ChemBioChem最新文献

Biocatalysis offers a sustainable alternative for chemical synthesis, but some enzymes, like lipases, still require conventional organic solvents, which are often flammable, toxic, and unsuitable for food or pharmaceutical applications. In this study, we present a systematic approach consisting of solvent selection, design of experiments optimization, and mass transfer analysis. As a case study, caffeic acid phenethyl ester (CAPE), a pharmacologically active compound derived from propolis, can be synthesized enzymatically. In this study, the Novozym 435-catalyzed esterification of caffeic acid and phenethyl alcohol was optimized using p-cymene, a bio-based solvent. To increase productivity, a system combining p-cymene as a solvent and dimethyl sulfoxide (DMSO) as a cosolvent was chosen. The optimal synthesis conditions were found to be 27 mM caffeic acid, 1460.5 mM phenethyl alcohol, and 73°C, achieving a 75.57% CAPE yield. Both external and internal mass transfer effects on the reaction rate were assessed. This study demonstrates the potential of using biocatalysts and green solvents for the sustainable synthesis of CAPE.

Biomineralization is a biological process through which organisms produce mineralized structures to improve their adaptation to environmental challenges. In nature, biomineralization primarily contributes to the formation of hard tissues such as teeth and bone. This process is remarkably complex and involves the self-assembly of proteins into scaffolds that play critical regulatory roles. For instance, collagen molecules self-assemble into fibrils, and their unique quarter-staggered arrangement provides both a spatial template and a defined chemical microenvironment for the nucleation and growth of hydroxyapatite (HAp). Such precise regulation ensures the formation of highly ordered and functional mineralized structures in bone and dentin. This exquisite mechanism highlights the interaction between biological systems and mineralization, demonstrating the refinement of natural evolution and offering inspiration for the development of artificial biomimetic systems. In this review, we first discuss the structural characteristics of self-assembled scaffolds formed by three representative proteins, and their distinct structures provide diverse templates for mineral deposition. Subsequently, we analyze the mineralization mechanisms regulated by these protein scaffolds. Finally, we summarize recent advances in the field of artificial biomimetic mineralization, with particular emphasis on their applications and potential in tissue repair. This overview aims to promote the integration of assembly biopolymer science with advanced biomedical applications.

Effective immune activation against infectious diseases is achieved through the coordinated interplay of peptide antigen presentation and adjuvant-mediated stimulation, including lipid antigen-type adjuvants. For optimal immune activation with viral antigens, we applied covalent antigen-adjuvant complexation at the molecular level, which enables refined modulation of immune responses. This antigen-complex strategy was applied to viral peptide antigens containing sequence regions with lower variability, a particularly valuable approach for rapidly mutating viruses such as influenza. To construct these conjugated antigen complex structures, we used α-GalCer as the lipid antigen adjuvant and optimized linker designs that markedly influenced immunomodulatory activity. Using the precisely synthesized antigen complexes along with human leukocyte antigen (HLA)-transgenic mice, we successfully demonstrated selective immune activation, particularly the peptide antigen-specific CD8⁺ T cell expansion. These methods can  be extended to other viral antigens and may facilitate the development of CD8⁺ T cell-based self-adjuvanting conjugate vaccines.

The studies reported a novel method for synthesizing hafnium-doped tungsten oxide as a sensing platform for clinically crucial serolytic agent, ambroxol. A carbon matrix decorated with synthesized nanostructures exhibited a synergistic effect, displaying high conductivity and a large surface area, which significantly enhanced the oxidative peak current compared to the bare carbon matrix. The analytical performance was evaluated electrochemically employing techniques such as cyclic voltammetry, electrochemical impedance spectroscopy, and square wave voltammetry. Under a wide linear range, the key highlight was low detection limit of 2.55 nM. The fabricated electrode was highly selective, reproducible, and suitable for long-term usage with good stability. Reasonable recovery rates from pharmaceutical and urine samples showed the accuracy and reliability of the sensor for real-world sample analysis. The proposed work is promising in quantifying ambroxol at trace levels, representing a cost-effective and a direct method for clinical analysis and pharmaceutical quantification.

Target identification of bioactive compounds is of significance in life sciences, ranging from molecular biology to drug development. Photoaffinity labeling (PAL), which utilizes ultraviolet (UV) light irradiation to generate a reactive species for covalent bond formation, is the gold standard method for labeling the binding target. However, requirements for UV light irradiation, which can potentially cause denaturation of biomolecules, and uncontrollable reactivity, resulting in nonproductive consumption of the active species, necessitate further improvement of the affinity labeling methodology. Here, we report our studies on the in situ generation of isocyanate from an α-amino hydroxamic acid and a sulfonyl fluoride for affinity labeling. Theoretical and experimental mechanistic studies of the reaction using various α-amino hydroxamic acid derivatives provided a design principle for efficient isocyanate formation. The best α-amino hydroxamic acid showed higher covalent bond-forming efficiency than PAL in model protein modifications.

Nicotinamide adenine dinucleotide (NAD⁺), as an endogenous donor for ADP-ribosylation, can modify DNA, RNA, and proteins, thereby participating in the regulation of the functions of these biomacromolecules. NAD⁺ serves as a reactant in both enzymatic and chemical synthesis. By employing a well-designed reaction process, the synthetic route can be significantly streamlined, enabling the preparation of structurally complex bioactive molecules in a step-saving and highly effective manner. This article reviews the latest research progress in this field. In the field of enzymatic synthesis, a strategy based on the HPF1/PARP1 complex has been developed. Earlier study shows that the recombinant HPF1/PARP1 complex can ADP-ribosylate a variety of substrates in vitro. In the field of chemical synthesis, the focus is on ionic liquid-mediated ADP-ribosylation reactions with controllable α/β configurations of products. These reactions help prepare biologically active ADP-ribosylated (ADPr) peptides from NAD⁺ and commercially available peptides. In addition, this article also outlines the applications of functional NAD⁺ derivatives in enzyme activity analysis and inhibitor development and discusses the challenges faced in this field, such as bio-compatible reaction conditions, synthesis for precise structural control, and structure-activity relationships between stereochemistry and biological functions of more ADPr derivatives.

Sesquiterpene synthases catalyse cyclisations and rearrangements of farnesyl diphosphate to produce a diverse array of sesquiterpenes generated by depronotation and/or water capture. However, the precise mechanisms and dynamics controlling the fate of the final carbocationic intermediate are not well understood. In our previous study, we engineered water capture in selina-4(15),7(11)-diene synthase (SpSdS) to produce selin-7(11)-en-4-ol as a major product at pH 6.0 by point mutation (G305E) in the K_helix region. To develop a more generalised protocol for this functional switch in sesquiterpene synthases, we identified and characterised a novel selina-3,7(11)-diene synthase (AsSdS) from Actinacidiphila soli through multiple sequence alignments which naturally contains glutamate at position 305 (E305). Through site-directed mutagenesis, creating variant G221T, we were able to instigate water capture in AsSdS to produce selin-7(11)-en-4-ol. Our findings identified two crucial regions in the active site pocket of selinadiene synthases: G/E305 in K_helix and T/G221 in H_helix, that have a reproducible effect on product outcome determination. We demonstrate that subtle, yet predictable changes to these residues impact the water capture as well as deprotonation capability of selinadiene synthases and this solvation aspect can be further exploited to engineer other terpene synthases to generate biocatalysts with unique product profiles for diverse applications.

Bacterial peptidoglycan fragments (PGNs) are key signaling molecules in mammalian hosts. A central aspect of understanding their biological functions is the biochemical characterization of PGN recognition by host receptors. Herein, we employed two fluorescent PGN probes, 940-NADA and 940-C1-NBD, to demonstrate their binding to human peptidoglycan recognition protein 1 (hPGRP1) using in vitro fluorescence polarization (FP) assay. Additionally, we used a diverse panel of chemically synthetized or isolated PGNs with varying stem peptide lengths, compositions, and amidation status, which reflect the structural diversity of bacterial peptidoglycan, to investigate their ability to displace 940-NADA from hPGRP1 in an FP displacement assay. Lastly, sequestration of PGNs by hPGRP1 attenuated NOD1 signaling in reporter cells and reduced the production of proinflammatory cytokines in THP-1 cells. Together, these results establish fluorescent PGN probes as a versatile platform for detecting hPGRP1-PGN interactions and provide functional evidence supporting the anti-inflammatory role of hPGRP1 in host innate immunity.

Interactions between peptides based on a region in the zinc finger translocation associated (ZFTA) protein and the Kelch domain of Kelch-like protein 20 (KLHL20^Kelch) have been characterised by biosensor analysis, supported by AlphaFold2-based structure predictions of peptides bound to the protein. Residues critical for the interaction were identified. The analysis showed that all peptides exhibited relatively weak and complex interactions with KLHL20^Kelch. The original ZFTA peptide had a much higher affinity for KLHL20^Kelch than for the Kelch domain of KLHL12 (KLHL12^Kelch), indicating a specificity for KLHL20^Kelch. The estimated K_D ^app of 35 µM was like that for a 21-mer peptide derived from death-associated protein kinase 1, a known KLHL20 substrate. Removal of flexible C-terminal residues generated a 12-mer, predicted to form a stable helix. This reduced the affinity 100-fold. Removal of N-terminal residues resulted in a 10-mer predicted to be flexible, which had a similar affinity as the original 16-mer. The similar affinities for peptides representing different regions of ZFTA suggest that the recognition is feature specific rather than sequence specific. The interaction mechanism reflects "fuzzy binding", consistent with the role of KLHL20 as an adaptor protein in the ubiquitination of disordered protein substrates by Cullin-3 E3 ubiquitin ligase.

This study presents the design to aim for an atom-efficient chemo-enzymatic synthesis route towards aromatic amino alcohols, based on an unspecific peroxygenase-catalysed oxyfunktionalisation of styrene and a highly atom-efficient conversion of the resulting epoxide with nucleophiles and electrophiles, respectively. This synthesis strategy features a simple two-step approach, and the practicality has been demonstrated at a semi-preparative scale. In a first step, the unspecific peroxygenase oxyfunctionalises the substrate, forming an epoxide. Due to its properties, the latter can serve as a starting material for the conversion into a wide range of products, thereby enabling the production of amino alcohols that are otherwise often difficult to synthesise. The shown concept features a one-pot two-step approach, depending on the respective ring-opening reagent. This method aims for a direct synthesis route for the pharmaceutical industry with good yields and high atom efficiency.