{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,7,30]],"date-time":"2025-07-30T10:06:37Z","timestamp":1753869997207,"version":"3.41.2"},"reference-count":39,"publisher":"Wiley","issue":"2","license":[{"start":{"date-parts":[[2025,1,25]],"date-time":"2025-01-25T00:00:00Z","timestamp":1737763200000},"content-version":"am","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"},{"start":{"date-parts":[[2025,1,25]],"date-time":"2025-01-25T00:00:00Z","timestamp":1737763200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000001","name":"National Science Foundation","doi-asserted-by":"publisher","award":["1129152"],"award-info":[{"award-number":["1129152"]}],"id":[{"id":"10.13039\/100000001","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Protein Science"],"published-print":{"date-parts":[[2025,2]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Hydrogen exchange mass spectrometry (HXMS) is a powerful tool to understand protein folding pathways and energetics. However, HXMS experiments to date have used exchange conditions termed EX1 or EX2 which limit the information that can be gained compared to the more general EXX exchange regime. If EXX behavior could be understood and analyzed, a single HXMS timecourse on an intact protein could fully map its folding landscape without requiring denaturation. To address this challenge, we developed a numerical simulation method called NumSimEX that models EXX exchange for arbitrarily complex folding pathways. NumSimEx fits protein folding dynamics to experimental HXMS data by iteratively comparing the simulated and experimental timecourses, allowing for determination of both kinetic and thermodynamic protein folding parameters. After analytically verifying NumSimEX's accuracy, we demonstrated its power on HXMS data from beta\u20102 microglobulin (\u03b22M), a protein involved in dialysis\u2010related amyloidosis. In particular, using NumSimEX, we identified three\u2010state kinetics that near\u2010perfectly matched experimental observation. This proof\u2010of\u2010principle application of NumSimEX sets the stage for harnessing HXMS to expand our understanding of proteins currently excluded from traditional protein folding methods. NumSimEX is freely available at <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" xlink:href=\"https:\/\/github.com\/JaswalLab\/NumSimEX_Public\">https:\/\/github.com\/JaswalLab\/NumSimEX_Public<\/jats:ext-link>.<\/jats:p>","DOI":"10.1002\/pro.70045","type":"journal-article","created":{"date-parts":[[2025,1,25]],"date-time":"2025-01-25T12:49:58Z","timestamp":1737809398000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["<scp>NumSimEX<\/scp>: A method using <scp>EXX<\/scp> hydrogen exchange mass spectrometry to map the energetics of protein folding landscapes"],"prefix":"10.1002","volume":"34","author":[{"given":"Jasper A. G.","family":"Flint","sequence":"first","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"University of Maryland School of Medicine  Baltimore Maryland USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0037-5999","authenticated-orcid":false,"given":"Jacob","family":"Witten","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"David H Koch Institute for Integrative Cancer Research Massachusetts Institute of Technology  Cambridge Massachusetts USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Isabella","family":"Han","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"Chicago Medical School Rosalind Franklin University of Medicine &amp; Science  North Chicago Illinois USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"John","family":"Strahan","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"Northwestern University  Evanston Illinois USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jovan","family":"Damjanovic","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"Novo Nordisk A\/S  Lexington Massachusetts USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nevon","family":"Song","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"Montefiore Medical Center  Bronx New York USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tim","family":"Poterba","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"E9 Genomics  Cambridge Massachusetts USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alexis Jaramillo","family":"Cartagena","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"Memorial Sloan Kettering Cancer Center  New York New York USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Angelika","family":"Hirsch","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"},{"name":"Stanford University  Palo Alto California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tony","family":"Ni","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Julie L.","family":"Sohl","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Amy S.","family":"Wagaman","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9105-3842","authenticated-orcid":false,"given":"Sheila S.","family":"Jaswal","sequence":"additional","affiliation":[{"name":"Amherst College  Amherst Massachusetts USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2025,1,25]]},"reference":[{"key":"e_1_2_9_2_1","doi-asserted-by":"publisher","DOI":"10.1006\/jmbi.2000.3859"},{"key":"e_1_2_9_3_1","doi-asserted-by":"publisher","DOI":"10.1006\/jmbi.1998.2338"},{"key":"e_1_2_9_4_1","doi-asserted-by":"publisher","DOI":"10.1002\/prot.340170110"},{"key":"e_1_2_9_5_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.7618079"},{"key":"e_1_2_9_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbapap.2012.10.009"},{"key":"e_1_2_9_7_1","doi-asserted-by":"publisher","DOI":"10.1002\/prot.23039"},{"key":"e_1_2_9_8_1","doi-asserted-by":"publisher","DOI":"10.1038\/nsb0996-782"},{"key":"e_1_2_9_9_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.tibs.2004.08.008"},{"key":"e_1_2_9_10_1","doi-asserted-by":"publisher","DOI":"10.1002\/prot.340170111"},{"key":"e_1_2_9_11_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.semcdb.2003.12.008"},{"key":"e_1_2_9_12_1","doi-asserted-by":"publisher","DOI":"10.1038\/nature04105"},{"key":"e_1_2_9_13_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jasms.2006.06.006"},{"key":"e_1_2_9_14_1","doi-asserted-by":"publisher","DOI":"10.1021\/j100540a008"},{"key":"e_1_2_9_15_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi0509548"},{"key":"e_1_2_9_16_1","first-page":"385","article-title":"Exchange of deuterium and 18O between water and other substances. 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