Biology subjects

Bista, M.

Publications and source records attributed to Bista, M..

4 recordsLinked to original sources

bioRxiv2025

A Robust Crystallographic Platform for High-Throughput β-Catenin Ligand Discovery

This study presents a robust crystallographic platform for assessing compounds binding to {beta}-catenin. We developed a standardized protein production protocol for the armadillo domain of {beta}-catenin (BC-ARM) and performed biophysical screens using Surface Plasmon Resonance (SPR) and Differential Scanning Fluorimetry (DSF). These findings led to the successful determination of the co-crystal structure of BC-ARM with compound 1 binding to previously reported site but distinct from known transcription factor binding sites. To broaden the search for novel BC binding sites, we utilized FragLites library with a cyclic peptide-stabilized BC-ARM construct. This yielded two high-resolution co-crystal structures identifying a previously unreported binding hotspot. Recognizing the limitations of the cyclic peptide-bound construct for general screening, we designed a novel, truncated BC-ARM construct. This new construct eliminates unstructured regions, reliably producing high-quality, diffracting crystals suitable for high-throughput crystallographic studies. In conclusion, the ligand-bound {beta}-catenin structures and this novel, robust BC-ARM construct establish a powerful platform for further {beta}-catenin investigation.

molecular biology↗

bioRxiv2025

Exploration of chemical probes and conformational flexibility of GID4 - the substrate receptor of human CTLH E3 ligase complex

The application of targeted protein degradation (TPD) is currently constrained by the limited availability of low-molecular-weight molecules that can recruit E3 ligases other than CRBN (Cereblon) or VHL (Von Hippel-Lindau ligase). In this study, we present the structure-based drug design (SBDD) of high-affinity ligands that engage E3 ligase GID4 (Glucose-induced degradation protein 4) in biophysical and cellular experiments. Through structural studies and molecular modeling, we identified three clusters of compounds that induce distinct conformations of GID4. We characterized potential exit vectors and used the most promising ligand as a building block to prepare bifunctional degraders in the form of proteolysis-targeting chimeras (PROTACs). Although ternary complex formation was successful in vitro, degradation of BRD4 was not observed, highlighting the need for further optimization of the degraders. Finally, we theoretically investigated the likelihood of the identified GID4 conformations participating in protein-protein interactions mediated by molecular glue mechanisms. We believe the expanded ligand diversity discovered in this study may pave the way for tuning the selectivity and efficacy of interactions involving GID4 and its neosubstrates. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=61 SRC="FIGDIR/small/662521v3_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@19187e8org.highwire.dtl.DTLVardef@171abd1org.highwire.dtl.DTLVardef@1c75a5forg.highwire.dtl.DTLVardef@fe17fc_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

Expression screen of TNFR1 R347A, MyD88, IRAK4 death domains in E. coli followed by purification and biophysical characterization of TNFR1 R347A death domain

Death domains play a crucial role in signaling pathways related to inflammation and programmed cell death, rendering them promising targets for therapeutic interventions. However, their expression as recombinant proteins often pose challenges. Here, we present expression screening of TNFR1, IRAK4, and MyD88 death domains in E. coli, followed by the biophysical characterization of TNFR1 death domain after subsequent construct optimization. The study also discusses the influence of pH and ionic strength on TNFR1R347A stability, providing statistical models to predict optimal conditions of the buffer to achieve the highest protein stability. HighlightsO_LIOptimization of expression conditions for TNFR1R347A, MyD88, IRAK4 death domains in E. coli BL21(DE3) cells. C_LIO_LIHigh-yield production of soluble monomeric TNFR1R347A death domain. C_LI

biochemistry↗

bioRxiv2019

Characterizing changes in land cover and forest fragmentation in Dhorpatan Hunting Reserve of Nepal from multi-temporal Landsat observations (1993-2018)

Recent centuries have experienced drastic changes in land cover around the world where Himalayan countries like Nepal have undergone changes in the past several decades because of increasing anthropogenic pressure, natural risks and climatic factors. Accordingly, forest fragmentation has also been increasing alarmingly, which is a matter of concern for natural resource management agencies and biodiversity conservation communities. In this study, we assessed land cover change and forest fragmentation trends in Dhorpatan Hunting Reserve of Nepal by implementing landscape fragmentation and recovery process models, and calculating landscape indices based on five-date land cover maps derived from Landsat satellite images from 1993 to 2018. Six land cover types including forest, grass land, barren land, agriculture & built-up, water bodies and snow & glaciers were determined after an intensive field survey. Diverse derived image features were fed to the Support Vector Machines classifier to create land cover maps, followed by a validation procedure using field samples and reference data. Land cover maps showed an increase in forest area from 37.32% (1993) to 39.26% (2018) and snow & glaciers from 1.72% (1993) to 2.15% (2018) while a decrease in grassland area from 38.78% (1993) to 36.41% (2018) and agriculture & built-up area from 2.39% (1993) to 1.80% (2018). Barren land and water body showed negligible changes. The spatial explicit process of forest fragmentation indicated that shrinkage was the most responsible factor of forest loss while expansion was dominant to increment for forest restoration. High dependency of people persists on the reserve for subsistence resources being a cause of forest fragmentation and posing threats to biodiversity. Focus should be made on strategies to decrease the anthropogenic pressure on the reserve. This requires approaches that provide sustainable alternative resources to the local people and innovations that will help them become less reliant on natural resources.

ecology↗