Switching head group selectivity in mammalian sphingolipid biosynthesis by active-site engineering of sphingomyelin synthases

DC FieldValueLanguage
dc.contributor.authorKol, Matthijs
dc.contributor.authorPanatala, Radhakrishnan
dc.contributor.authorNordmann, Mirjana
dc.contributor.authorSwart, Leoni
dc.contributor.authorvan Suijlekom, Leonie
dc.contributor.authorCabukusta, Birol
dc.contributor.authorHilderink, Angelika
dc.contributor.authorGrabietz, Tanja
dc.contributor.authorMina, John G. M.
dc.contributor.authorSomerharju, Pentti
dc.contributor.authorKorneev, Sergei
dc.contributor.authorTafesse, Fikadu G.
dc.contributor.authorHolthuis, Joost C. M.
dc.date.accessioned2021-12-23T16:19:38Z-
dc.date.available2021-12-23T16:19:38Z-
dc.date.issued2016
dc.identifier.issn00222275
dc.identifier.urihttps://osnascholar.ub.uni-osnabrueck.de/handle/unios/13234-
dc.description.abstractSM is a fundamental component of mammalian cell membranes that contributes to mechanical stability, signaling, and sorting. Its production involves the transfer of phosphocholine from phosphatidylcholine onto ceramide, a reaction catalyzed by SM synthase (SMS) 1 in the Golgi and SMS2 at the plasma membrane. Mammalian cells also synthesize trace amounts of the SM analog ceramide phosphoethanolamine (CPE), but the physiological relevance of CPE production is unclear. Previous work revealed that SMS2 is a bifunctional enzyme producing both SM and CPE, whereas a closely related enzyme, sphingomyelin synthase-related protein (SMSr)/SAMD8, acts as a monofunctional CPE synthase in the endoplasmatic reticulum. Using domain swapping and site-directed mutagenesis on enzymes expressed in defined lipid environments, we here identified structural determinants that mediate head group selectivity of SMS family members. Notably, a single residue adjacent to the catalytic histidine in the third exoplasmic loop profoundly influenced enzyme specificity, with glutamic acid permitting SMS-catalyzed CPE production and aspartic acid confining the enzyme to produce SM. An exchange of exoplasmic residues with SMSr proved sufficient to convert SMS1 into a bulk CPE synthase. This allowed us to establish mammalian cells that produce CPE rather than SM as the principal phosphosphingolipid and provide a model of the molecular interactions that impart catalytic specificity among SMS enzymes.-Kol, M., R. Panatala, M. Nordmann, L. Swart, L. van Suijlekom, B. Cabukusta, A. Hilderink, T. Grabietz, J. G. M. Mina, P. Somerharju, S. Korneev, F. G. Tafesse, and J. C. M. Holthuis. Switching head group selectivity in mammalian sphingolipid biosynthesis by active-site engineering of sphingomyelin synthases.
dc.description.sponsorshipMarie Curie Intra-European FellowshipEuropean Commission; European UnionEuropean Commission [289278]; Deutsche ForschungsgemeinschaftGerman Research Foundation (DFG) [Sonderforschungsbereich/SFB944-P14]; Faculty of Biology/Chemistry from the University of Osnabruck; This work was supported by a Marie Curie Intra-European Fellowship (J.G.M.M.), the European Union Seventh Framework Programme Marie-Curie ITN `Sphingonet' Grant (289278), the Deutsche Forschungsgemeinschaft Sonderforschungsbereich/SFB944-P14 (J.C.M.H.), and an Incentive Award of the Faculty of Biology/Chemistry from the University of Osnabruck (M.K.). The authors declare that they have no conflicts of interest with the contents of this article.
dc.language.isoen
dc.publisherAMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
dc.relation.ispartofJOURNAL OF LIPID RESEARCH
dc.subjectBiochemistry & Molecular Biology
dc.subjectcell-free expression
dc.subjectCELLS
dc.subjectceramide phosphoethanolamine
dc.subjectCERAMIDE PHOSPHOETHANOLAMINE BIOSYNTHESIS
dc.subjectCHOLESTEROL
dc.subjectclick chemistry
dc.subjectenzyme mechanisms
dc.subjectFAMILY
dc.subjectFUNCTIONAL-CHARACTERIZATION
dc.subjectGolgi apparatus
dc.subjectlipid biochemistry
dc.subjectlipidomics
dc.subjectMICE
dc.subjectmodel membranes
dc.subjectPLASMA-MEMBRANE SPHINGOMYELIN
dc.subjectprotein engineering
dc.subjectPROTEIN SMSR
dc.subjectPURIFICATION
dc.subjectTRANSMEMBRANE DOMAINS
dc.titleSwitching head group selectivity in mammalian sphingolipid biosynthesis by active-site engineering of sphingomyelin synthases
dc.typejournal article
dc.identifier.doi10.1194/jlr.M068692
dc.identifier.isiISI:000379653700016
dc.description.volume57
dc.description.issue7
dc.description.startpage1273
dc.description.endpage1285
dc.contributor.orcid0000-0003-0429-1370
dc.contributor.orcid0000-0003-3663-8236
dc.contributor.orcid0000-0002-8575-4164
dc.contributor.researcheridAAL-9157-2020
dc.contributor.researcheridN-8622-2014
dc.contributor.researcheridAAG-2675-2021
dc.contributor.researcheridAAH-7284-2021
dc.identifier.eissn15397262
dc.publisher.place9650 ROCKVILLE PIKE, BETHESDA, MD 20814-3996 USA
dcterms.isPartOf.abbreviationJ. Lipid Res.
dcterms.oaStatusGreen Published, Green Submitted, hybrid, Green Accepted
crisitem.author.deptFB 05 - Biologie/Chemie-
crisitem.author.deptidfb05-
crisitem.author.parentorgUniversität Osnabrück-
crisitem.author.netidKoSe681-
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