UniProt UniProt http://www.uniprot.org Structural elucidation of the <i>cis</i>-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation. Structural elucidation of the <i>cis</i>-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation. Proceedings of the National Academy of Sciences of the United States of America 2020 imex curation Edani BH., Grabińska KA., Zhang R., Park EJ., Siciliano B., Surmacz L., Ha Y., Sessa WC. william.sessa@yale.edu Only protein-protein interactions imex curation ecoli Escherichia coli (strain K12) molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column weight by comassie molecular weight estimation by coomasie staining weight by comassie Edani BH., Grabińska KA., Zhang R., Park EJ., Siciliano B., Surmacz L., Ha Y., Sessa WC. The His-SUMO protein complex was purified on a nickel affinity column,followed by size exclusion chromatography (Superdex 200) The complex was then subjected to cleavage with SUMO protease to remove the His-SUMO tag and again reapplied to a nickel affinity columnwith the flow-through then subjected to size exclusion chromatography (Superdex 200) william.sessa@yale.edu Accepted 2021-MAY-18 AT 09:50 BST AT 09:50 BST by MGT Proceedings of the National Academy of Sciences of the United States of America 2020 Only protein-protein interactions imex curation imex curation Structural elucidation of the <i>cis</i>-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation. Structural elucidation of the <i>cis</i>-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation. Proceedings of the National Academy of Sciences of the United States of America 2020 imex curation Edani BH., Grabińska KA., Zhang R., Park EJ., Siciliano B., Surmacz L., Ha Y., Sessa WC. william.sessa@yale.edu Only protein-protein interactions imex curation human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. anti tag coip anti tag coimmunoprecipitation anti tag coip anti tag western anti tag western blot anti tag western Edani BH., Grabińska KA., Zhang R., Park EJ., Siciliano B., Surmacz L., Ha Y., Sessa WC. william.sessa@yale.edu Accepted 2021-MAY-18 AT 09:50 BST AT 09:50 BST by MGT Proceedings of the National Academy of Sciences of the United States of America 2020 Only protein-protein interactions imex curation imex curation Structural elucidation of the <i>cis</i>-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation. Structural elucidation of the <i>cis</i>-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation. Proceedings of the National Academy of Sciences of the United States of America 2020 imex curation Edani BH., Grabińska KA., Zhang R., Park EJ., Siciliano B., Surmacz L., Ha Y., Sessa WC. william.sessa@yale.edu Only protein-protein interactions imex curation ecoli Escherichia coli (strain K12) x-ray diffraction x-ray crystallography X-ray X-ray x-ray diffraction predetermined predetermined participant predetermined Edani BH., Grabińska KA., Zhang R., Park EJ., Siciliano B., Surmacz L., Ha Y., Sessa WC. william.sessa@yale.edu Accepted 2021-MAY-18 AT 09:50 BST AT 09:50 BST by MGT Proceedings of the National Academy of Sciences of the United States of America 2020 Only protein-protein interactions imex curation imex curation Mock publication and experiment for abstract interactions that are not interaction evidences. Mock publication and experiment for abstract interactions that are not interaction evidences. unspecified method Structural elucidation of the <i>cis</i>-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation. Structural elucidation of the <i>cis</i>-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation. Proceedings of the National Academy of Sciences of the United States of America 2020 imex curation Edani BH., Grabińska KA., Zhang R., Park EJ., Siciliano B., Surmacz L., Ha Y., Sessa WC. william.sessa@yale.edu Only protein-protein interactions imex curation in vitro In vitro enzymatic study enzymatic study predetermined predetermined participant predetermined Edani BH., Grabińska KA., Zhang R., Park EJ., Siciliano B., Surmacz L., Ha Y., Sessa WC. william.sessa@yale.edu Accepted 2021-MAY-18 AT 09:50 BST AT 09:50 BST by MGT Proceedings of the National Academy of Sciences of the United States of America 2020 Only protein-protein interactions imex curation imex curation ngbr_human Dehydrodolichyl diphosphate synthase complex subunit NUS1 NUS1 C6orf68 NGBR Nuclear undecaprenyl pyrophosphate synthase 1 homolog Nogo-B receptor Cis-prenyltransferase subunit NgBR protein protein human Homo sapiens Human MTGLYELVWRVLHALLCLHRTLTSWLRVRFGTWNWIWRRCCRAASAAVLAPLGFTLRKPPAVGRNRRHHRHPRGGSCLAAAHHRMRWRADGRSLEKLPVHMGLVITEVEQEPSFSDIASLVVWCMAVGISYISVYDHQGIFKRNNSRLMDEILKQQQELLGLDCSKYSPEFANSNDKDDQVLNCHLAVKVLSPEDGKADIVRAAQDFCQLVAQKQKRPTDLDVDTLASLLSSNGCPDPDLVLKFGPVDSTLGFLPWHIRLTEIVSLPSHLNISYEDFFSALRQYAACEQRLGK DIP protein Q96E22 original sequence version: 105 mint Acts as a specific receptor for the N-terminus of Nogo-B, a neural and cardiovascular regulator. Able to regulate vascular remodeling and angiogenesis. Its similarity with UPP synthetase proteins suggests that it may act as a scaffold for the binding of isoprenyl lipids and/or prenylated proteins homomint E6A0C9D9B39D4BCA 2-trans,6-trans-farnesyl diphosphate(3-) 2-trans,6-trans-farnesyl diphosphate(3-) (2-trans,6-trans)-farnesyl diphosphate(3-) (2E,6E)-3,7,11-trimethyldodeca-2,6,10-trien-1-yl (phosphonatooxy)phosphonate (2E,6E)-farnesyl diphosphate (2E,6E)-farnesyl diphosphate(3-) (E,E)-farnesyl diphosphate(3-) farnesyl pyrophosphate(3-) trans,trans-farnesyl diphosphate(3-) (2E,6E)-3,7,11-trimethyldodeca-2,6,10-trien-1-yl diphosphate small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) pi Phosphatidylinositol phosphatidylinositols PI PtdIns small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) magnesium(2+) Magnesium(2+) small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) dhdds_human Dehydrodolichyl diphosphate synthase complex subunit DHDDS DHDDS Cis-isoprenyltransferase Cis-prenyltransferase subunit hCIT Epididymis tissue protein Li 189m HDS protein protein human Homo sapiens Human MSWIKEGELSLWERFCANIIKAGPMPKHIAFIMDGNRRYAKKCQVERQEGHSQGFNKLAETLRWCLNLGILEVTVYAFSIENFKRSKSEVDGLMDLARQKFSRLMEEKEKLQKHGVCIRVLGDLHLLPLDLQELIAQAVQATKNYNKCFLNVCFAYTSRHEISNAVREMAWGVEQGLLDPSDISESLLDKCLYTNRSPHPDILIRTSGEVRLSDFLLWQTSHSCLVFQPVLWPEYTFWNLFEAILQFQMNHSVLQKARDMYAEERKRQQLERDQATVTEQLLREGLQASGDAQLRRTRLHKLSARREERVQGFLQALELKRADWLARLGTASA 12EF3A15437A2583 dhdds-nus1-1 3 6 unspecified role unspecified role neutral component neutral component purified purified region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 10 unspecified role unspecified role neutral component neutral component purified purified region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain direct interaction direct interaction Supp.Table 1 dhdds-nus1-2 1 6 unspecified role unspecified role neutral component neutral component purified purified region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 10 unspecified role unspecified role neutral component neutral component purified purified region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain direct interaction direct interaction Fig. 1B dhdds-nus1-3 2 6 unspecified role unspecified role prey prey over-expressed over expressed level over-expressed cell lysate cell lysate Q96E22:p.Lys243Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain K A Q96E22:p.Asp248Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain D A Q96E22:p.Gly252Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain G A Q96E22:p.Phe253Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain F A Q96E22:p.Pro255Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain P A Q96E22:p.Leu260Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain L A Q96E22:p.Ile263Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain I A Q96E22:p.Gly292Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain G A c-terminal ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range Q96E22:p.Gln180Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain Q A Q96E22:p.Gly196Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain G A Q96E22:p.Lys197Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain K A Q96E22:p.Ile200Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain I A Q96E22:p.Leu226Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain L A Q96E22:p.Leu230Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain L A Q96E22:p.Pro238Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain P A 10 unspecified role unspecified role bait bait over-expressed over expressed level over-expressed cell lysate cell lysate n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range physical association physical association Fig.2C dds_human-farnesyl triphosphate(3_)-1 5 43 enzyme enzyme unspecified role unspecified role purified purified Q96E22:p.Gly91Cys mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate certain certain sequence position certain certain certain sequence position certain G C 45 Q86SQ9:p.[Trp12Ala;Phe15Ala;Ile19Ala] mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain W A 47 certain certain sequence position certain certain certain sequence position certain F A 47 certain certain sequence position certain certain certain sequence position certain I A 47 ecoli Escherichia coli (strain K12) 7 enzyme target enzyme target substrate substrate unspecified role unspecified role enzymatic reaction enzymatic reaction Biochemical reaction Biochemical reaction Figs.1C, 4C, 5D 2_trans,6_trans_farnesyl diphosphate(3_)-dds_human-1 5 43 enzyme enzyme unspecified role unspecified role purified purified Q96E22:p.Gly91Cys mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate certain certain sequence position certain certain certain sequence position certain G C 45 Q86SQ9:p.[Trp12Ala;Phe15Ala;Ile19Ala] mutation decreasing rate mutation decreasing interaction rate mutation decreasing rate certain certain sequence position certain certain certain sequence position certain F A 47 certain certain sequence position certain certain certain sequence position certain W A 47 certain certain sequence position certain certain certain sequence position certain I A 47 ecoli Escherichia coli (strain K12) 8 enzyme regulator enzyme regulator unspecified role unspecified role 7 enzyme target enzyme target substrate substrate unspecified role unspecified role enzymatic reaction enzymatic reaction Biochemical reaction Biochemical reaction Fig.1C, 5D The activity of the NgBR/DHDDS complex was potently stimulated by phosphatidylinositol dds_human Cis-PTase complex NgBR-DHDDS complex 2xDHDDS:2xNUS1 Cis-IPTase complex Dehydrodolichyl diphosphate synthase complex DDS complex Cis-prenyltransferase complex 6 unspecified role unspecified role dhdds binding region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 6 unspecified role unspecified role dhdds binding region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 9 cofactor cofactor coenzyme dhdds binding region binding region binding-associated region binding region undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 10 enzyme enzyme dhdds binding region binding region binding-associated region binding region undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined nus1 binding region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain magnesium binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain 9 cofactor cofactor coenzyme dhdds binding region binding region binding-associated region binding region undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 10 enzyme enzyme dhdds binding region binding region binding-associated region binding region undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined nus1 binding region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain magnesium binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain 56 62 67 57 63 59 66 61 65 68 physical association physical association human Homo sapiens Human stable complex stable complex physical interaction evidence used in manual assertion physical interaction evidence used in manual assertion IPI inferred from physical interaction A cis-prenyltransferase which catalyzes the rate-limiting step in the synthesis of glycosyl carrier lipids required for protein glycosylation in the lumen of endoplasmic reticulum. Essential for the synthesis of dolichol phosphate (CHEBI:23875), an indispensable lipid carrier for protein N-glycosylation, O-mannosylation, C-mannosylation, and GPI-anchor formation, catalyzing the formation of its precursor dehydrodolichyl diphosphate (ditrans,polycis-polyprenyl diphosphate(3-), CHEBI:136960), a long-chain isoprenoid, by chain elongation of farnesyl diphosphate (CHEBI:175763) via multiple condensations with isopentenyl pyrophosphate (CHEBI:128769). The DHDDS-NUS1 heterodimer is formed through a large interaction interface with the C-terminus of NUS1 which contributes to enzymatic activity through its C-terminal -RXG- motif, extending across the dimerization interface. The heterotetrameric assembly is formed via a dimer-of-heterodimers mechanism, mainly through homotypic interface formation between the C-terminal helix-turn-helix motifs of DHDDS, and interaction between this motif and the NUS1 protein in the adjoining heterodimer. NUS1 lacks catalytic activity due to an absence of substrate binding sites. Heterotetramer Retinitis pigmentosa (RP) [Orphanet:791]: an inherited retinal dystrophy leading to progressive loss of the photoreceptors and retinal pigment epithelium and resulting in blindness usually after several decades. Congenital disorder of glycosylation type I [EFO:0005545]: A congenital disorder of glycosylation involve disrupted synthesis of the lipid-linked oligosaccharide precursor. Developmental delay and seizures with or without movement abnormalities (DEDSM) [MONDO:0044326]: a neurodevelopmental disorder characterized by global developmental delay, variable intellectual disability, and early-onset seizures with a myoclonic component. Most patients have delayed motor development and show abnormal movements, including ataxia, dystonia, and tremor. Global developmental delay - neuro-ophthalmological abnormalities - seizures - intellectual disability syndrome [Orphanet:488613]: a form of mental retardation, a disorder characterized by significantly below average general intellectual functioning associated with impairments in adaptive behavior and manifested during the developmental period. MRD55 patients suffer from seizures appearing during the first years of life. Accepted 2021-MAY-25 by BMELDAL