thus contributing to the understanding of the connection of microscopic structures with macroscopic properties.