{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T18:56:46Z","timestamp":1785437806718,"version":"3.56.0"},"posted":{"date-parts":[[2026,4,17]]},"group-title":"Biophysics","reference-count":90,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,4,17]],"date-time":"2026-04-17T00:00:00Z","timestamp":1776384000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"award":["5R01HL138579"],"award-info":[{"award-number":["5R01HL138579"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R35GM142868"],"award-info":[{"award-number":["R35GM142868"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["#25PRE1377151"],"award-info":[{"award-number":["#25PRE1377151"]}],"id":[{"id":"https:\/\/ror.org\/013kjyp64","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,4,17]]},"abstract":"<jats:label>2.<\/jats:label>\n                <jats:title>ABSTRACT<\/jats:title>\n                <jats:p>\n                  Calmodulin (CaM) is a highly conserved Ca\n                  <jats:sup>2+<\/jats:sup>\n                  sensor that regulates hundreds of cellular targets through Ca\n                  <jats:sup>2+<\/jats:sup>\n                  -dependent conformational dynamics. Despite its central role in Ca\n                  <jats:sup>2+<\/jats:sup>\n                  signaling and disease, its evolutionary conservation and structural flexibility have suggested that CaM is resistant to rational redesign. Here, using the cardiac Ca\n                  <jats:sup>2+<\/jats:sup>\n                  release channel Ryanodine receptor 2 (RyR2) as a model system, we tested whether incorporating conformational dynamics into computational protein design enables functional reengineering of CaM. We first applied a static structure\u2013based redesign to increase CaM\u2013RyR2 affinity. Although the resulting variant bound more tightly to both the RyR2 peptide and the intact channel in vitro, it distorted peptide geometry and worsened Ca\n                  <jats:sup>2+<\/jats:sup>\n                  leak in cardiomyocytes ex vivo. Guided by molecular dynamics simulations, we then developed a dynamic-structure redesign strategy that preserves conformational integrity while strengthening binding. The resulting CaM variant exhibited increased RyR2 affinity and reduced pathological Ca\n                  <jats:sup>2+<\/jats:sup>\n                  leak in a disease-relevant model. These findings show that improved binding affinity alone is insufficient to enhance physiological regulation and that successful CaM redesign requires preservation of conformational dynamics. More broadly, they demonstrate that integrating conformational dynamics into protein redesign can enable functionally predictive engineering of flexible regulatory protein\u2013protein interactions.\n                <\/jats:p>","DOI":"10.64898\/2026.04.14.717973","type":"posted-content","created":{"date-parts":[[2026,4,17]],"date-time":"2026-04-17T17:35:21Z","timestamp":1776447321000},"source":"Crossref","is-referenced-by-count":0,"title":["Dynamic-Structure Redesign of Calmodulin Reveals Mechanistic Constraints on Ryr2 Regulation"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9287-5303","authenticated-orcid":false,"given":"Vladimir","family":"Bogdanov","sequence":"first","affiliation":[{"name":"The Dorothy M. Davis Heart and Lung Research Institute, The Ohio State University Wexner Medical Center, Columbus, Ohio, 43210"},{"name":"Department of Physiology and Cell Biology, College of Medicine, The Ohio State University, Columbus, Ohio, 43210"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Svetlana","family":"Tikunova","sequence":"additional","affiliation":[{"name":"The Dorothy M. Davis Heart and Lung Research Institute, The Ohio State University Wexner Medical Center, Columbus, Ohio, 43210"},{"name":"Department of Physiology and Cell Biology, College of Medicine, The Ohio State University, Columbus, Ohio, 43210"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nicolas","family":"Fadell","sequence":"additional","affiliation":[{"name":"Department of Physiology and Cell Biology, College of Medicine, The Ohio State University, Columbus, Ohio, 43210"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robyn T.","family":"Rebbeck","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minnesota, Minneapolis, 55455"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Melanie L.","family":"Aprahamian","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, Ohio State University, Columbus, Ohio 43210, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Md Nure Alam","family":"Afsar","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Mississippi State University, Starkville, Mississippi, 39759"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Aleksei","family":"Chekodanov","sequence":"additional","affiliation":[{"name":"Independent Software Developer, Moscow, Russia, 119049"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Daniel J.","family":"Blackwell","sequence":"additional","affiliation":[{"name":"Vanderbilt Center for Arrhythmia Research and Therapeutics, Division of Clinical Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee, 37212"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4956-9735","authenticated-orcid":false,"given":"Bjorn C.","family":"Knollmann","sequence":"additional","affiliation":[{"name":"Vanderbilt Center for Arrhythmia Research and Therapeutics, Division of Clinical Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee, 37212"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5739-0992","authenticated-orcid":false,"given":"Razvan L.","family":"Cornea","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minnesota, Minneapolis, 55455"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7286-3022","authenticated-orcid":false,"given":"Pete M.","family":"Kekenes-Huskey","sequence":"additional","affiliation":[{"name":"Department of Cell and Molecular Physiology, Stritch School of Medicine, Loyola University Chicago, Maywood, Illinois, 60153"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Steffen","family":"Lindert","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, The University of California, Los Angeles, California, 90095"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christopher N.","family":"Johnson","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Mississippi State University, Starkville, Mississippi, 39759"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sandor","family":"Gy\u00f6rke","sequence":"additional","affiliation":[{"name":"The Dorothy M. Davis Heart and Lung Research Institute, The Ohio State University Wexner Medical Center, Columbus, Ohio, 43210"},{"name":"Department of Physiology and Cell Biology, College of Medicine, The Ohio State University, Columbus, Ohio, 43210"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jonathan P.","family":"Davis","sequence":"additional","affiliation":[{"name":"The Dorothy M. 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