Antimicrobial peptides emerge as compounds that can alleviate the global health hazard of antimicrobial resistance, prompting a need for novel computational approaches to peptide generation. Here, we propose HydrAMP, a conditional variational autoencoder that learns lower-dimensional, continuous representation of peptides and captures their antimicrobial properties. The model disentangles the learnt representation of a peptide from its antimicrobial conditions and leverages parameter-controlled creativity. HydrAMP is the first model that is directly optimized for diverse tasks, including unconstrained and analogue generation and outperforms other approaches in these tasks. An additional preselection procedure based on ranking of generated peptides and molecular dynamics simulations increases experimental validation rate. Wet-lab experiments on five bacterial strains confirm high activity of nine peptides generated as analogues of clinically relevant prototypes, as well as six analogues of an inactive peptide. HydrAMP enables generation of diverse and potent peptides, making a step towards resolving the antimicrobial resistance crisis.

S-adenosylmethionine (SAM) is one of the most important enzyme substrates. It is vital for the function of various proteins, including large group of methyltransferases (MTs). Intriguingly, some bacterial and eukaryotic MTs, while catalysing the same reaction, possess significantly different topologies, with the former being a knotted one. Here, we conducted a comprehensive analysis of SAM conformational space and factors that affect its vastness. We investigated SAM in two forms: free in water (via NMR studies and explicit solvent simulations) and bound to proteins (based on all data available in the PDB and on all-atom molecular dynamics simulations in water). We identified structural descriptors—angles which show the major differences in SAM conformation between unknotted and knotted methyltransferases. Moreover, we report that this is caused mainly by a characteristic for knotted MTs compact binding site formed by the knot and the presence of adenine-binding loop. Additionally, we elucidate conformational restrictions imposed on SAM molecules by other protein groups in comparison to conformational space in water. The topology of a folded polypeptide chain has great impact on the resulting protein function and its interaction with ligands. Interestingly, topological constraints appear to affect binding of one of the most ubiquitous substrates in the cell, S-adenosylmethionine (SAM), to its target proteins. Here, we demonstrate how binding sites of specific proteins restrict SAM conformational freedom in comparison to its unbound state, with a special interest in proteins with non-trivial topology, including an exciting group of knotted methyltransferases. Using a vast array of computational methods combined with NMR experiments, we identify key structural features of knotted methyltransferases that impose unorthodox SAM conformations. We compare them with the characteristics of standard, unknotted SAM binding proteins. These results are significant for understanding differences between analogous, yet topologically different enzymes, as well as for future rational drug design.

Constantly advancing computer simulations of biomolecules provide huge amounts of data that are difficult to interpret. In particular, obtaining insights into functional aspects of macromolecular dynamics, often related to cascades of transient events, calls for methodologies that depart from the well-grounded framework of equilibrium statistical physics. One of the approaches toward the analysis of complex temporal data which has found applications in the fields of neuroscience and econometrics is Granger causality analysis. It allows determining which components of multidimensional time series are most influential for the evolution of the entire system, thus providing insights into causal relations within the dynamic structure of interest. In this work, we apply Granger analysis to a long molecular dynamics trajectory depicting repetitive folding and unfolding of a mini β-hairpin protein, CLN025. We find objective, quantitative evidence indicating that rearrangements within the hairpin turn region are determinant for protein folding and unfolding. On the contrary, interactions between hairpin arms score low on the causality scale. Taken together, these findings clearly favor the concept of zipperlike folding, which is one of two postulated β-hairpin folding mechanisms. More importantly, the results demonstrate the possibility of a conclusive application of Granger causality analysis to a biomolecular system.

Significance: Cooperativity in substrate binding to a protein often involves some kind of interaction between distant binding sites mediated by the protein structure. Experimental studies indicate that the cooperative effect in protein kinase A relies on a shift in conformational equilibrium of the catalytic subunit, but its exact molecular basis remains unclear. Our molecular dynamics simulations provide atomistic insight into conformational states visited by the enzyme and its complexes and suggest a plausible mechanistic model contributing to cooperative substrate binding. Intriguingly, an important role seems to be played by water molecules that occupy two conserved hydration sites within the catalytic subunit.

Cholesterol-rich lipid rafts are found to facilitate membrane fusion, central to processes like viral entry, fertilization, and neurotransmitter release. While the fusion process involves local, transient membrane dehydration, the impact of reduced hydration on cholesterol's structural organization in biological membranes remains unclear. Here, we employ confocal fluorescence microscopy and atomistic molecular dynamics simulations to investigate cholesterol behavior in phase-separated lipid bilayers under controlled hydration. We unveiled that dehydration prompts cholesterol release from raft-like domains into the surrounding fluid phase. Unsaturated phospholipids undergo more significant dehydration-induced structural changes and lose more hydrogen bonds with water than sphingomyelin. The results suggest that cholesterol redistribution is driven by the equalization of biophysical properties between phases and the need to satisfy lipid hydrogen bonds. This underscores the role of cholesterol-phospholipid-water interplay in governing cholesterol affinity for a specific lipid type, providing a new perspective on the regulatory role of cell membrane heterogeneity during membrane fusion.