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Here, we investigate how protein backbone topology influences phase behavior using ubiquitin C-terminal hydrolase L1 (UCH-L1), a highly neuron-enriched deubiquitinase in the ubiquitin\u2013proteasome system harboring a rare, evolutionarily conserved knotted backbone topology, and its Parkinson\u2019s disease\u2013associated I93M mutant. Through multiscale molecular dynamics (MD) simulations of single-chain and multichain systems, we show that knot integrity acts as a conformational constraint that limits access to expanded states, and suppresses LLPS propensity. Destabilization of the native knotted ensemble in I93M reshapes the conformational ensemble, enhancing intermolecular contacts, strengthening hydrophobic interaction, and reducing solvation penalties, thereby stabilizing protein-rich phases. Within condensates, these changes lead to persistent interchain contacts, increased topological entanglement, and slower relaxation dynamics, indicative of a transition toward viscoelastic assemblies, whereas intact topology maintains dynamic, liquid-like behavior. Our results identify topological integrity as a key physical determinant of protein phase behavior and establish a mechanistic link between topological stability and condensate material properties, with implications for aggregation-associated neurodegeneration.<\/jats:p>","DOI":"10.64898\/2026.06.12.731949","type":"posted-content","created":{"date-parts":[[2026,6,14]],"date-time":"2026-06-14T05:40:17Z","timestamp":1781415617000},"source":"Crossref","is-referenced-by-count":1,"title":["Tightly Knotted Enzymes Inhibit Protein-Protein Aggregation"],"prefix":"10.64898","author":[{"given":"Susmita","family":"Sarkar","sequence":"first","affiliation":[{"name":"Institute for Computational Molecular Science, Temple University, Philadelphia, Pennsylvania 19122, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hemanth Mandya","family":"Nagaiah","sequence":"additional","affiliation":[{"name":"Institute for Computational Molecular Science, Temple University, Philadelphia, Pennsylvania 19122, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Michael L.","family":"Klein","sequence":"additional","affiliation":[{"name":"Institute for Computational Molecular Science, Temple University, Philadelphia, Pennsylvania 19122, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Vincenzo","family":"Carnevale","sequence":"additional","affiliation":[{"name":"Institute for Computational Molecular Science, Temple University, Philadelphia, Pennsylvania 19122, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026061607351474000_2026.06.12.731949v1.1","doi-asserted-by":"publisher","DOI":"10.1038\/nm1066"},{"key":"2026061607351474000_2026.06.12.731949v1.2","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-biochem-061516-045115"},{"key":"2026061607351474000_2026.06.12.731949v1.3","doi-asserted-by":"publisher","DOI":"10.1038\/nrm3810"},{"issue":"3","key":"2026061607351474000_2026.06.12.731949v1.4","doi-asserted-by":"crossref","first-page":"e147","DOI":"10.1038\/emm.2014.117","article-title":"Degradation of misfolded proteins in neurodegenerative diseases: therapeutic targets and strategies","volume":"47","year":"2015","journal-title":"Experimental & molecular medicine"},{"key":"2026061607351474000_2026.06.12.731949v1.5","doi-asserted-by":"publisher","DOI":"10.1038\/nrm2731"},{"issue":"1","key":"2026061607351474000_2026.06.12.731949v1.6","first-page":"2023","article-title":"Non-functional ubiquitin c-terminal hydrolase l1 drives podocyte injury through impairing proteasomes in autoimmune glomerulonephritis","volume":"14","year":"2114","journal-title":"Nature communications"},{"issue":"1","key":"2026061607351474000_2026.06.12.731949v1.7","first-page":"2022","article-title":"Structural basis for specific inhibition of the deubiquitinase uchl1","volume":"13","year":"5950","journal-title":"Nature Communications"},{"issue":"1","key":"2026061607351474000_2026.06.12.731949v1.8","first-page":"2015","article-title":"Uchl1 provides diagnostic and antimetastatic strategies due to its deubiquitinating effect on hif-1\u03b1","volume":"6","year":"6153","journal-title":"Nature communications"},{"key":"2026061607351474000_2026.06.12.731949v1.9","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.0806474106"},{"key":"2026061607351474000_2026.06.12.731949v1.10","doi-asserted-by":"publisher","DOI":"10.1126\/science.1141448"},{"key":"2026061607351474000_2026.06.12.731949v1.11","doi-asserted-by":"publisher","DOI":"10.1038\/nm.4001"},{"key":"2026061607351474000_2026.06.12.731949v1.12","doi-asserted-by":"publisher","DOI":"10.1126\/science.2530630"},{"key":"2026061607351474000_2026.06.12.731949v1.13","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1222732110"},{"key":"2026061607351474000_2026.06.12.731949v1.14","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.0510403103"},{"key":"2026061607351474000_2026.06.12.731949v1.15","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.0911516107"},{"key":"2026061607351474000_2026.06.12.731949v1.16","doi-asserted-by":"crossref","unstructured":"Daewon Lee , Eunju Yoon , Su Jin Ham , Kunwoo Lee , Hansaem Jang , Daihn Woo , Da Hyun Lee , Sehyeon Kim , Sekyu Choi , and Jongkyeong Chung . 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