000010913 001__ 10913 000010913 005__ 20251007025610.0 000010913 02470 $$ahttps://doi.org/10.1126/sciadv.abj8633$$2doi 000010913 037__ $$aTEXTUAL 000010913 037__ $$bArticle 000010913 041__ $$aeng 000010913 245__ $$aCln5 represents a new type of cysteine-based S-depalmitoylase linked to neurodegeneration 000010913 269__ $$a2022-04-15 000010913 336__ $$aArticle 000010913 520__ $$aGenetic CLN5 variants are associated with childhood neurodegeneration and Alzheimer’s disease; however, the molecular function of ceroid lipofuscinosis neuronal protein 5 (Cln5) is unknown. We solved the Cln5 crystal structure and identified a region homologous to the catalytic domain of members of the N1pC/P60 superfamily of papain-like enzymes. However, we observed no protease activity for Cln5; and instead, we discovered that Cln5 and structurally related PPPDE1 and PPPDE2 have efficient cysteine palmitoyl thioesterase (S-depalmitoylation) activity using fluorescent substrates. Mutational analysis revealed that the predicted catalytic residues histidine-166 and cysteine-280 are critical for Cln5 thioesterase activity, uncovering a new cysteine-based catalytic mechanism for S-depalmitoylation enzymes. Last, we found that Cln5-deficient neuronal progenitor cells showed reduced thioesterase activity, confirming live cell function of Cln5 in setting S-depalmitoylation levels. Our results provide new insight into the function of Cln5, emphasize the importance of S-depalmitoylation in neuronal homeostasis, and disclose a new, unexpected enzymatic function for the N1pC/P60 superfamily of proteins. 000010913 536__ $$oDeutsche Forschungsgemeinschaft$$cGA 354/14-1 000010913 536__ $$oDeutsche Forschungsgemeinschaft$$cEXC 2067/1 000010913 536__ $$oSwiss National Science Foundation$$c310030 185298 000010913 536__ $$oNational Institutes of Health$$cR35 GM119840 000010913 540__ $$a<p>Copyright © 2022 The Authors</p> <p>This is an open-access article distributed under the terms of the&nbsp;<a href="https://creativecommons.org/licenses/by-nc/4.0/">Creative Commons Attribution-NonCommercial license</a>, which permits use, distribution, and reproduction in any medium, so long as the resultant use is&nbsp;<span>not</span>&nbsp;for commercial advantage and provided the original work is properly cited.</p> 000010913 542__ $$fCC BY-NC 000010913 594__ $$aAll data needed to evaluate the conclusions in the paper are present in the paper and/or the Supplementary Materials. 000010913 690__ $$aPhysical Sciences Division 000010913 691__ $$aChemistry 000010913 7001_ $$1https://orcid.org/0000-0003-2867-9468$$2ORCID$$aLuebben, Anna V.$$uUniversity of Göttingen 000010913 7001_ $$1https://orcid.org/0000-0002-2975-4186$$2ORCID$$aBender, Daniel$$uUniversity of Zurich 000010913 7001_ $$aBecker, Stefan$$uMax Planck Institute for Biophysical Chemistry 000010913 7001_ $$aCrowther, Lisa M.$$uUniversity of Zurich 000010913 7001_ $$1https://orcid.org/0000-0002-1095-6861$$2ORCID$$aErven, Ilka$$uUniversity of Cologne 000010913 7001_ $$1https://orcid.org/0000-0002-2289-9083$$2ORCID$$aHofmann, Kay$$uUniversity of Cologne 000010913 7001_ $$1https://orcid.org/0000-0001-9642-8244$$2ORCID$$aSöding, Johannes$$uMax-Planck Institute for Biophysical Chemistry 000010913 7001_ $$1https://orcid.org/0000-0002-3147-3017$$2ORCID$$aKlemp, Henry$$uUniversity of Göttingen 000010913 7001_ $$1https://orcid.org/0000-0002-3999-2807$$2ORCID$$aBellotti, Cristina$$uUniversity of Zurich 000010913 7001_ $$1University of Zurich$$2ORCID$$aStäuble, Andreas$$uhttps://orcid.org/0000-0001-9691-1690 000010913 7001_ $$1https://orcid.org/0000-0002-7168-0715$$2ORCID$$aQiu, Tian$$uUniversity of Chicago 000010913 7001_ $$1https://orcid.org/0000-0002-9159-2413$$2ORCID$$aKathayat, Rahul S.$$uUniversity of Chicago 000010913 7001_ $$1https://orcid.org/0000-0002-9616-1911$$2ORCID$$aDickinson, Bryan C.$$uUniversity of Chicago 000010913 7001_ $$aGärtner, Jutta$$uUniversity of Göttingen 000010913 7001_ $$aSheldrick, George M.$$uUniversity of Göttingen 000010913 7001_ $$aKrätzner, Ralph$$uUniversity of Göttingen 000010913 7001_ $$1https://orcid.org/0000-0003-0956-6601$$2ORCID$$aSteinfeld, Robert$$uUniversity of Zurich 000010913 773__ $$tScience Advances 000010913 8564_ $$ySupplementary materials$$9e7fadaa5-4876-49cf-b826-dc3d11867e5f$$s830498$$uhttps://knowledge.uchicago.edu/record/10913/files/sciadv.abj8633_sm.pdf$$ePublic 000010913 8564_ $$yArticle$$93741c51f-862a-4a4a-8124-ea92022228ef$$s1743552$$uhttps://knowledge.uchicago.edu/record/10913/files/sciadv.abj8633.pdf$$ePublic 000010913 908__ $$aI agree 000010913 909CO $$ooai:uchicago.tind.io:10913$$pGLOBAL_SET 000010913 983__ $$aArticle