{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,6]],"date-time":"2026-05-06T17:21:14Z","timestamp":1778088074549,"version":"3.51.4"},"reference-count":36,"publisher":"Wiley","issue":"22","license":[{"start":{"date-parts":[[2018,9,26]],"date-time":"2018-09-26T00:00:00Z","timestamp":1537920000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"DOI":"10.13039\/501100001502","name":"Department of Atomic Energy, Government of India","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100001502","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["febs.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["The FEBS Journal"],"published-print":{"date-parts":[[2018,11]]},"abstract":"<jats:sec><jats:label\/><jats:p>Human cathepsin K (CTSK) is a collagenolytic lysosomal cysteine protease that plays an important role in bone turnover. Mutation in <jats:italic>CTSK<\/jats:italic> gene is associated with loss of collagenolytic activity of CTSK leading to an autosomal recessive bone disorder called pycnodysostosis. Although a number of pycnodysostotic missense mutations have been reported, underlying mechanism of the disease is not clear. In this study, we investigated <jats:italic>in\u00a0vitro<\/jats:italic> six recombinant pycnodysostosis\u2010related mutants of human CTSK (G79E, I249T, G243E, G303E, G319C and Q187P). While all the mutants, like wild\u2010type, show similar high levels of expression in <jats:italic>Escherichia\u00a0coli<\/jats:italic>, four of them (G79E, G303E, G319C and Q187P) are inactive, unstable and spontaneously degrade during purification process. In contrast, proteolytic\/collagenolytic activity, zymogen activation kinetics and stability of G243E and I249T mutants are nominally affected. Crystal structure of I249T at 1.92\u00a0\u00c5 resolution shows that the mutation in R\u2010domain causes conformational changes of a surface loop in the L\u2010domain although the catalytic cleft remains unaltered. Molecular simulation, normal mode analysis and fluorescence lifetime measurement eliminated the possibility that the change in L\u2010domain surface loop orientation is a crystallization artefact. CD\u2010based thermal melting profile indicates that stability of I249T is significantly higher than wild\u2010type. Our studies first time reports that pycnodysostosis\u2010related mutations do not always lead to complete loss of general proteolytic activity or specific collagenolytic activity of CTSK. The first crystal structure of a pycnodysostotic mutant (I249T) provides critical information that may pave new avenues towards understanding the disease at molecular level.<\/jats:p><\/jats:sec><jats:sec><jats:title>Database<\/jats:title><jats:p>The atomic co\u2010ordinates and structure factors for I249T mutant of human CTSK (codes 5Z5O) have been deposited in the Protein Data Bank (<jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" xlink:href=\"http:\/\/wwpdb.org\/\">http:\/\/wwpdb.org\/<\/jats:ext-link>)<\/jats:p><\/jats:sec>","DOI":"10.1111\/febs.14655","type":"journal-article","created":{"date-parts":[[2018,9,10]],"date-time":"2018-09-10T17:58:49Z","timestamp":1536602329000},"page":"4265-4280","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Not all pycnodysostosis\u2010related mutants of human cathepsin K are inactive \u2013 crystal structure and biochemical studies of an active mutant I249T"],"prefix":"10.1111","volume":"285","author":[{"given":"Sumana","family":"Roy","sequence":"first","affiliation":[{"name":"Crystallography and Molecular Biology Division Saha Institute of Nuclear Physics Kolkata India"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sudeshna","family":"Das Chakraborty","sequence":"additional","affiliation":[{"name":"Chemical Sciences Division Saha Institute of Nuclear Physics Kolkata India"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sampa","family":"Biswas","sequence":"additional","affiliation":[{"name":"Crystallography and Molecular Biology Division Saha Institute of Nuclear Physics Kolkata India"},{"name":"Homi Bhaba National Institute Mumbai India"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2018,9,26]]},"reference":[{"key":"e_1_2_9_2_1","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkr987"},{"key":"e_1_2_9_3_1","doi-asserted-by":"publisher","DOI":"10.1515\/bchm3.1995.376.6.379"},{"key":"e_1_2_9_4_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.271.21.12511"},{"key":"e_1_2_9_5_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.271.21.12517"},{"key":"e_1_2_9_6_1","doi-asserted-by":"publisher","DOI":"10.1002\/ar.1092240220"},{"key":"e_1_2_9_7_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M204004200"},{"key":"e_1_2_9_8_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.addr.2004.12.013"},{"key":"e_1_2_9_9_1","doi-asserted-by":"publisher","DOI":"10.4155\/fmc.09.4"},{"key":"e_1_2_9_10_1","doi-asserted-by":"publisher","DOI":"10.1517\/13543780902832661"},{"key":"e_1_2_9_11_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.273.5279.1236"},{"key":"e_1_2_9_12_1","doi-asserted-by":"publisher","DOI":"10.1186\/1750-1172-6-20"},{"key":"e_1_2_9_13_1","doi-asserted-by":"publisher","DOI":"10.1186\/1750-1172-9-60"},{"key":"e_1_2_9_14_1","doi-asserted-by":"publisher","DOI":"10.1111\/febs.13980"},{"key":"e_1_2_9_15_1","doi-asserted-by":"publisher","DOI":"10.1007\/s12013-017-0838-x"},{"key":"e_1_2_9_16_1","doi-asserted-by":"publisher","DOI":"10.1038\/nrm2745"},{"key":"e_1_2_9_17_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.272.21.13955"},{"key":"e_1_2_9_18_1","doi-asserted-by":"publisher","DOI":"10.1038\/nsb0297-105"},{"key":"e_1_2_9_19_1","doi-asserted-by":"publisher","DOI":"10.1042\/BJ20100337"},{"key":"e_1_2_9_20_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi992251u"},{"key":"e_1_2_9_21_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M310349200"},{"key":"e_1_2_9_22_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2008.07.038"},{"key":"e_1_2_9_23_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1414126111"},{"key":"e_1_2_9_24_1","doi-asserted-by":"publisher","DOI":"10.1038\/ncomms4287"},{"key":"e_1_2_9_25_1","doi-asserted-by":"publisher","DOI":"10.1002\/humu.9490"},{"key":"e_1_2_9_26_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00223-015-9963-y"},{"key":"e_1_2_9_27_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.procbio.2012.06.029"},{"key":"e_1_2_9_28_1","doi-asserted-by":"publisher","DOI":"10.1007\/s10895-017-2053-y"},{"key":"e_1_2_9_29_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444909047337"},{"key":"e_1_2_9_30_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444909052925"},{"key":"e_1_2_9_31_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444904019158"},{"key":"e_1_2_9_32_1","first-page":"1281","article-title":"Substrate\u2010zymography: a still worthwhile method for gelatinases analysis in biological samples","volume":"54","author":"Ricci S","year":"2016","journal-title":"Clin Chem Lab Med"},{"key":"e_1_2_9_33_1","doi-asserted-by":"publisher","DOI":"10.1110\/ps.8.2.283"},{"key":"e_1_2_9_34_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi9822271"},{"key":"e_1_2_9_35_1","doi-asserted-by":"publisher","DOI":"10.1042\/BJ20140809"},{"key":"e_1_2_9_36_1","doi-asserted-by":"publisher","DOI":"10.1172\/JCI653"},{"key":"e_1_2_9_37_1","doi-asserted-by":"publisher","DOI":"10.1006\/bbrc.2001.4394"}],"container-title":["The FEBS Journal"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/api.wiley.com\/onlinelibrary\/tdm\/v1\/articles\/10.1111%2Ffebs.14655","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1111\/febs.14655","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/full-xml\/10.1111\/febs.14655","content-type":"application\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/febs.onlinelibrary.wiley.com\/doi\/pdf\/10.1111\/febs.14655","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,9,10]],"date-time":"2023-09-10T09:36:06Z","timestamp":1694338566000},"score":1,"resource":{"primary":{"URL":"https:\/\/febs.onlinelibrary.wiley.com\/doi\/10.1111\/febs.14655"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2018,9,26]]},"references-count":36,"journal-issue":{"issue":"22","published-print":{"date-parts":[[2018,11]]}},"alternative-id":["10.1111\/febs.14655"],"URL":"https:\/\/doi.org\/10.1111\/febs.14655","archive":["Portico"],"relation":{},"ISSN":["1742-464X","1742-4658"],"issn-type":[{"value":"1742-464X","type":"print"},{"value":"1742-4658","type":"electronic"}],"subject":[],"published":{"date-parts":[[2018,9,26]]},"assertion":[{"value":"2018-03-09","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2018-09-07","order":1,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2018-09-26","order":2,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}]}}