ChemBioChem最新文献

Peptides and proteins are important functional biomolecules both inside and outside of living organisms. The ability to prepare various types of functionalized peptides and proteins is essential for understanding fundamental biological processes, such as protein folding and post-translational modifications (PTMs), and for developing new therapeutics for many diseases, such as cancers and neurodegenerative diseases. The o-aminoaniline moiety was first proposed for activation to a thioester precursor and used for native chemical ligation to prepare large peptides and proteins. In the past decade, the function of o-aminoaniline has been greatly expanded to facilitate the preparation of homogeneously modified peptide and protein samples, where the modifications can include cyclization, C-terminus diversification, etc. Many o-aminoaniline derivatives have also been developed to overcome the inherent limitations of previous versions. In this review, we attempt to summarize the recent developments of different o-aminoaniline derivatives, focusing on their application to the preparation of functional peptide and protein molecules.

Polyphosphate kinases (PPK) play crucial roles in various biological processes, including energy storage and stress responses, through their interaction with inorganic polyphosphate (polyP) and the intracellular nucleotide pool. Members of the PPK family 2 (PPK2s) catalyse polyP‑consuming phosphorylation of nucleotides. In this study, we characterised two PPK2 enzymes from Bacillus cereus (BcPPK2) and Lysinibacillus fusiformis (LfPPK2) to investigate their substrate specificity and potential for selective nucleotide synthesis. Both enzymes exhibited a broad substrate scope, selectively converting over 85% of pyrimidine nucleoside monophosphates (NMPs) to nucleoside diphosphates (NDPs), while nucleoside triphosphate (NTP) formation was observed only with purine NMPs. Preparative enzymatic synthesis of cytidine diphosphate (CDP) was applied to achieve an yield of 49%. Finally, structural analysis of five crystal structures of BcPPK2 and LfPPK2 provided insights into their active sites and substrate interactions. This study highlights PPK2-II enzymes as promising biocatalysts for the efficient and selective synthesis of pyrimidine NDPs.

The Rous sarcoma virus (RSV) is an onco-retrovirus that infects avian species such as the chicken (Gallus gallus). RSV is the first oncovirus to be described, and the oncogenic activity of this virus is related to the expression of a tyrosine kinase that induces carcinogenic transformation. Interestingly, we have noted that the RSV genome contains various potential G4-forming sequences. Among these, two sequences in the GAG and POL genes show high G4-forming potential. Additionally, the SRC oncogene also harbours a putative G4 forming sequence. In this study, we have characterised the G4 formation and topology in these three loci in the RSV-DNA. We have found that these sequences form dynamic G4 structures in physiological conditions, and such dynamicity may be associated with their cellular functions. Further, we have also established that these G4s are recognised by G4 interacting small-molecule ligands and the G4-stabilising protein nucleolin. The binding of these ligands induces structural shifts in the G4s, leading to changes in structure and stability. Thus, the RSV-DNA G4s may be further studied as targets to control its infection and oncogenic effects.

Bacterial infections, particularly those caused by drug-resistant bacteria, represent a pressing global health challenge. During the interaction between pathogen infection and host defense, bacterial infections initiate the host's immune response, which involves the activation of proteases that play a critical role in antibacterial defense. Granzyme B (GzmB), a key immune-related biomarker associated with cytotoxic T lymphocytes (CTLs), plays a pivotal role in this process. Therefore, detecting the activity of GzmB is crucial for understanding the host immune response to bacterial infections and for developing therapeutic strategies to overcome bacterial virulence. In this study, we designed and synthesized three granzyme B-activated near-infrared molecular probes. Among them, the probe HCy-F demonstrates in situ imaging capability, enabling precise quantification of GzmB activity. This development offers a valuable tool for monitoring immune responses and optimizing immunotherapy approaches for combating drug-resistant pathogens.

Vitamin D receptor (VDR) plays a critical role in regulating multiple biological processes, including bone metabolism and cell differentiation, by mediating transcriptional activation in response to ligand binding. We have constructed an environmentally fluorescent probe 2 for VDR to facilitate real-time observation of its ligand-dependent conformational changes in living cells. This probe 2 was synthesized by introducing a dansyl fluorophore via an ethynyl group at the C11 position of 1α,25(OH)₂D₃. Probe 2 exhibited strong environmentally responsive fluorescence, showing increased intensity and a blue shift of the peak wavelength upon binding to VDR due to the increased hydrophobicity of the environment.

Alzheimer's disease (AD) is a progressive neurodegenerative condition characterized by the deposition of amyloid-β (Aβ) peptides, which aggregate into toxic structures such as oligomers, fibrils, and plaques. The presence of these Aβ aggregates in the brain plays a crucial role in the pathophysiology, leading to synaptic dysfunction and cognitive impairment. Understanding how physiological factors affect Aβ aggregation is essential, and therefore, exploring their influence in vitro will likely provide insights into their role in AD pathology. In this study, we investigated the effects of physiological, free amino acids on Aβ aggregation dynamics. We focused on positively charged amino acids, particularly lysine, and employed a chemical modification, methylation, to neutralize its charge. Our analyses revealed that modified lysine significantly reduced Aβ aggregation, indicating that charge distribution of amino acids plays a crucial role in modulating Aβ aggregation behavior. These findings enhance our understanding of the regulatory factors influencing Aβ aggregation and highlight important considerations for future research on Aβ.

Since the building blocks of DNA are nonfluorescent, various external fluorescence reporters have been employed to investigate the structure, dynamics, and function of DNA G-quadruplexes (GQs) and i-motifs (iMs), which play an important role in gene regulation and expression. However, most of those fluorescence reporters lack the ability to provide site-specific structural information of interest. Therefore, it is necessary to develop fluorescent nucleoside analogues that can be covalently inserted into oligonucleotides, which not only serve this purpose, but minimize any potential perturbation towards the native structure of the DNA systems in question. Herein, we characterize the spectroscopic utility of a high quantum yield fluorescent nucleoside analogue, 4-cyanoindole-2'-deoxyribonucleoside (4CNI-NS). We show that (1) incorporation of 4CNI-NS into various oligonucleotides does not alter their ability to fold into their respective native structures, nor does it affect the overall stability of those structures and (2) the fluorescence property of 4CNI-NS is sensitive to its local environment, and the fluorescence intensity and decay kinetics of the 4CNI-NS-containing oligonucleotides exhibit a clear dependence on their secondary structure formation. Collectively, our results demonstrate that 4CNI-NS can be used as a sensitive, isomorphic probe in the study of noncanonical DNA structures.

In recent years, antimicrobial peptides (AMPs) have emerged as a potent weapon against the growing threat of antibiotic resistance. Among AMPs, the ones containing tryptophan (W) and arginine (R) exhibit enhanced antimicrobial properties, benefiting from the unique physicochemical features of the two amino acids. Herein, we designed three hexapeptides, including WR, DWR (_D-isomer), and RF, derived from the original sequence, RWWRWW-NH₂ (RW). By combining sum frequency generation vibrational spectroscopy (SFG-VS) and molecular dynamics (MD) simulation, we examined AMPs' interactions with model bacterial membrane at the molecular level. Our findings revealed the innate different structural features associated with molecular aggregation and membrane activity between _L-(WR, RF and RW) and _D-isomer. The _D-isomer was demonstrated to aggregate via intermolecular hydrogen bonding, which reduced its membrane adsorption quantity and consequently weakened its disruptive effect on the model membrane; while _L-isomers rarely aggregated and thus could fully interact with the model membrane. _D-isomer was proven to lack a stable helical structure, while _L-isomers adopted helical structures, which was believed to be the reason for DWR's tendency to aggregate easily. This study should contribute to designing novel short-chain AMPs with high efficiency, especially in the case that _D-isomers will be used.

Photodynamic therapy (PDT) has emerged as an innovative approach in cancer treatment, effectively inducing tumor cell death through light-triggered reactive oxygen species (ROS) generation. Additionally, PDT can also trigger antitumor immune responses, thereby reducing the risk of postoperative tumor recurrence. However, the development of highly efficient photosensitizers aimed at activating immune responses for comprehensive tumor eradication remains at an early stage. In this study, we developed a new organic photosensitizer, ThC, which exhibits excellent mitochondrial-targeting effect and significantly enhanced ROS production compared to traditional photosensitizers, such as Chlorin e6 (Ce6). Our findings demonstrate that ThC robustly induces immunogenic cell death (ICD) process in hepatic cancer cells, which could effectively transform immunologically "cold" tumors into "hot" tumors. Through in situ injection and subsequent white light irradiation, ThC achieved superior efficacy in eliminating subcutaneous hepatic tumors compared to Ce6 treatment. Immunological analyses revealed that ThC treatment led to elevated levels of CD4+ and CD8+ T cells, along with a reduction in immunosuppressive cell populations (Tregs and tumor-associated macrophages) within the tumor microenvironment. This study provides a novel therapeutic agent with significant potential for clinical translation in the treatment of malignant tumors.

This study describes an enzymatic pathway to produce high purity 4-O-methylglucaric acid from xylan, an underutilized fraction of lignocellulosic biomass. Beechwood xylan was enzymatically hydrolysed using a commercial xylanase and an α-glucuronidase from Amphibacillus xylanus to form 4-O-methylglucuronic acid, which was then purified by anion exchange chromatography and subsequently oxidized to 4-O-methylglucaric acid using a recombinantly produced uronic acid oxidase from Citrus sinensis. Enzymatic oxidation with uronic acid oxidase afforded 95 % yield in 72 hours which is considerably higher than yields previously achieved using a glucooligosaccharide oxidase from Sarocladium strictum. 4-O-methylglucaric acid was isolated by precipitation and purified by recrystallization. Characterization by liquid chromatography mass spectrometry and nuclear magnetic resonance spectroscopy confirmed product identity and high purity (97.8 % w/w). 4-O-methylglucaric acid's performance as a detergent builder was compared to commercial glucaric acid. At 10 : 1 molar ratios of detergent builder to calcium, 4-O-methylglucaric acid provided similar calcium sequestration performance to glucaric acid (less than 5 % difference) at pH 7 and pH 10 in the presence of surfactants, including sodium dodecylbenzene sulfonate. Given their similar calcium sequestration performance, 4-O-methylglucaric acid could effectively substitute for glucaric acid in detergent formulations.