{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,7]],"date-time":"2026-03-07T11:13:51Z","timestamp":1772882031867,"version":"3.50.1"},"reference-count":36,"publisher":"Oxford University Press (OUP)","issue":"10","license":[{"start":{"date-parts":[[2019,7,8]],"date-time":"2019-07-08T00:00:00Z","timestamp":1562544000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/journals\/pages\/open_access\/funder_policies\/chorus\/standard_publication_model"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["R01-GM037539"],"award-info":[{"award-number":["R01-GM037539"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["R01-GM084383"],"award-info":[{"award-number":["R01-GM084383"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2019,9,20]]},"abstract":"<jats:title>Abstract<\/jats:title>\n               <jats:p>Skp1 is hydroxylated by an O2-dependent prolyl hydroxylase (PhyA) that contributes to O2-sensing in the social amoeba Dictyostelium and the mammalian pathogen Toxoplasma gondii. HO-Skp1 is subject to glycosylation and the resulting pentasaccharide affects Skp1 conformation in a way that influences association of Skp1 with F-box proteins, and potentially the assembly of E3(SCF) ubiquitin ligase complexes that mediate the polyubiquitination of target proteins that are degraded in the 26S-proteasome. To investigate the conservation and specificity of these modifications, we analyzed proteins from the oomycete Pythium ultimum, an important crop plant pathogen. Putative coding sequences for Pythium\u2019s predicted PhyA and first glycosyltransferase in the predicted five-enzyme pathway, a GlcNAc-transferase (Gnt1), predict a bifunctional enzyme (Phgt) that, when expressed in Dictyostelium, rescued a knockout of phyA but not gnt1. Though recombinant Phgt was also unable to glycosylate Dictyostelium HO-Skp1, it could hydrolyze UDP-GlcNAc and modify a synthetic hydroxypeptide from Dictyostelium Skp1. Pythium encodes two highly similar Skp1 isoforms, but only Skp1A was efficiently hydroxylated and glycosylated in vitro. While kinetic analysis revealed no evidence for processive processing of Skp1, the physical linkage of the two activities implies dedication to Skp1 in vivo. These findings indicate a widespread occurrence of the Skp1 modification pathway across protist phylogeny, suggest that both Gnt1 and PhyA are specific for Skp1 and indicate that the second Skp1 provides a bypass mechanism for O2-regulation in Pythium and other protists that conserve this gene.<\/jats:p>","DOI":"10.1093\/glycob\/cwz049","type":"journal-article","created":{"date-parts":[[2019,7,5]],"date-time":"2019-07-05T15:08:52Z","timestamp":1562339332000},"page":"705-714","source":"Crossref","is-referenced-by-count":13,"title":["Skp1 isoforms are differentially modified by a dual function prolyl 4-hydroxylase\/N-acety lglucosaminyltransferase in a plant pathogen"],"prefix":"10.1093","volume":"29","author":[{"given":"Hanke","family":"van der Wel","sequence":"first","affiliation":[{"name":"Department of Biochemistry & Molecular Biology, Center for Tropical & Emerging Global Diseases, Complex Carbohydrate Research Center, University of Georgia, Athens, GA, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Elisabet","family":"Gas-Pascual","sequence":"additional","affiliation":[{"name":"Department of Biochemistry & Molecular Biology, Center for Tropical & Emerging Global Diseases, Complex Carbohydrate Research Center, University of Georgia, Athens, GA, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9077-965X","authenticated-orcid":false,"given":"Christopher M","family":"West","sequence":"additional","affiliation":[{"name":"Department of Biochemistry & Molecular Biology, Center for Tropical & Emerging Global Diseases, Complex Carbohydrate Research Center, University of Georgia, Athens, GA, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"286","published-online":{"date-parts":[[2019,10,2]]},"reference":[{"key":"2019102301191039400_ref1","doi-asserted-by":"crossref","first-page":"463","DOI":"10.1146\/annurev-micro-091014-104100","article-title":"Lytic cycle of Toxoplasma gondii: 15 years later","volume":"69","author":"Blader","year":"2015","journal-title":"Annu Rev Microbiol."},{"key":"2019102301191039400_ref2","doi-asserted-by":"crossref","first-page":"3932","DOI":"10.1074\/jbc.273.7.3932","article-title":"The RPA32 subunit of human replication protein A contains a single-stranded DNA-binding domain","volume":"273","author":"Bochkareva","year":"1998","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref3","doi-asserted-by":"crossref","first-page":"399","DOI":"10.1146\/annurev.biochem.78.101807.093809","article-title":"RING domain E3 ubiquitin ligases","volume":"78","author":"Deshaies","year":"2009","journal-title":"Annu Rev Biochem."},{"key":"2019102301191039400_ref4","doi-asserted-by":"crossref","first-page":"918","DOI":"10.1093\/glycob\/cwp068","article-title":"Molecular analysis of a UDP-GlcNAc:polypeptide alpha-N-acetylglucosaminyltransferase implicated in the initiation of mucin-type O-glycosylation in Trypanosoma cruzi","volume":"19","author":"Heise","year":"2009","journal-title":"Glycobiology."},{"key":"2019102301191039400_ref5","doi-asserted-by":"crossref","first-page":"585","DOI":"10.1146\/annurev-biochem-061516-044724","article-title":"2-oxoglutarate-dependent oxygenases","volume":"87","author":"Islam","year":"2018","journal-title":"Annu Rev Biochem."},{"key":"2019102301191039400_ref6","doi-asserted-by":"crossref","first-page":"3022","DOI":"10.1074\/jbc.270.7.3022","article-title":"Characterization of FP21, a cytosolic glycoprotein from Dictyostelium","volume":"270","author":"Kozarov","year":"1995","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref7","doi-asserted-by":"crossref","first-page":"R73","DOI":"10.1186\/gb-2010-11-7-r73","article-title":"Genome sequence of the necrotrophic plant pathogen Pythium ultimum reveals original pathogenicity mechanisms and effector repertoire","volume":"11","author":"L\u00e9vesque","year":"2010","journal-title":"Genome Biol."},{"key":"2019102301191039400_ref8","doi-asserted-by":"crossref","first-page":"43701","DOI":"10.1074\/jbc.M111.309096","article-title":"Mimivirus collagen is modified by bifunctional lysyl hydroxylase and glycosyltransferase enzyme","volume":"286","author":"Luther","year":"2011","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref9","doi-asserted-by":"crossref","first-page":"4268","DOI":"10.1074\/jbc.M115.703751","article-title":"The E3 ubiquitin ligase adaptor protein Skp1 is glycosylated by an evolutionarily conserved pathway that regulates protist growth and development","volume":"291","author":"Rahman","year":"2016","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref10","doi-asserted-by":"crossref","first-page":"18644","DOI":"10.1074\/jbc.M117.809301","article-title":"Characterization of a cytoplasmic glucosyltransferase that extends the core trisaccharide of the Toxoplasma Skp1 E3 ubiquitin ligase subunit","volume":"292","author":"Rahman","year":"2017","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref11","doi-asserted-by":"crossref","first-page":"1657","DOI":"10.1021\/bi401707y","article-title":"Glycosylation of Skp1 affects its conformation and promotes binding to a model F-box protein","volume":"53","author":"Sheikh","year":"2014","journal-title":"Biochemistry."},{"key":"2019102301191039400_ref12","doi-asserted-by":"crossref","first-page":"66","DOI":"10.1074\/mcp.M114.044560","article-title":"Glycosylation of Skp1 promotes formation of Skp1\/Cullin-1\/F-box protein complexes in Dictyostelium","volume":"14","author":"Sheikh","year":"2015","journal-title":"Mol Cell Proteomics."},{"key":"2019102301191039400_ref13","doi-asserted-by":"crossref","first-page":"206","DOI":"10.1093\/glycob\/cww114","article-title":"Rapid screening of sugar-nucleotide donor specificities of putative glycosyltransferases","volume":"27","author":"Sheikh","year":"2017","journal-title":"Glycobiology."},{"key":"2019102301191039400_ref14","doi-asserted-by":"crossref","first-page":"18897","DOI":"10.1074\/jbc.M117.809160","article-title":"O2 sensing\u2013associated glycosylation exposes the F-box\u2013combining site of the Dictyostelium Skp1 subunit in E3 ubiquitin ligases","volume":"292","author":"Sheikh","year":"2017","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref15","doi-asserted-by":"crossref","first-page":"583","DOI":"10.1007\/s12010-013-0326-8","article-title":"Identification of amino acid residues responsible for the binding to anti-FLAG\u2122 M2 antibody using a phage display combinatorial peptide library","volume":"171","author":"Srila","year":"2013","journal-title":"Appl Biochem Biotechnol."},{"key":"2019102301191039400_ref16","doi-asserted-by":"crossref","first-page":"36392","DOI":"10.1074\/jbc.274.51.36392","article-title":"Identification of a UDP-GlcNAc:Skp1-hydroxyproline GlcNAc-transferase in the cytoplasm of Dictyostelium","volume":"274","author":"Teng-umnuay","year":"1999","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref17","doi-asserted-by":"crossref","first-page":"46527","DOI":"10.1074\/jbc.M208824200","article-title":"A bifunctional diglycosyltransferase forms the Fucalpha1,2Galbeta1,3-disaccharide on Skp1 in the cytoplasm of Dictyostelium","volume":"277","author":"van der Wel","year":"2002","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref18","doi-asserted-by":"crossref","first-page":"46328","DOI":"10.1074\/jbc.M208024200","article-title":"Molecular cloning and expression of a UDP-N-acetylglucosamine (GlcNAc):Hydroxyproline polypeptide GlcNAc-transferase that modifies Skp1 in the cytoplasm of Dictyostelium","volume":"277","author":"van der Wel","year":"2002","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref19","doi-asserted-by":"crossref","first-page":"14645","DOI":"10.1074\/jbc.M500600200","article-title":"The Skp1 prolyl hydroxylase from Dictyostelium is related to the hypoxia-inducible factor-alpha class of animal prolyl 4-hydroxylases","volume":"280","author":"van der Wel","year":"2005","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref20","doi-asserted-by":"crossref","first-page":"1700","DOI":"10.1021\/bi101977w","article-title":"Requirements for Skp1 processing by cytosolic prolyl 4(trans)-hydroxylase and alpha-N-acetylglucosaminyltransferase enzymes involved in O2 signaling in Dictyostelium","volume":"50","author":"van der Wel","year":"2011","journal-title":"Biochemistry."},{"key":"2019102301191039400_ref21","doi-asserted-by":"crossref","first-page":"110","DOI":"10.1016\/j.plasmid.2008.11.003","article-title":"A new set of small, extrachromosomal expression vectors for Dictyostelium discoideum","volume":"61","author":"Veltman","year":"2009","journal-title":"Plasmid."},{"key":"2019102301191039400_ref22","doi-asserted-by":"crossref","first-page":"18568","DOI":"10.1074\/jbc.M201389200","article-title":"Identification of amino acids important for the catalytic activity of the collagen glucosyltransferase associated with the multifunctional lysyl hydroxylase 3 (LH3)","volume":"277","author":"Wang","year":"2002","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref23","doi-asserted-by":"crossref","first-page":"28896","DOI":"10.1074\/jbc.M109.022574","article-title":"Role of a cytoplasmic dual-function glycosyltransferase in O2-regulation of development in Dictyostelium","volume":"284","author":"Wang","year":"2009","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref24","doi-asserted-by":"crossref","first-page":"283","DOI":"10.1016\/j.ydbio.2010.10.013","article-title":"Prolyl hydroxylation- and glycosylation-dependent functions of Skp1 in O2-regulated development of Dictyostelium","volume":"349","author":"Wang","year":"2011","journal-title":"Dev Biol."},{"key":"2019102301191039400_ref25","doi-asserted-by":"crossref","first-page":"204","DOI":"10.1016\/j.sbi.2019.03.031","article-title":"Nucleocytoplasmic O-glycosylation in protists","volume":"56","author":"West","year":"2019","journal-title":"Curr Opin Struct Biol."},{"key":"2019102301191039400_ref26","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/S0378-1119(97)00194-7","article-title":"The cytosolic glycoprotein FP21 of Dictyostelium discoideum is encoded by two genes resulting in a polymorphism at a single amino acid position","volume":"200","author":"West","year":"1997","journal-title":"Gene."},{"key":"2019102301191039400_ref27","doi-asserted-by":"crossref","first-page":"3349","DOI":"10.1242\/dev.000893","article-title":"Prolyl 4-hydroxylase-1 mediates O2 signaling during development of Dictyostelium","volume":"134","author":"West","year":"2007","journal-title":"Development."},{"key":"2019102301191039400_ref28","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1016\/j.mib.2015.04.006","article-title":"Oxygen sensing by protozoans: How they catch their breath","volume":"26","author":"West","year":"2015","journal-title":"Curr Opin Microbiol."},{"key":"2019102301191039400_ref29","doi-asserted-by":"crossref","first-page":"927","DOI":"10.1007\/978-4-431-54841-6_170","volume-title":"Glycoscience: Biology and Medicine","author":"West","year":"2015"},{"key":"2019102301191039400_ref30","volume-title":"Essentials of Glycobiology [Internet]","author":"West","year":"2017","edition":"3rd"},{"key":"2019102301191039400_ref31","doi-asserted-by":"crossref","first-page":"759","DOI":"10.1016\/j.pt.2018.07.006","article-title":"Apicomplexa cell cycles: Something old, borrowed, lost, and new","volume":"34","author":"White","year":"2018","journal-title":"Trends Parasitol."},{"key":"2019102301191039400_ref32","doi-asserted-by":"crossref","first-page":"133","DOI":"10.1016\/j.bbamcr.2004.09.027","article-title":"A hitchhiker's guide to the cullin ubiquitin ligases: SCF and its kin","volume":"1695","author":"Willems","year":"2004","journal-title":"Biochim Biophys Acta."},{"key":"2019102301191039400_ref33","doi-asserted-by":"crossref","first-page":"25098","DOI":"10.1074\/jbc.M112.355446","article-title":"The Skp1 protein from Toxoplasma is modified by a cytoplasmic prolyl 4-hydroxylase associated with oxygen sensing in the social amoeba Dictyostelium","volume":"287","author":"Xu","year":"2012","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref34","doi-asserted-by":"crossref","first-page":"511","DOI":"10.1021\/acs.biochem.7b01033","article-title":"Glycosylation promotes the random coil to helix transition in a region of a protist Skp1 associated with F-box binding","volume":"57","author":"Xu","year":"2018","journal-title":"Biochemistry."},{"key":"2019102301191039400_ref35","doi-asserted-by":"crossref","first-page":"2006","DOI":"10.1074\/jbc.M111.314021","article-title":"Skp1 prolyl 4-hydroxylase of Dictyostelium mediates glycosylation-independent and -dependent responses to O2 without affecting Skp1 stability","volume":"287","author":"Zhang","year":"2012","journal-title":"J Biol Chem."},{"key":"2019102301191039400_ref36","doi-asserted-by":"crossref","first-page":"14.1","DOI":"10.1146\/annurev-biochem-060815-014922","article-title":"Ubiquitin ligases: Structure, function, and regulation","volume":"86","author":"Zheng","year":"2017","journal-title":"Annu Rev Biochem."}],"container-title":["Glycobiology"],"original-title":[],"language":"en","link":[{"URL":"http:\/\/academic.oup.com\/glycob\/advance-article-pdf\/doi\/10.1093\/glycob\/cwz049\/29808355\/cwz049.pdf","content-type":"application\/pdf","content-version":"am","intended-application":"syndication"},{"URL":"http:\/\/academic.oup.com\/glycob\/article-pdf\/29\/10\/705\/30262053\/cwz049.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"http:\/\/academic.oup.com\/glycob\/article-pdf\/29\/10\/705\/30262053\/cwz049.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2019,10,23]],"date-time":"2019-10-23T01:19:23Z","timestamp":1571793563000},"score":1,"resource":{"primary":{"URL":"https:\/\/academic.oup.com\/glycob\/article\/29\/10\/705\/5528792"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2019,9,20]]},"references-count":36,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2019,10,2]]},"published-print":{"date-parts":[[2019,9,20]]}},"URL":"https:\/\/doi.org\/10.1093\/glycob\/cwz049","relation":{},"ISSN":["1460-2423"],"issn-type":[{"value":"1460-2423","type":"electronic"}],"subject":[],"published-other":{"date-parts":[[2019,10]]},"published":{"date-parts":[[2019,9,20]]}}}