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Inward rectifying potassium (Kir) channels play a critical role in maintaining the resting membrane potential and cellular homeostasis. The high-resolution crystal structure of homotetrameric KirBac1.1 in detergent micelles provides a snapshot of the closed state. Similar to micelles, KirBac1.1 is reported to be in the inactive/closed conformation in POPC membranes. The slide helix of KirBac1.1 is an important structural motif that regulates channel gating. Despite the importance of slide helix in lipid-dependent gating, conflicting models have emerged for the location of slide helix and its structural dynamics in membrane mimetics is poorly understood. Here, we monitored the structural dynamics of the slide helix (residues 46-57) of KirBac1.1 in both DM micelles and POPC membranes utilizing various site-directed fluorescence approaches. We show, using ACMA-based liposome-flux assay, the cysteine mutants of the slide helix are not functional, ensuring the inactive/closed conformation in POPC membranes similar to wild-type channel. Time-resolved fluorescence and water accessibility measurements of NBD-labeled single-cysteine mutants of slide-helix residues suggest that the location of the slide helix at the interfacial region might be shallower in membranes compared to micelles. Interestingly, the slide helix of KirBac1.1 is more dynamic in the physiologically relevant membrane environment, which is accompanied by a differential hydration dynamics throughout the slide helix. Further, REES and lifetime distribution analyses suggest significant changes in conformational heterogeneity of the slide helix in membrane mimetics. Overall, our results give an insight into how membrane mimetics affect the organization and dynamics of slide helix of the closed state of KirBac1.1, and highlight the importance of lipid-protein interactions in membranes., Competing Interests: Declarations. Conflict of interest: The authors declare no competing interests., (© 2025. The Author(s).)

MicroRNAs (miRNAs) are considered reliable biomarkers for a variety of diseases. However, their low abundance in organisms and high sequence similarity of homologous miRNAs make their accurate detection challenging. Here, we constructed a novel fluorescent biosensor for the detection of miRNA-155, a potential biomarker of neuroinflammation, based on duplex-specific nuclease (DSN) assisted amplification and DNA-templated silver nanoclusters (DNA-AgNCs) as fluorescence signal probes. DSN-assisted amplification can transform unstable miRNA into stable DNA and amplify the miRNA signal at the same time. Using DNA-AgNCs as fluorescence signal probes for biosensors can avoid complex labeling processes and reduce costs. The biosensor shows excellent selectivity, reproducibility, a wide linear range (1–600 nM) with a detection limit of 0.86 nM, and potentiality for real sample detection. This work provides a potential universal biosensing platform for miRNA detection. [ABSTRACT FROM AUTHOR]

Persistence of long-term hyperglycemia results in the glyco-oxidation of plasma proteins, which is considered to be a significant factor in metabolic dysfunction, linking hyperglycemia to the emergence of vascular complications. Methylglyoxal (MGO), a dicarbonyl species formed excessively under diabetes, elevates the oxidative stress, enhancing the generation of superoxide anion, which ultimately reacts with nitric oxide (NO•) to form peroxynitrite (PON). PON, being a powerful nitro-oxidizing agent distorts protein structure, hampering its function. This article describes the binding mechanism of thymoquinone (TQ) to fibrinogen (Fg) and its protective effects under simultaneous glyco-nitro-oxidation. Thermodynamic investigations revealed hydrogen bonding and Vander Waal interactions stabilise the complex, confirming its spontaneity and exothermic nature. TQ-induced micro-environmental and structural alterations in fibrinogen were observed by synchronous, 3-D fluorescence maps, and red edge excitation shift (REES). Molecular docking confirmed the wet lab experiments. Previous studies have shown that glycation, as well as nitro-oxidation, modifies the key residues of fibrinogen, leading to its aggregation. Our findings showed that TQ prevented MGO + PON-induced damage to fibrinogen. The current study analyzed the protective effects of TQ on glyco-nitro-oxidized fibrinogen using various biochemical, spectroscopic, and computational methods. NBT assay and carbonyl content revealed glyco-nitro-oxidation-mediated oxidative stress, which was effectively mitigated by TQ in a concentration-dependent manner. The secondary structural alterations in fibrinogen were prevented by TQ as observed by circular dichroism (CD) and Fourier transform infrared spectroscopy (FTIR). Moreover, multiple assays and electron microscopy confirmed structural perturbations leading to the development of fibrillar aggregates that were reduced in TQ treated samples. Our findings project TQ as a potent protective agent against hyperglycemia and related human complications., Competing Interests: Declaration of competing interest The authors have no relevant financial or non-financial interests to disclose., (Copyright © 2025 Elsevier B.V. All rights reserved.)