The receptor-binding domain (RBD) of the SARS-CoV-2 spike protein (RBD_CoV2) has a higher binding affinity to the human receptor angiotensin-converting enzyme 2 (ACE2) than the SARS-CoV RBD (RBD_CoV). Here, we performed molecular dynamics (MD) simulations, binding free energy (BFE) calculations, and interface residue contact network (IRCN) analysis to explore the mechanistic origin of different ACE2-binding affinities of the two RBDs. The results demonstrate that, when compared to the RBD_CoV2-ACE2 complex, RBD_CoV-ACE2 features enhanced dynamicsand inter-protein positional movements and increased conformational entropy and conformational diversity. Although the inter-protein electrostatic attractive interactions are the primary determinant for the high ACE2-binding affinities of both RBDs, the significantly enhanced electrostatic attractive interactions between ACE2 and RBD_CoV2 determine the higher ACE2-binding affinity of RBD_CoV2 than of RBD_CoV. Comprehensive comparative analyses of the residue BFE components and IRCNs between the two complexes reveal that it is the residue changes at the RBD interface that lead to the overall stronger inter-protein electrostatic attractive force in RBD_CoV2-ACE2, which not only tightens the interface packing and suppresses the dynamics of RBD_CoV2-ACE2, but also enhances the ACE2-binding affinity of RBD_CoV2. Since the RBD residue changes involving gain/loss of the positively/negatively charged residues can greatly enhance the binding affinity, special attention should be paid to the SARS-CoV-2 variants carrying such mutations, particularly those near or at the binding interfaces with the potential to form hydrogen bonds and/or salt bridges with ACE2.

The indispensable role of the SARS-CoV-2 main protease (Mpro) in the viral replication cycle and its dissimilarity to human proteases make Mpro a promising drug target. In order to identify the non-covalent Mpro inhibitors, we performed a comprehensive study using a combined computational strategy. We first screened the ZINC purchasable compound database using the pharmacophore model generated from the reference crystal structure of Mpro complexed with the inhibitor ML188. The hit compounds were then filtered by molecular docking and predicted parameters of drug-likeness and pharmacokinetics. The final molecular dynamics (MD) simulations identified three effective candidate inhibitors (ECIs) capable of maintaining binding within the substrate-binding cavity of Mpro. We further performed comparative analyses of the reference and effective complexes in terms of dynamics, thermodynamics, binding free energy (BFE), and interaction energies and modes. The results reveal that, when compared to the inter-molecular electrostatic forces/interactions, the inter-molecular van der Waals (vdW) forces/interactions are far more important in maintaining the association and determining the high affinity. Given the un-favorable effects of the inter-molecular electrostatic interactions—association destabilization by the competitive hydrogen bond (HB) interactions and the reduced binding affinity arising from the un-compensable increase in the electrostatic desolvation penalty—we suggest that enhancing the inter-molecular vdW interactions while avoiding introducing the deeply buried HBs may be a promising strategy in future inhibitor optimization.

The human transmembrane protease, serine 2 (TMPRSS2), essential for SARS-CoV-2 entry, is a key antiviral target. Here, we computationally profiled the TMPRSS2-binding affinities of 15 antiviral compounds. Molecular dynamics (MD) simulations for the docked complexes revealed that three compounds exited the substrate-binding cavity (SBC), suggesting noncompetitive inhibition. Of the remaining compounds, five charged ones exhibited reduced binding stability due to competing electrostatic interactions and increased solvent exposure, while seven neutral compounds showed stronger binding affinity driven by van der Waals (vdW) interactions compensating for unfavorable electrostatic effects (including electrostatic interactions and desolvation penalties). Positive and negative hotspot residues were identified as uncharged and charged, respectively, both lining the SBC. Despite forming diverse interactions with compounds, the burial of positive hotspots led to strong vdW interactions that overcompensated for unfavorable electrostatic effects, whereas negative hotspots incurred high desolvation penalties, negating any favorable contributions. Charged residues at the SBC’s outer rim can reduce binding affinity significantly when forming hydrogen bonds or salt bridges. These findings underscore the importance of enhancing vdW interactions with uncharged residues and minimizing the unfavorable electrostatic effects of charged residues, providing valuable insights for designing effective TMPRSS2 inhibitors.

Colletotrichum spp. are widespread fungal pathogens that cause anthracnose in numerous economically important crops and, exhibiting extensive taxonomic, host plant, and lifestyle diversity. Here, we analyzed the genome sequences of 150 strains representing 97 species across 15 species complexes and four singletons, including and integrating both newly assembled and publicly available genomes. Phylogenomic investigation clarified the taxonomy of Colletotrichum and resolved misidentifications. We identified variations in genome architecture contributed by phylogenetic lineages, host types, and lifestyles, with transposable element proliferation playing significant roles. Interestingly, codon usage bias followed phylogenetic patterns, with species complexes forming distinct clusters and exhibiting a significant bistable co-evolutionary relationship with tRNA genes. Functional gene repertoires displayed coordinated shifts, with higher abundance in broad host-range species complexes and in strains associated with woody or dicotyledonous hosts. Although most functional categories retained strong phylogenetic signals, co-occurrence analysis of weak-signal categories identified modules related to host cell wall disruption, fungal cell wall remodeling, and virulence that were significantly associated with ecological differentiation. Evolutionary trajectories and gene family dynamics further revealed divergent ecological strategies, with oxidative versus rapid-response detoxification in woody- and herbaceous-associated lineages, respectively. The diversifications were accompanied by woody-specific expansion of GH39 and alkaline proteases and progressive differentiation of pectin-degrading capacity, including contraction in woody lineages and divergence between dicot- and monocot-associated herbaceous lineages. The C. gloeosporioides species complex emerged with a comprehensive expansion of detoxification and cell wall-degrading capacities, likely contributing to its broad host range. In contrast, endophytic lineages exhibited convergent gene family contraction in adhesion and cell wall remodeling. Together, this study revealed concordance of codon usage and functional gene abundance with phylogeny, along with diverse host- and lifestyle-associated adaptive strategies in this important group of plant pathogens.