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The structure of HtrA2 with an intact catalytic triad was determined, revealing a conformational change in the active site loops, involving mainly the regulatory LD loop, which resulted in burial of the catalytic serine relative to the previously reported structure of the proteolytically inactive mutant. Mutations in the loops surrounding the active site that significantly restricted their mobility, reduced proteolytic activity both <jats:italic>in vitro<\/jats:italic> and in cells, suggesting that regulation of HtrA2 activity cannot be explained by a simple transition to an activated conformational state with enhanced active site accessibility. Manipulation of solvent viscosity highlighted an unusual bi-phasic behavior of the enzymatic activity, which together with MD calculations supports the importance of motion in the regulation of the activity of HtrA2. HtrA2 is an unusually thermostable enzyme (<jats:italic>T<\/jats:italic><jats:sub>M<\/jats:sub>=97.3\u2009\u00b0C), a trait often associated with structural rigidity, not dynamic motion. We suggest that this thermostability functions to provide a stable scaffold for the observed loop motions, allowing them a relatively free conformational search within a rather restricted volume.<\/jats:p>","DOI":"10.1038\/cddis.2017.487","type":"journal-article","created":{"date-parts":[[2017,10,12]],"date-time":"2017-10-12T12:43:02Z","timestamp":1507812182000},"page":"e3119-e3119","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":24,"title":["Molecular motion regulates the activity of the Mitochondrial Serine Protease HtrA2"],"prefix":"10.1038","volume":"8","author":[{"given":"Matthew","family":"Merski","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"C\u00e1tia","family":"Moreira","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rui MV","family":"Abreu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maria Jo\u00e3o","family":"Ramos","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pedro A","family":"Fernandes","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"L Miguel","family":"Martins","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pedro Jos\u00e9 Barbosa","family":"Pereira","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7698-1170","authenticated-orcid":false,"given":"Sandra","family":"Macedo-Ribeiro","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2017,10,12]]},"reference":[{"key":"BFcddis2017487_CR1","doi-asserted-by":"publisher","first-page":"152","DOI":"10.1038\/nrm3065","volume":"12","author":"T Clausen","year":"2011","unstructured":"Clausen T, Kaiser M, Huber R, Ehrmann M . 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