{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,20]],"date-time":"2026-10-20T02:58:41Z","timestamp":1792465121375,"version":"4.5.0"},"reference-count":31,"publisher":"National Academy of Sciences","issue":"44","content-domain":{"domain":["www.pnas.org"],"crossmark-restriction":true},"short-container-title":["Proc. Natl. Acad. Sci. U.S.A."],"published-print":{"date-parts":[[2012,10,30]]},"abstract":"<jats:p>We measure the stability and folding rate of a mutant of the enzyme phosphoglycerate kinase (PGK) inside bone tissue cells as a function of temperature from 38 to 48\u2009\u00b0C. To facilitate measurement in individual living cells, we developed a rapid laser temperature stepping method capable of measuring complete thermal melts and kinetic traces in about two min. We find that this method yields improved thermal melts compared to heating a sample chamber or microscope stage. By comparing results for six cells with in vitro data, we show that the protein is stabilized by about 6\u00a0kJ\/mole in the cytoplasm, but the temperature dependence of folding kinetics is similar to in vitro. The main difference is a slightly steeper temperature dependence of the folding rate in some cells that can be rationalized in terms of temperature-dependent crowding, local viscosity, or hydrophobicity. The observed rate coefficients can be fitted within measurement uncertainty by an effective two-state model, even though PGK folds by a multistate mechanism. We validate the effective two-state model with a three-state free energy landscape of PGK to illustrate that the effective fitting parameters can represent a more complex underlying free energy landscape.<\/jats:p>","DOI":"10.1073\/pnas.1201797109","type":"journal-article","created":{"date-parts":[[2012,6,5]],"date-time":"2012-06-05T10:05:21Z","timestamp":1338890721000},"page":"17863-17867","update-policy":"https:\/\/doi.org\/10.1073\/pnas.cm10313","source":"Crossref","is-referenced-by-count":133,"title":["Temperature dependence of protein folding kinetics in living cells"],"prefix":"10.1073","volume":"109","author":[{"given":"Minghao","family":"Guo","sequence":"first","affiliation":[{"name":"Departments of Physics,"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yangfan","family":"Xu","sequence":"additional","affiliation":[{"name":"Chemical and Biomolecular Engineering, and"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Martin","family":"Gruebele","sequence":"additional","affiliation":[{"name":"Departments of Physics,"},{"name":"Chemistry and Center for Biophysics and Computational Biology, University of Illinois, Urbana, IL 61801"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"341","published-online":{"date-parts":[[2012,6,4]]},"reference":[{"key":"e_1_3_3_1_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.282.5389.740"},{"key":"e_1_3_3_2_2","doi-asserted-by":"publisher","DOI":"10.1038\/nature01160"},{"key":"e_1_3_3_3_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.1208351"},{"key":"e_1_3_3_4_2","doi-asserted-by":"publisher","DOI":"10.1093\/emboj\/19.15.3870"},{"key":"e_1_3_3_5_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.bpj.2008.10.014"},{"key":"e_1_3_3_6_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1006760107"},{"key":"e_1_3_3_7_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.bpj.2010.08.066"},{"key":"e_1_3_3_8_2","doi-asserted-by":"publisher","DOI":"10.1002\/prot.340210302"},{"key":"e_1_3_3_9_2","doi-asserted-by":"publisher","DOI":"10.1021\/jz101729z"},{"key":"e_1_3_3_10_2","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.1435"},{"key":"e_1_3_3_11_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.bpj.2011.05.071"},{"key":"e_1_3_3_12_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.96.11.6031"},{"key":"e_1_3_3_13_2","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M601915200"},{"key":"e_1_3_3_14_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2003.08.011"},{"key":"e_1_3_3_15_2","volume-title":"Structure and Mechanism in Protein Science: A Guide to Enzyme Catalysis and Protein Folding","author":"Fersht AR","year":"1999","unstructured":"AR Fersht Structure and Mechanism in Protein Science: A Guide to Enzyme Catalysis and Protein Folding (W.H. 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