Melanin binding of small molecule drugs can lead to targeted disposition to the pigmented tissues and prolonged pharmacological responses in the eye Melanin binding of drugs in vitro correlates with in vivo binding but current workflows for binding affinity require multiple slow steps and analytical method development and or they ...More |Related Solutions: μPulse^®

Melanin binding of small molecule drugs can lead to targeted disposition to the pigmented tissues and prolonged pharmacological responses in the eye. Melanin binding of drugs in vitro correlates with in vivo binding, but current workflows for binding affinity require multiple slow steps, and analytical method development, and/or they may result in high data variability. We developed tangential flow filtration-based methodology to produce size-specific fractions of water-soluble melanin nanoparticles (MNPs), reducing production time from 2-3 days to just a few hours and yielding MNPs with enhanced fluorescence signal. Improved MNPs enabled modifications to a previously published microscale thermophoresis-based melanin binding protocol, shifting analytical focus toward thermophoretic behavior, reducing data variability, and improving reproducibility. The process was tested with nine compounds with varying melanin binding affinities, and the results were consistent with literature, confirming the ability of the method to differentiate compounds based on melanin binding. Fast and reliable workflow will be useful in screening binding affinity for therapeutics and new drug candidates to melanin thereby facilitating ocular drug discovery and construction of predictive pharmacokinetic simulation models. Less |Related Solutions: μPulse^®

SummaryBACH is a transcriptional regulator that modulates various cytoprotective pathways Among these pathways BACH regulates the cellular oxidative stress responses by suppressing the expression of cytoprotective genes Dysregulated BACH activity has been implicated in a range of pathologies including chronic inflammatory diseases fibrosis and cancer making it a promising therapeutic ...More |Related Solutions: Tempest^®

SummaryBACH1 is a transcriptional regulator that modulates various cytoprotective pathways. Among these pathways BACH1 regulates the cellular oxidative stress responses by suppressing the expression of cytoprotective genes. Dysregulated BACH1 activity has been implicated in a range of pathologies, including chronic inflammatory diseases, fibrosis, and cancer, making it a promising therapeutic target. However, BACH1 remains an underexploited drug target, with limited pharmacological inhibitors available. We have developed a novel luciferase-based reporter cell line enabling quantitative, high-throughput assessment of BACH1 inhibition. Using this platform, we rigorously screened two small-molecule libraries with 2,046 compounds and identified four structurally distinct compounds that robustly inhibit BACH1 function. Notably, these compounds simultaneously activate transcription factor NRF2, suggesting the potential for a broader modulation of oxidative stress pathways.Importantly, we demonstrate that commonly used 2D migration assays may fail to detect phenotypes consistent with BACH1 inhibition, resulting in false negatives. In contrast, we establish that 3D invasion assays more robustly capture anti-invasive effects of BACH1 functional inhibition. Using this 3D system, we validate the identified compounds as potent suppressors of lung cancer cell invasion in vitro.This study delivers a novel screening platform for BACH1-targeted drug discovery, and challenges current in vitro standards by establishing 3D invasion assays as a more accurate functional readout for BACH1-targeting compounds. Additionally, it identifies new dual functional BACH1 inhibitors/NRF2 activators, offering novel chemical scaffolds for the development of anti-metastatic therapies and potentially treatments for diseases driven by oxidative stress and inflammation. Less |Related Solutions: Tempest^®

The precise and selective transport of protons across cellular membranes relies on the dynamic formation and dissipation of hydrogen-bonding networks involving water molecules protein sidechains and backbone carbonyls As in aqueous solution protons are conducted over long distances along chains of hydrogen-bonded water molecules within narrow protein pores To engineer ...More |Related Solutions: NT8^®

The precise and selective transport of protons across cellular membranes relies on the dynamic formation and dissipation of hydrogen-bonding networks involving water molecules, protein sidechains, and backbone carbonyls. As in aqueous solution, protons are conducted over long distances along chains of hydrogen-bonded water molecules within narrow protein pores. To engineer proton-conductive pathways, therefore, we must explicitly account for the dynamic behavior of these networks. In previous work, we showed that incorporation of polar Gln residues into hydrophobic pores drives formation of transient, single-file water wires that enable proton-selective transport. Here, we sought to enhance conduction by introducing targeted Ile-to-Ser substitutions to extend connectivity across the pore. We find that the position of Ser relative to Gln modulates sidechain dynamics and, in turn, channel hydration. Although increased polarity reduces hydrophobic length and enhances hydration, these effects alone do not explain the observed conduction rates. Instead, asymmetry in the arrangement and dynamics of polar sidechains emerges as a key determinant of proton conductivity. Together, these results demonstrate that proton conduction is governed not only by pore polarity and hydration, but also by the dynamic and asymmetric organization of hydrogen-bonding networks. This work establishes design principles for engineering proton-selective channels and reveals how asymmetry enables efficient proton transport across biological membranes. Less |Related Solutions: NT8^®

The evolution of transcription factor TF DNA-binding specificity is a major driver of gene regulatory innovation Unlike most TFs which diversify through gene duplication and neofunctionalization the plant-specific LEAFY LFY TF evolved novel binding specificities without extensive duplication Here we combine experimental structural determination and biochemical assays to reveal how ...More |Related Solutions: Rock Imager^®

The evolution of transcription factor (TF) DNA-binding specificity is a major driver of gene regulatory innovation. Unlike most TFs, which diversify through gene duplication and neofunctionalization, the plant-specific LEAFY (LFY) TF evolved novel binding specificities without extensive duplication. Here, we combine experimental structural determination and biochemical assays to reveal how LFY’s dimerization and DNA-binding preferences shifted during the water-to-land transition. We present crystal structures of the LFY DNA-binding domain (DBD) from the hornwort Nothoceros aenigmaticus and the alga Interfilum paradoxum bound to DNA, demonstrating two distinct dimerization mechanisms: one mediated by direct protein-protein interactions and another driven by DNA-mediated cooperativity. In the ancestral state, LFY likely bound DNA as a dimer through DNA-mediated cooperativity, with protein-protein dimerization emerging later, enforcing new DNA-binding preferences. Our findings support a revised evolutionary scenario for LFY, highlighting the dynamic interplay between protein-DNA and protein-protein interactions as key drivers of TF binding specificity. This work deepens our understanding of how structural adaptations in TFs underpin evolutionary transitions in gene regulation. Less |Related Solutions: Rock Imager^®

The richness of our somatosensory experience is reflected in the functional diversity of somatic sensory neurons Single-cell RNA sequencing of sensory neurons has revealed a molecular basis for such diversity However sensory neuron diversity has yet to be captured at the level of the proteome Here we combined electrophysiology with ...More |Related Solutions: Mantis^®

The richness of our somatosensory experience is reflected in the functional diversity of somatic sensory neurons. Single-cell RNA sequencing of sensory neurons has revealed a molecular basis for such diversity1,2,3. However, sensory neuron diversity has yet to be captured at the level of the proteome. Here, we combined electrophysiology with deep visual proteomics 4 to quantify over 6000 proteins from phenotypically-defined sensory neurons in mice and identified proteomic markers of sensory neuron subtypes. Comparative analysis revealed both concordance and meaningful divergence between transcriptomes and proteomes. We further show that up to 3000 proteins can be quantified from one-fourth of a single neuron, demonstrating subset-specific protein signatures. In culture, nociceptive neurons can be acutely sensitized to mechanical stimuli by nerve growth factor (NGF) which normally drives inflammatory pain in vivo5. Indeed, overnight exposure of peptidergic nociceptors to NGF and a protein kinase C (PKC) activator produced functional sensitization associated with proteome changes. Functional knockdown experiments identified the up-regulated B3GNT2 enzyme as a potential effector of nociceptor sensitization. In summary, we present a high-resolution proteomic resource linking molecular identity to function, enabling the discovery of mechanisms underlying somatic sensation and pain sensitization. Less |Related Solutions: Mantis^®