SUMMARY: MOLEonline is an interactive, web-based tool designed to detect and analyse channels (pores and tunnels) within protein structures. The latest version of MOLEonline addresses the limitations of its predecessor by integrating the Mol* viewer for visualization and offering a streamlined, fully interactive user experience. The new features include colouring tunnels in the 3D viewer based on their physicochemical properties. A 2D representation of the protein structure and calculated tunnels is generated using 2DProts. Users can now store tunnels directly in the mmCIF file format, facilitating sharing via the community-standard FAIR format for structural data. In addition, the ability to store and load computation settings ensures the reproducibility of tunnel computation results. Integration with the ChannelsDB 2.0 database allows users to access precomputed tunnels. AVAILABILITY AND IMPLEMENTATION: The MOLEonline application is freely available at https://moleonline.cz with no login requirement, its source code is stored at GitHub under the MIT licence at https://github.com/sb-ncbr/moleonline-web, and archived at Figshare at https://doi.org/10.6084/m9.figshare.29816174.

• MeSH
• Databases, Protein MeSH
• Internet MeSH
• Protein Conformation MeSH
• Proteins * chemistry   MeSH
• Software * MeSH
• User-Computer Interface MeSH
• Computational Biology * methods   MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Proteins * MeSH

Enzymes with buried active sites utilize molecular tunnels to exchange substrates, products, and solvent molecules with the surface. These transport mechanisms are crucial for protein function and influence various properties. As proteins are inherently dynamic, their tunnels also vary structurally. Understanding these dynamics is essential for elucidating structure-function relationships, drug discovery, and bioengineering. Caver Web 2.0 is a user-friendly web server that retains all Caver Web 1.0 functionalities while introducing key improvements: (i) generation of dynamic ensembles via automated molecular dynamics with YASARA, (ii) analysis of dynamic tunnels with CAVER 3.0, (iii) prediction of ligand trajectories in multiple snapshots with CaverDock 1.2, and (iv) customizable ligand libraries for virtual screening. Users can assess protein flexibility, identify and characterize tunnels, and predict ligand trajectories and energy profiles in both static and dynamic structures. Additionally, the platform supports virtual screening with FDA/EMA-approved drugs and user-defined datasets. Caver Web 2.0 is a versatile tool for biological research, protein engineering, and drug discovery, aiding the identification of strong inhibitors or new substrates to bind to the active sites or tunnels, and supporting drug repurposing efforts. The server is freely accessible at https://loschmidt.chemi.muni.cz/caverweb.

• MeSH
• Internet MeSH
• Catalytic Domain MeSH
• Protein Conformation MeSH
• Ligands MeSH
• Drug Discovery MeSH
• Proteins * chemistry   metabolism   MeSH
• Molecular Dynamics Simulation MeSH
• Software * MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Ligands MeSH
• Proteins * MeSH

ChannelsDB 2.0 is an updated database providing structural information about the position, geometry and physicochemical properties of protein channels-tunnels and pores-within deposited biomacromolecular structures from PDB and AlphaFoldDB databases. The newly deposited information originated from several sources. Firstly, we included data calculated using a popular CAVER tool to complement the data obtained using original MOLE tool for detection and analysis of protein tunnels and pores. Secondly, we added tunnels starting from cofactors within the AlphaFill database to enlarge the scope of the database to protein models based on Uniprot. This has enlarged available channel annotations ∼4.6 times as of 1 September 2023. The database stores information about geometrical features, e.g. length and radius, and physico-chemical properties based on channel-lining amino acids. The stored data are interlinked with the available UniProt mutation annotation data. ChannelsDB 2.0 provides an excellent resource for deep analysis of the role of biomacromolecular tunnels and pores. The database is available free of charge: https://channelsdb2.biodata.ceitec.cz.

• MeSH
• Amino Acids MeSH
• Databases, Protein * MeSH
• Protein Conformation MeSH
• Proteins * chemistry   MeSH
• Software * MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Amino Acids MeSH
• Proteins * MeSH