{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,9]],"date-time":"2026-10-09T16:27:32Z","timestamp":1791563252962,"version":"4.3.4"},"reference-count":28,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2020,3,3]],"date-time":"2020-03-03T00:00:00Z","timestamp":1583193600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Galactosemia Initiative Germany","award":["--"],"award-info":[{"award-number":["--"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cells"],"abstract":"<jats:p>Genetic defects of human galactose-1-phosphate uridyltransferase (hGALT) and the partial loss of enzyme function result in an altered galactose metabolism with serious long-term developmental impairment of organs in classic galactosemia patients. In search for cellular pathomechanisms induced by the stressor galactose, we looked for ways to induce metabolically a galactosemia-like phenotype by hGALT inhibition in HEK293 cells. In kinetic studies, we provide evidence for 2-fluorinated galactose-1-phosphate (F-Gal-1-P) to competitively inhibit recombinant hGALT with a KI of 0.9 mM. Contrasting with hepatic cells, no alterations of N-glycoprofiles in MIG (metabolic induction of galactosemia)-HEK293 cells were revealed for an inducible secretory netrin-1 probe by MALDI-MS. Differential fluorescence-activated cell sorting demonstrated reduced surface expression of N-glycosylated CD109, EGFR, DPP4, and rhMUC1. Membrane raft proteomes exhibited dramatic alterations pointing to an affection of the unfolded protein response, and of targeted protein traffick. Most prominent, a negative regulation of oxidative stress was revealed presumably as a response to a NADPH pool depletion during reduction of Gal\/F-Gal. Cellular perturbations induced by fluorinated galactoses in normal epithelial cells resemble proteomic changes revealed for galactosemic fibroblasts. In conclusion, the metabolic induction of galactosemia-like phenotypes in healthy epithelial\/neuronal cells could support studies on the molecular pathomechanisms in classic galactosemia, in particular under conditions of low galactose stress and residual GALT activity.<\/jats:p>","DOI":"10.3390\/cells9030607","type":"journal-article","created":{"date-parts":[[2020,3,3]],"date-time":"2020-03-03T13:06:23Z","timestamp":1583240783000},"page":"607","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":12,"title":["Fluorinated Galactoses Inhibit Galactose-1-Phosphate Uridyltransferase and Metabolically Induce Galactosemia-like Phenotypes in HEK-293 Cells"],"prefix":"10.3390","volume":"9","author":[{"given":"Verena","family":"Janes","sequence":"first","affiliation":[{"name":"Institute of Biochemistry II, Medical Faculty, University of Cologne, 50931 Koeln, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Simona","family":"Grabany","sequence":"additional","affiliation":[{"name":"Institute of Biochemistry II, Medical Faculty, University of Cologne, 50931 Koeln, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Julien","family":"Delbrouck","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Laboratory of Bio-organic Chemistry, University of Namur (FUNDP), B-5000 Namur, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6258-9058","authenticated-orcid":false,"given":"Stephane P.","family":"Vincent","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Laboratory of Bio-organic Chemistry, University of Namur (FUNDP), B-5000 Namur, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Johannes","family":"Gottschalk","sequence":"additional","affiliation":[{"name":"Helmholtz Institute for Biomedical Engineering, 52074 Aachen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lothar","family":"Elling","sequence":"additional","affiliation":[{"name":"Helmholtz Institute for Biomedical Engineering, 52074 Aachen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Franz-Georg","family":"Hanisch","sequence":"additional","affiliation":[{"name":"Institute of Biochemistry II, Medical Faculty, University of Cologne, 50931 Koeln, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,3,3]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"516","DOI":"10.1007\/s10545-006-0382-0","article-title":"Classical galactosaemia revisited","volume":"29","author":"Bosch","year":"2006","journal-title":"J. Inherit. Metab. Dis."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"351","DOI":"10.1093\/glycob\/8.4.351","article-title":"Defective galactosylation of serum transferrin in galactosemia","volume":"8","author":"Charlwood","year":"1998","journal-title":"Glycobiology"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"906","DOI":"10.1021\/pr200711w","article-title":"Differential glycomics of epithelial membrane glycoproteins from urinary exovesicles reveals shifts toward complex-type N-glycosylation in classical galactosemia","volume":"11","author":"Staubach","year":"2012","journal-title":"J. Proteome Res."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1754","DOI":"10.1021\/acs.jproteome.5b00902","article-title":"Differential proteomics of urinary exovesicles from classical galactosemic patients reveals subclinical kidney insuffiency","volume":"15","author":"Staubach","year":"2016","journal-title":"J. Proteome Res."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"516","DOI":"10.1021\/acs.jproteome.6b00658","article-title":"Classical galactosemia: Insight into molecular pathomechanisms by differential membrane proteomics of fibroblasts under galactose stress","volume":"16","author":"Staubach","year":"2017","journal-title":"J. Proteome Res."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"78","DOI":"10.1016\/j.ymgme.2007.06.005","article-title":"Involvement of endoplasmic reticulum stress in a novel Classic Galactosemia model","volume":"92","author":"Slepak","year":"2007","journal-title":"Mol. Genet. Metab."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"C1","DOI":"10.1152\/ajpcell.00333.2005","article-title":"Role of N- and O-glycans in polarized biosynthetic sorting","volume":"290","author":"Potter","year":"2006","journal-title":"Am. J. Physiol. Cell. Physiol."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Shaw, G., Morse, S., Ararat, M., and Graham, F.L. (2002). Preferential transformation of human neuronal cells by human adenoviruses and the origin of HEK 293 cells. FASEB J.","DOI":"10.1096\/fj.01-0995fje"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1298","DOI":"10.1002\/biot.201600265","article-title":"Multiplexed capillary electrophoresis as analytical tool for fast optimization of multi-enzyme cascade reactions\u2014Synthesis of nucleotide sugars","volume":"11","author":"Wahl","year":"2016","journal-title":"Biotechnol. J."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"51","DOI":"10.1016\/j.jbiotec.2017.03.025","article-title":"Characterization of a new UDP-sugar pyrophosphorylase from Hordeum vulgare (barley)","volume":"258","author":"Wahl","year":"2017","journal-title":"Biotechnol. J."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Fisch\u00f6der, T., Laaf, D., Dey, C., and Elling, L. (2017). Enzymatic Synthesis of N-Acetyllactosamine (LacNAc) Type 1 Oligomers and Characterization as Multivalent Galectin Ligand. Molecules, 22.","DOI":"10.3390\/molecules22081320"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"6704","DOI":"10.1002\/chem.200800734","article-title":"Fragmentation of carbohydrate anomeric alkoxyl radicals: New synthesis of chiral 1-fluoro-1-halo-1-iodoalditols","volume":"14","author":"Francisco","year":"2008","journal-title":"Chem. Eur. J."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"392","DOI":"10.1002\/ijch.201400148","article-title":"\u03b2-Selective One-Pot Fluorophosphorylation of d,d-Heptosylglycals Mediated by Selectfluor","volume":"55","author":"Vincent","year":"2015","journal-title":"Isr. J. Chem."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"15208","DOI":"10.1002\/chem.201404180","article-title":"Selectfluor and NFSI exo-glycal fluorination strategies applied to the enhancement of the binding affinity of galactofuranosyl transferase GalfT2 inhibitors","volume":"20","author":"Dumitrescu","year":"2014","journal-title":"Chem. Eur. J."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"9530","DOI":"10.1002\/chem.200801279","article-title":"Stereoselective glycal fluorophosphorylation: Synthesis of ADP-2-fluoroheptose, an inhibitor of the LPS biosynthesis","volume":"14","author":"Dohi","year":"2008","journal-title":"Chem. Eur. J."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1111","DOI":"10.1093\/glycob\/cwi099","article-title":"Transmembrane and secreted MUC1 probes show trafficking-dependent changes in O-glycan core profiles","volume":"15","author":"Engelmann","year":"2005","journal-title":"Glycobiology"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"22847","DOI":"10.1074\/jbc.M001053200","article-title":"Functional consequence of substitutions at residue 171 in human galactose-1-phosphate uridylyltransferase","volume":"275","author":"Crews","year":"2000","journal-title":"J. Biol. Chem."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"20160103","DOI":"10.1098\/rsif.2016.0103","article-title":"Neutral glycans from sandfish skin can reduce friction of polymers","volume":"13","author":"Vihar","year":"2016","journal-title":"J. R. Soc. Interface"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"217","DOI":"10.1016\/0014-5793(91)81433-9","article-title":"2-Deoxy-2-fluoro-D-galactose protein N-glycosylation","volume":"294","author":"Loch","year":"1991","journal-title":"FEBS Lett."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"821","DOI":"10.1042\/bj2850821","article-title":"Inhibition of protein N-glycosylation by 2-deoxy-2-fluoro-D-galactose","volume":"285","author":"Gross","year":"1992","journal-title":"Biochem. J."},{"key":"ref_21","first-page":"55","article-title":"The unfolded protein response has a protective role in yeast models of classic galactosemia","volume":"7","author":"Pimentel","year":"2014","journal-title":"Dis. Models Mech."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"618","DOI":"10.1242\/dmm.005041","article-title":"A Drosophila Melanogaster model of classic galactosemia","volume":"3","author":"Kushner","year":"2010","journal-title":"Dis. Models Mech."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1172","DOI":"10.1038\/ejhg.2014.12","article-title":"Subfertility and growth restriction in a new galactose-1-phosphate uridyltransferase (GALT)-deficient mouse model","volume":"22","author":"Tang","year":"2014","journal-title":"Eur. J. Hum. Genet."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"325","DOI":"10.1007\/s10545-017-0029-3","article-title":"Sweet and sour: An update on classic galactosemia","volume":"40","author":"Coelho","year":"2017","journal-title":"J. Inherit. Metab. Dis."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"786","DOI":"10.1016\/j.cell.2014.01.024","article-title":"Regulation of TORC1 in response to amino acid starvation via lysosomal recruitment of TSC2","volume":"156","author":"Demetriades","year":"2014","journal-title":"Cell"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1177","DOI":"10.1534\/genetics.111.133363","article-title":"Target of rapamycin (TOR) in nutrient signaling and growth control","volume":"189","author":"Loewith","year":"2011","journal-title":"Genetics"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"7215","DOI":"10.1073\/pnas.0500297102","article-title":"Insights into TOR function and rapamycin response: Chemical genomic profiling by using a high-density cell array method","volume":"102","author":"Xie","year":"2005","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1021\/acs.jproteome.9b00586","article-title":"Redox proteomes in human physiology and disease mechanisms","volume":"19","author":"Mannaa","year":"2020","journal-title":"J. Proteome Res."}],"container-title":["Cells"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4409\/9\/3\/607\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T09:03:51Z","timestamp":1760173431000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4409\/9\/3\/607"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,3,3]]},"references-count":28,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2020,3]]}},"alternative-id":["cells9030607"],"URL":"https:\/\/doi.org\/10.3390\/cells9030607","relation":{},"ISSN":["2073-4409"],"issn-type":[{"value":"2073-4409","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,3,3]]}}}