{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,3]],"date-time":"2026-08-03T18:48:13Z","timestamp":1785782893513,"version":"3.56.0"},"reference-count":58,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2025,4,25]],"date-time":"2025-04-25T00:00:00Z","timestamp":1745539200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Polish National Science Centre","award":["UMO-2020\/37\/B\/NZ6\/01476"],"award-info":[{"award-number":["UMO-2020\/37\/B\/NZ6\/01476"]}]},{"name":"Polish National Science Centre","award":["KP-06-N72\/3\/2023, AIDA"],"award-info":[{"award-number":["KP-06-N72\/3\/2023, AIDA"]}]},{"name":"Polish National Science Centre","award":["KP-06-COST-20\/2023, INSISTT"],"award-info":[{"award-number":["KP-06-COST-20\/2023, INSISTT"]}]},{"name":"Bulgarian National Science Fund","award":["UMO-2020\/37\/B\/NZ6\/01476"],"award-info":[{"award-number":["UMO-2020\/37\/B\/NZ6\/01476"]}]},{"name":"Bulgarian National Science Fund","award":["KP-06-N72\/3\/2023, AIDA"],"award-info":[{"award-number":["KP-06-N72\/3\/2023, AIDA"]}]},{"name":"Bulgarian National Science Fund","award":["KP-06-COST-20\/2023, INSISTT"],"award-info":[{"award-number":["KP-06-COST-20\/2023, INSISTT"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>The SARS-CoV-2 spike receptor-binding motif is crucial for viral entry via interaction with the human ACE2 receptor. Mutations N501Y and E484K, found in several variants of concern, impact viral transmissibility and immune escape, but experimental data on their binding effects remain inconsistent. Using isothermal titration calorimetry (ITC) and molecular dynamics (MD) simulations, we analyzed the thermodynamic and structural effects of these mutations. ITC confirmed that N501Y increases ACE2 affinity by 2.2-fold, while E484K enhances binding by 5.8-fold. The Beta\/Gamma variant (carrying both mutations) showed the strongest affinity, with a 15-fold increase. E484K was enthalpy-driven, while N501Y introduced entropy-driven effects, suggesting hydrophobic interactions and conformational changes. MD simulations revealed distinct binding poses, with Beta\/Gamma peptides interacting with a secondary ACE2 site. A strong correlation was found between entropy contributions and hydrophobic contacts. Additionally, a convolutional neural network was used to estimate the free binding energy of these complexes. Our findings confirm that N501Y and E484K enhance ACE2 binding, with the greatest effect when combined, providing insights into SARS-CoV-2 variant evolution and potential therapeutic strategies.<\/jats:p>","DOI":"10.3390\/ijms26094064","type":"journal-article","created":{"date-parts":[[2025,4,25]],"date-time":"2025-04-25T04:08:24Z","timestamp":1745554104000},"page":"4064","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Impact of SARS-CoV-2 RBM Mutations N501Y and E484K on ACE2 Binding: A Combined Computational and Experimental Study"],"prefix":"10.3390","volume":"26","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-8790-3541","authenticated-orcid":false,"given":"Agnieszka","family":"Rombel-Bryzek","sequence":"first","affiliation":[{"name":"Institute of Medical Sciences, University of Opole, 45-052 Opole, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0420-9480","authenticated-orcid":false,"given":"Peicho","family":"Petkov","sequence":"additional","affiliation":[{"name":"Faculty of Physics, Sofia University \u201cSt. Kliment Ohridski\u201d, 1164 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2008-710X","authenticated-orcid":false,"given":"Elena","family":"Lilkova","sequence":"additional","affiliation":[{"name":"Institute of Information and Communication Technologies, Bulgarian Academy of Sciences, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2372-2319","authenticated-orcid":false,"given":"Nevena","family":"Ilieva","sequence":"additional","affiliation":[{"name":"Institute of Information and Communication Technologies, Bulgarian Academy of Sciences, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8511-6883","authenticated-orcid":false,"given":"Leandar","family":"Litov","sequence":"additional","affiliation":[{"name":"Faculty of Physics, Sofia University \u201cSt. Kliment Ohridski\u201d, 1164 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6602-2742","authenticated-orcid":false,"given":"Mariusz","family":"Kubus","sequence":"additional","affiliation":[{"name":"Centre of Education and Mathematics Applications, Opole University of Technology, 45-758 Opole, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6346-7543","authenticated-orcid":false,"given":"Danuta","family":"Witkowska","sequence":"additional","affiliation":[{"name":"Institute of Health Sciences, University of Opole, 45-060 Opole, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,4,25]]},"reference":[{"key":"ref_1","unstructured":"WHO (2024, January 15). Tracking SARS-CoV-2 Variants. Available online: https:\/\/www.who.int\/en\/activities\/tracking-SARS-CoV-2-variants\/."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"403","DOI":"10.1093\/procel\/pwae007","article-title":"Mutations in the SARS-CoV-2 Spike Receptor Binding Domain and Their Delicate Balance between ACE2 Affinity and Antibody Evasion","volume":"15","author":"Xue","year":"2024","journal-title":"Protein Cell"},{"key":"ref_3","unstructured":"WHO COVID-19 Dashboard (2024, January 15). COVID-19 Deaths. Available online: https:\/\/covid19.who.int."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"33","DOI":"10.5604\/01.3001.0054.4076","article-title":"Differences in treatment and post-COVID symptoms in elderly and young patients: A cross-sectional study","volume":"17","author":"Paskaleva","year":"2024","journal-title":"Med. Sci. Pulse"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"10","DOI":"10.5604\/01.3001.0053.8890","article-title":"Resilience index and mental stress of nursing staff working in intensive care units during the SARS-CoV-2 coronavirus pandemic","volume":"17","author":"Januszek","year":"2023","journal-title":"Med. Sci. Pulse"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"396","DOI":"10.1038\/s41392-021-00809-8","article-title":"The glycosylation in SARS-CoV-2 and its receptor ACE2","volume":"6","author":"Gong","year":"2021","journal-title":"Signal Transduct. Target. Ther."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1735","DOI":"10.1016\/j.cell.2020.11.032","article-title":"Structure, Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein","volume":"183","author":"Walls","year":"2020","journal-title":"Cell"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1603","DOI":"10.1126\/science.abc4730","article-title":"Adaptation of SARS-CoV-2 in BALB\/c mice for testing vaccine efficacy","volume":"369","author":"Gu","year":"2020","journal-title":"Science"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"215","DOI":"10.1038\/s41586-020-2180-5","article-title":"Structure of the SARS-CoV-2 spike receptor-binding domain bound to the ACE2 receptor","volume":"581","author":"Lan","year":"2020","journal-title":"Nature"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Witkowska, D. (2020). Mass Spectrometry and Structural Biology Techniques in the Studies on the Coronavirus-Receptor Interaction. Molecules, 25.","DOI":"10.3390\/molecules25184133"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"294","DOI":"10.1038\/s41586-021-04245-0","article-title":"The N501Y spike substitution enhances SARS-CoV-2 infection and transmission","volume":"602","author":"Liu","year":"2022","journal-title":"Nature"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1587","DOI":"10.4014\/jmb.2308.08020","article-title":"Distinctive Combinations of RBD Mutations Contribute to Antibody Evasion in the Case of the SARS-CoV-2 Beta Variant","volume":"33","author":"Kim","year":"2023","journal-title":"J. Microbiol. Biotechnol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"620","DOI":"10.1038\/s41591-021-01270-4","article-title":"Neutralization of SARS-CoV-2 spike 69\/70 deletion, E484K and N501Y variants by BNT162b2 vaccine-elicited sera","volume":"27","author":"Xie","year":"2021","journal-title":"Nat. Med."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Laffeber, C., De Koning, K., Kanaar, R., and Lebbink, J. (2021). Experimental Evidence for Enhanced Receptor Binding by Rapidly Spreading SARS-CoV-2 Variants. J. Mol. Biol., 433.","DOI":"10.1101\/2021.02.22.432357"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"e70658","DOI":"10.7554\/eLife.70658","article-title":"Effects of common mutations in the SARS-CoV-2 Spike RBD and its ligand, the human ACE2 receptor on binding affinity and kinetics","volume":"10","author":"Barton","year":"2021","journal-title":"eLife"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1022","DOI":"10.1016\/j.csbj.2023.01.024","article-title":"SARS-CoV-2 and its impact on the cardiovascular and digestive systems\u2014The interplay between new virus variants and human cells","volume":"21","author":"Szpulak","year":"2023","journal-title":"Comput. Struct. Biotechnol. J."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"e69091","DOI":"10.7554\/eLife.69091","article-title":"N501Y mutation of spike protein in SARS-CoV-2 strengthens its binding to receptor ACE2","volume":"10","author":"Tian","year":"2021","journal-title":"eLife"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"720","DOI":"10.1038\/s41422-021-00496-8","article-title":"The basis of a more contagious 501Y.V1 variant of SARS-CoV-2","volume":"31","author":"Liu","year":"2021","journal-title":"Cell Res."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"859","DOI":"10.1007\/s12257-021-0327-3","article-title":"SARS-CoV-2 Variants: Mutations and Effective Changes","volume":"26","author":"Park","year":"2021","journal-title":"Biotechnol. Bioprocess Eng."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"e283","DOI":"10.1016\/S2666-5247(21)00068-9","article-title":"SARS-CoV-2 spike E484K mutation reduces antibody neutralisation","volume":"2","author":"Jangraa","year":"2021","journal-title":"Lancet Microbe"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"1605","DOI":"10.1002\/jcc.20084","article-title":"UCSF Chimera\u2014A visualization system for exploratory research and analysis","volume":"25","author":"Pettersen","year":"2004","journal-title":"J. Comput. Chem."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Isobe, A., Arai, Y., Kuroda, D., Okumura, N., Ono, T., Ushiba, S., Nakakita, S.i., Daidoji, T., Suzuki, Y., and Nakaya, T. (2022). ACE2 N-glycosylation modulates interactions with SARS-CoV-2 spike protein in a site-specific manner. Commun. Biol., 5.","DOI":"10.1038\/s42003-022-04170-6"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"58","DOI":"10.1016\/j.jpha.2021.09.006","article-title":"Assessment of the binding interactions of SARS-CoV-2 spike glycoprotein variants","volume":"12","author":"Raghu","year":"2022","journal-title":"Ournal Pharm. Anal."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"111","DOI":"10.1111\/all.15065","article-title":"In vitro data suggest that Indian delta variant B.1.617 of SARS-CoV-2 escapes neutralization by both receptor affinity and immune evasion","volume":"77","author":"Augusto","year":"2022","journal-title":"Allergy"},{"key":"ref_25","unstructured":"Ramanathan, M., Ferguson, I., Miao, W., and Khavari, P. (2024, January 15). SARS-CoV-2 B.1.1.7 and B.1.351 Spike Variants Bind Human ACE2 with Increased Affinity. Available online: https:\/\/www.biorxiv.org\/content\/10.1101\/2021.02.22.432359v1."},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Pr\u00e9vost, J., Richard, J., Gasser, R., Ding, S., Fage, C., Anand, S.P., Adam, D., Gupta Vergara, N., Tauzin, A., and Benlarbi, M. (2021). Impact of temperature on the affinity of SARS-CoV-2 Spike glycoprotein for host ACE2. J. Biol. Chem., 297.","DOI":"10.1101\/2021.07.09.451812"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"2201","DOI":"10.1016\/j.cell.2021.02.033","article-title":"Reduced neutralization of SARS-CoV-2 B.1.1.7 variant by convalescent and vaccine sera","volume":"184","author":"Supasa","year":"2021","journal-title":"Cell"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"e70002","DOI":"10.7554\/eLife.70002","article-title":"The alpha\/B.1.1.7 SARS-CoV-2 variant exhibits significantly higher affinity for ACE-2 and requires lower inoculation doses to cause disease in K18-hACE2 mice","volume":"10","author":"Johnsen","year":"2021","journal-title":"eLife"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"143","DOI":"10.1111\/all.15002","article-title":"Molecular definition of severe acute respiratory syndrome coronavirus 2 receptor-binding domain mutations: Receptor affinity versus neutralization of receptor interaction","volume":"77","author":"Vogel","year":"2022","journal-title":"Allergy"},{"key":"ref_30","unstructured":"Tripathi, T., and Dubey, V.K. (2022). Chapter 8-Experimental methods to study the kinetics of protein\u2013protein interactions. Advances in Protein Molecular and Structural Biology Methods, Academic Press."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"108424","DOI":"10.1016\/j.intimp.2021.108424","article-title":"E484K and N501Y SARS-CoV 2 spike mutants Increase ACE2 recognition but reduce affinity for neutralizing antibody","volume":"102","author":"Chakraborty","year":"2022","journal-title":"Int. Immunopharmacol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1007\/s13167-021-00267-w","article-title":"Volpicella, M.; De Grassi, A.; Pierri, C.L. Modeling SARS-CoV-2 spike\/ACE2 protein\u2013protein interactions for predicting the binding affinity of new spike variants for ACE2, and novel ACE2 structurally related human protein targets, for COVID-19 handling in the 3PM context","volume":"13","author":"Tragni","year":"2022","journal-title":"EPMA J."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"7045","DOI":"10.1002\/jcp.30367","article-title":"Higher infectivity of the SARS-CoV-2 new variants is associated with K417N\/T, E484K, and N501Y mutants: An insight from structural data","volume":"236","author":"Khan","year":"2021","journal-title":"J. Cell. Physiol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"12760","DOI":"10.1080\/07391102.2021.1975569","article-title":"Understanding the molecular interaction of SARS-CoV-2 spike mutants with ACE2 (angiotensin converting enzyme 2)","volume":"40","author":"Istifli","year":"2022","journal-title":"J. Biomol. Struct. Dyn."},{"key":"ref_35","unstructured":"Nelson, G., Buzko, O., Spilman, P., Niazi, K., Rabizadeh, S., and Soon-Shiong, P. (2024, January 15). Molecular Dynamic Simulation Reveals E484K Mutation Enhances Spike RBD-ACE2 Affinity and the Combination of E484K, K417N and N501Y Mutations (501Y.V2 Variant) Induces Conformational Change Greater than N501Y Mutant Alone, Potentially Resulting in an Escape Mutant. Available online: https:\/\/www.biorxiv.org\/content\/10.1101\/2021.01.13.426558v1."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1188","DOI":"10.1038\/s41564-021-00954-4","article-title":"SARS-CoV-2 variant prediction and antiviral drug design are enabled by RBD in vitro evolution","volume":"6","author":"Marciano","year":"2021","journal-title":"Nat. Microbiol."},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Zhu, X., Mannar, D., Srivastava, S.S., Berezuk, A.M., Demers, J.P., Saville, J.W., Leopold, K., Li, W., Dimitrov, D.S., and Tuttle, K.S. (2021). Cryo-electron microscopy structures of the N501Y SARS-CoV-2 spike protein in complex with ACE2 and 2 potent neutralizing antibodies. PLoS Biol., 19.","DOI":"10.1371\/journal.pbio.3001237"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"136","DOI":"10.1038\/s41586-021-03412-7","article-title":"Sensitivity of SARS-CoV-2 B.1.1.7 to mRNA vaccine-elicited antibodies","volume":"593","author":"Collier","year":"2021","journal-title":"Nature"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Tanaka, S., Nelson, G., Olson, C.A., Buzko, O., Higashide, W., Shin, A., Gonzalez, M., Taft, J., Patel, R., and Buta, S. (2021). An ACE2 Triple Decoy that neutralizes SARS-CoV-2 shows enhanced affinity for virus variants. Sci. Rep., 11.","DOI":"10.1038\/s41598-021-91809-9"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"254","DOI":"10.1016\/j.bbapap.2017.10.005","article-title":"Analysis of Substrate Binding in Individual Active Sites of Bifunctional Human ATIC","volume":"1866","author":"Witkowska","year":"2018","journal-title":"Biochim. Et Biophys. Acta BBA-Proteins Proteom."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"3","DOI":"10.1016\/bs.mie.2015.08.021","article-title":"Isothermal Titration Calorimetry Measurements of Metal Ions Binding to Proteins","volume":"Volume 567","author":"Quinn","year":"2016","journal-title":"Methods in Enzymology"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"17996","DOI":"10.1074\/jbc.M311191200","article-title":"ACE2 X-Ray Structures Reveal a Large Hinge-bending Motion Important for Inhibitor Binding and Catalysis","volume":"279","author":"Towler","year":"2004","journal-title":"J. Biol. Chem."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"586","DOI":"10.1016\/j.chom.2020.08.004","article-title":"Virus-Receptor Interactions of Glycosylated SARS-CoV-2 Spike and Human ACE2 Receptor","volume":"28","author":"Zhao","year":"2020","journal-title":"Cell Host Microbe"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"320","DOI":"10.1093\/glycob\/cwz003","article-title":"CHARMM-GUI Glycan Modeler for modeling and simulation of carbohydrates and glycoconjugates","volume":"29","author":"Park","year":"2018","journal-title":"Glycobiology"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"1859","DOI":"10.1002\/jcc.20945","article-title":"CHARMM-GUI: A web-based graphical user interface for CHARMM","volume":"29","author":"Jo","year":"2008","journal-title":"J. Comput. Chem."},{"key":"ref_46","unstructured":"Schrodinger, LLC (2024, January 15). The PyMOL Molecular Graphics System. Version 1.7.4. Available online: https:\/\/www.pymol.org."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"19","DOI":"10.1016\/j.softx.2015.06.001","article-title":"GROMACS: High performance molecular simulations through multi-level parallelism from laptops to supercomputers","volume":"1\u20132","author":"Abraham","year":"2015","journal-title":"SoftwareX"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1038\/nmeth.4067","article-title":"CHARMM36m: An improved force field for folded and intrinsically disordered proteins","volume":"14","author":"Huang","year":"2016","journal-title":"Nat. Methods"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"3162","DOI":"10.1021\/ct200328p","article-title":"CHARMM additive all-atom force field for carbohydrate derivatives and its utility in polysaccharide and carbohydrate\u2013protein modeling","volume":"7","author":"Guvench","year":"2011","journal-title":"J. Chem. Theory Comput."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"014101","DOI":"10.1063\/1.2408420","article-title":"Canonical sampling through velocity rescaling","volume":"126","author":"Bussi","year":"2007","journal-title":"J. Chem. Phys."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"1196","DOI":"10.1103\/PhysRevLett.45.1196","article-title":"Crystal structure and pair potentials: A molecular-dynamics study","volume":"45","author":"Parrinello","year":"1980","journal-title":"Phys. Rev. Lett."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"8577","DOI":"10.1063\/1.470117","article-title":"A smooth particle mesh Ewald method","volume":"103","author":"Essmann","year":"1995","journal-title":"J. Chem. Phys."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"116","DOI":"10.1021\/ct700200b","article-title":"P-LINCS: A parallel linear constraint solver for molecular simulation","volume":"4","author":"Hess","year":"2008","journal-title":"J. Chem. Theory Comput."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"148","DOI":"10.1016\/0021-9991(74)90010-2","article-title":"Quiet high-resolution computer models of a plasma","volume":"14","author":"Hockney","year":"1974","journal-title":"J. Comput. Phys."},{"key":"ref_55","unstructured":"(2024, January 15). GROMACS Development Team. GROMACS Documentation Release 2021.1. Available online: https:\/\/zenodo.org\/records\/4561625."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"33","DOI":"10.1016\/0263-7855(96)00018-5","article-title":"VMD\u2013Visual Molecular Dynamics","volume":"14","author":"Humphrey","year":"1996","journal-title":"J. Mol. Graph."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"2778","DOI":"10.1021\/ci200227u","article-title":"LigPlot+: Multiple ligand-protein interaction diagrams for drug discovery","volume":"51","author":"Laskowski","year":"2011","journal-title":"J. Chem. Inf. Model."},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"2787","DOI":"10.1093\/bioinformatics\/btx312","article-title":"PROXiMATE: A database of mutant protein\u2013protein complex thermodynamics and kinetics","volume":"33","author":"Yugandhar","year":"2017","journal-title":"Bioinformatics"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/9\/4064\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T17:21:15Z","timestamp":1760030475000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/9\/4064"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,4,25]]},"references-count":58,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2025,5]]}},"alternative-id":["ijms26094064"],"URL":"https:\/\/doi.org\/10.3390\/ijms26094064","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,4,25]]}}}