The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Manfred Wuhrer - One of the best experts on this subject based on the ideXlab platform.
-
acpa igg Galactosylation associates with disease activity in pregnant patients with rheumatoid arthritis
Annals of the Rheumatic Diseases, 2018Co-Authors: Albert Bondt, Manfred Wuhrer, Lise Hafkenscheid, David Falck, Martijn T Kuijper, Yoann Rombouts, Johanna M W Hazes, Radboud J E M DolhainAbstract:Objectives Patients with autoantibody-positive rheumatoid arthritis (RA) are less likely to experience pregnancy-induced improvement of RA disease activity (DAS28-C reactive protein (CRP)) compared with patients with autoantibody-negative RA. Anti-citrullinated protein antibodies (ACPAs) are the most specific autoantibodies for RA. We previously demonstrated that disease improvement is associated with changes in total IgG glycosylation, which regulate antibody effector function. Therefore, we sought to analyse the ACPA-IgG glycosylation profile during pregnancy with the aim to understand the lower change of pregnancy-induced improvement of the disease in patients with autoantibody-positive RA. Methods ACPA-IgGs were purified from ACPA-positive patient sera (n=112) of the Pregnancy-induced Amelioration of Rheumatoid Arthritis cohort, a prospective study designed to investigate pregnancy-associated improvement of RA. The fragment crystallisable (Fc)glycosylation profile of ACPA-IgGs was characterised by mass spectrometry and compared with that of total IgG derived from the same patients or from ACPA-negative patients. Results All ACPA-IgG subclasses display significant changes in the level of Galactosylation and sialylation during pregnancy, although less pronounced than in total IgG. The pregnancy-induced increase in ACPA-IgG Galactosylation, but not sialylation, associates with lower DAS28-CRP. In ACPA-positive patients, no such association was found with changes in the Galactosylation of total IgG, whereas in ACPA-negative patients changes in disease activity correlated well with changes in the Galactosylation of total IgG. Conclusions In ACPA-positive RA, the pregnancy-induced change in Galactosylation of ACPA-IgG, and not that of total IgG, associates with changes in disease activity. These data may indicate that in ACPA-positive patients the Galactosylation of ACPA-IgG is of more pathogenic relevance than that of total IgG.
-
the n glycosylation of mouse immunoglobulin g igg fragment crystallizable differs between igg subclasses and strains
Frontiers in Immunology, 2017Co-Authors: Noortje De Haan, Karli R Reiding, Jasminka Kristic, Agnes Hipgrave L Ederveen, Gordan Lauc, Manfred WuhrerAbstract:N-linked glycosylation of the fragment crystallizable (Fc)-region of immunoglobulin G (IgG) is known to have a large influence on the activity of the antibody, an effect reported to be IgG subclass specific. This situation applies both to humans and mice. The mouse is often used as experimental animal model to study the effects of Fc-glycosylation on IgG effector functions, and results are not uncommonly translated back to the human situation. However, while human IgG Fc-glycosylation has been extensively characterized in both health and disease, this is not the case for mice. To characterize the glycosylation profile of murine IgG-Fc and in addition evaluate the systematic glycosylation differences between mouse strains, sexes, and IgG subclasses, we used nanoliquid chromatography mass spectrometry (nanoLC-MS(/MS)) to look at the subclass-specific IgG Fc-glycopeptides of male and female mice from the strains BALB/c, C57BL/6, CD-1, and Swiss Webster. The structural analysis revealed the presence of predominantly fucosylated, diantennary glycans, with varying amounts of Galactosylation and α2,6-sialylation. In addition, we report glycosylation features not previously reported in an Fc-specific way on murine IgG, including monoantennary, hybrid, and high mannose structures, as well as diantennary structures without a core fucose, with a bisecting N-acetylglucosamine, or with α1,3-Galactosylation. Pronounced differences were detected between strains and the IgG subclasses within each strain. Especially the large spread in Galactosylation and sialylation levels found between both strains and subclasses may vastly influence IgG effector functions. Mouse strain-based and subclass-specific glycosylation differences should be taken into account when designing and interpreting immunological and glycobiological mouse studies involving IgG effector functions.
-
Galactosylation and sialylation levels of igg predict relapse in patients with pr3 anca associated vasculitis
EBioMedicine, 2017Co-Authors: Michael J Kemna, Rosina Plomp, Pieter Van Paassen, Carolien A M Koeleman, Bas C Jansen, Jan Damoiseaux, Jan Willem Cohen Tervaert, Manfred WuhrerAbstract:Abstract Objective The objective of our study is to investigate the Fc glycosylation profiles of both antigen-specific IgG targeted against proteinase 3 (PR3-ANCA) and total IgG as prognostic markers of relapse in patients with Granulomatosis with Polyangiitis (GPA). Methods Seventy-five patients with GPA and a PR3-ANCA rise during follow-up were included, of whom 43 patients relapsed within a median period of 8 (2–16) months. The N-glycan at Asn297 of affinity-purified and denatured total IgG and PR3-ANCA was determined by mass spectrometry of glycopeptides in samples obtained at the time of the PR3-ANCA rise and at the time of the relapse or time-matched during remission. Results Patients with total IgG1 exhibiting low Galactosylation or low sialylation were highly prone to relapse after an ANCA rise (HR 3.46 [95%-CI 1.73–6.96], p In relapsing patients, total IgG1 Galactosylation, sialylation and bisection significantly decreased and fucosylation significantly increased from the time of the PR3-ANCA rise to the relapse (p Conclusion While Fc glycosylation profiles have been associated with clinically manifest autoimmune diseases, in the present study we show that low Galactosylation and sialyation in total IgG1 but not PR3-ANCA IgG1 predicts disease reactivation in patients with GPA who experience an ANCA rise during follow-up. We postulate that glycosylation profiles may be useful in pre-emptive therapy studies using ANCA rises as guideline.
-
skewed fc glycosylation profiles of anti proteinase 3 immunoglobulin g1 autoantibodies from granulomatosis with polyangiitis patients show low levels of bisection Galactosylation and sialylation
Journal of Proteome Research, 2015Co-Authors: Manfred Wuhrer, Carolien A M Koeleman, Maurice H J Selman, Kathrin Stavenhagen, Lorraine Harper, Bart C Jacobs, Caroline O S Savage, Roy Jefferis, Andre M Deelder, Matthew D MorganAbstract:Granulomatosis with polyangiitis (GPA) is associated with circulating immunoglobulin (Ig) G anti-proteinase 3 specific (anti-PR3) anti-neutrophil cytoplasm antibodies (ANCA), which activate cytokine primed neutrophils via Fcgamma receptors. ANCA are class switched IgG antibodies implying T cell help in their production. Glycosylation of IgG Fc, under the control of T cell cytokines, determines the interaction between IgG and its receptors. Previous studies have reported aberrant glycosylation of Ig Fc in GPA patients. We investigated whether aberrant Fc glycosylation was present on anti-PR3 ANCA as well as whole IgG subclass preparations compared to healthy controls and whether this correlated with Birmingham vasculitis activity scores (BVAS), serum cytokines, and time to remission. Here, IgG Fc glycosylation of GPA patients and controls and anti-PR3 ANCA Fc glycosylation were determined by mass spectrometry of glycopeptides. IgG1 and IgG2 subclasses from GPA patients showed reduced Galactosylation, sialylation, and bisection compared to healthy controls. Anti-PR3 IgG1 ANCA Fc Galactosylation, sialylation, and bisection were reduced compared to total IgG1 in GPA. Galactosylation of anti-PR3 ANCA Fc correlated with inflammatory cytokines and time to remission but not BVAS. Bisection of anti-PR3 ANCA Fc correlated with BVAS. Total IgG1 and anti-PR3 IgG1 Fc Galactosylation were weakly correlated, while bisection of IgG1 and anti-PR3 showed no correlation. Our data indicate that aberrant ANCA Galactosylation may be driven in an antigen-specific manner.
-
association between Galactosylation of immunoglobulin g and improvement of rheumatoid arthritis during pregnancy is independent of sialylation
Journal of Proteome Research, 2013Co-Authors: Albert Bondt, Manfred Wuhrer, Maurice H J Selman, Andre M Deelder, Johanna M W Hazes, Sten P Willemsen, Radboud J E M DolhainAbstract:Rheumatoid arthritis (RA) is known to improve during pregnancy and to flare after delivery. Changes in the glycosylation of immunoglobulin G (IgG)'s fragment crystallizable (Fc) have been suggested to play a role herein. Recent animal studies indicate that not Galactosylation but mainly sialylation is important in this respect. We aim to find new associations between IgG-Fc N-glycosylation and improvement of RA during pregnancy and the flare after delivery. Sera of RA patients (n = 251 pregnancies) and healthy controls (n = 32), all participating in a prospective cohort study on RA and pregnancy (PARA study), were collected before conception, during pregnancy, and after delivery. Using a recently developed fast and robust nanoRP-HPLC-sheath-flow-ESI-MS method the glycosylation of IgG Fc-glycopeptides was measured in a subclass specific manner, with relative standard deviations of <4% for the 8 most abundant IgG Fc glycopeptides during the entire measurement period of over 3 weeks. In patients and controls, several glycosylation changes were observed during pregnancy. In depth analysis of the association of these glycosylation changes with disease activity revealed that Galactosylation, independent of sialylation, is associated with improvement of RA during pregnancy. Functional studies in human cell systems should be performed to obtain more insight into this matter.
Makoto Kiso - One of the best experts on this subject based on the ideXlab platform.
-
extended applications of di tert butylsilylene directed α predominant Galactosylation compatible with c2 participating groups toward the assembly of various glycosides
Chemistry: A European Journal, 2006Co-Authors: Akihiro Imamura, Akiyoshi Kimura, Hiromune Ando, Hideharu Ishida, Makoto KisoAbstract:The high versatility of di-tert-butylsilylene(DTBS)-directed α-predominant Galactosylation have been extended to the construction of difficult glycan sequences. First, to investigate the compatibility of the α-predominant reaction with various glycosylation systems a variety of 4,6-O-DTBS-tethered galactosaminyl or galactosyl donors were synthesized efficiently, which have C2-participating groups with a wide variety of leaving groups such as alkylsulfenyl, halide, trichloroacetimidate groups. The results of the detailed examination of the glycosylation reaction using the glycosyl donors showed the wide scope of the 4,6-DTBS-directed α-Galactosylation. In the next step, the stereoselective construction of α-GalN-Ser/Thr sequences was examined by employing the DTBS-directed glycosylation. As a result, various types of serine and threonine derivatives were glycosylated α-selectively, producing α-GalN-Ser/Thr sequences in high yields. Moreover, the DTBS-directed Galactosylation was successfully applied for the synthesis of α-tetrasaccharyl-Ser segment of glycophorin A.
-
a novel synthetic route to α galactosyl ceramides and igb3 using dtbs directed α selective Galactosylation
Synlett, 2006Co-Authors: Akiyoshi Kimura, Akihiro Imamura, Hiromune Ando, Hideharu Ishida, Makoto KisoAbstract:A novel synthetic route to α-galactosyl ceramides and isoglobotrihexosylceramide (iGb3), which can activate NKT cells, was developed by exploiting a di-tert-butylsilylene-directed α-selective Galactosylation procedure.
-
di tert butylsilylene dtbs group directed α selective Galactosylation unaffected by c 2 participating functionalities
ChemInform, 2003Co-Authors: Akihiro Imamura, Hiromune Ando, Hideharu Ishida, Satomi Korogi, Genzoh Tanabe, Osamu Muraoka, Makoto KisoAbstract:We have discovered an unusual α-Galactosylation using phenylthioglycoside of 4,6-O-di-tert-butylsilylene (DTBS)-protected galactose derivatives as a glycosyl donor, which was not hampered by the neighboring participation of C-2 acyl functionality such as NTroc and OBz. The power of the DTBS effect has been exemplified by the coupling reaction with various glycosyl acceptors.
Andre M Deelder - One of the best experts on this subject based on the ideXlab platform.
-
skewed fc glycosylation profiles of anti proteinase 3 immunoglobulin g1 autoantibodies from granulomatosis with polyangiitis patients show low levels of bisection Galactosylation and sialylation
Journal of Proteome Research, 2015Co-Authors: Manfred Wuhrer, Carolien A M Koeleman, Maurice H J Selman, Kathrin Stavenhagen, Lorraine Harper, Bart C Jacobs, Caroline O S Savage, Roy Jefferis, Andre M Deelder, Matthew D MorganAbstract:Granulomatosis with polyangiitis (GPA) is associated with circulating immunoglobulin (Ig) G anti-proteinase 3 specific (anti-PR3) anti-neutrophil cytoplasm antibodies (ANCA), which activate cytokine primed neutrophils via Fcgamma receptors. ANCA are class switched IgG antibodies implying T cell help in their production. Glycosylation of IgG Fc, under the control of T cell cytokines, determines the interaction between IgG and its receptors. Previous studies have reported aberrant glycosylation of Ig Fc in GPA patients. We investigated whether aberrant Fc glycosylation was present on anti-PR3 ANCA as well as whole IgG subclass preparations compared to healthy controls and whether this correlated with Birmingham vasculitis activity scores (BVAS), serum cytokines, and time to remission. Here, IgG Fc glycosylation of GPA patients and controls and anti-PR3 ANCA Fc glycosylation were determined by mass spectrometry of glycopeptides. IgG1 and IgG2 subclasses from GPA patients showed reduced Galactosylation, sialylation, and bisection compared to healthy controls. Anti-PR3 IgG1 ANCA Fc Galactosylation, sialylation, and bisection were reduced compared to total IgG1 in GPA. Galactosylation of anti-PR3 ANCA Fc correlated with inflammatory cytokines and time to remission but not BVAS. Bisection of anti-PR3 ANCA Fc correlated with BVAS. Total IgG1 and anti-PR3 IgG1 Fc Galactosylation were weakly correlated, while bisection of IgG1 and anti-PR3 showed no correlation. Our data indicate that aberrant ANCA Galactosylation may be driven in an antigen-specific manner.
-
association between Galactosylation of immunoglobulin g and improvement of rheumatoid arthritis during pregnancy is independent of sialylation
Journal of Proteome Research, 2013Co-Authors: Albert Bondt, Manfred Wuhrer, Maurice H J Selman, Andre M Deelder, Johanna M W Hazes, Sten P Willemsen, Radboud J E M DolhainAbstract:Rheumatoid arthritis (RA) is known to improve during pregnancy and to flare after delivery. Changes in the glycosylation of immunoglobulin G (IgG)'s fragment crystallizable (Fc) have been suggested to play a role herein. Recent animal studies indicate that not Galactosylation but mainly sialylation is important in this respect. We aim to find new associations between IgG-Fc N-glycosylation and improvement of RA during pregnancy and the flare after delivery. Sera of RA patients (n = 251 pregnancies) and healthy controls (n = 32), all participating in a prospective cohort study on RA and pregnancy (PARA study), were collected before conception, during pregnancy, and after delivery. Using a recently developed fast and robust nanoRP-HPLC-sheath-flow-ESI-MS method the glycosylation of IgG Fc-glycopeptides was measured in a subclass specific manner, with relative standard deviations of <4% for the 8 most abundant IgG Fc glycopeptides during the entire measurement period of over 3 weeks. In patients and controls, several glycosylation changes were observed during pregnancy. In depth analysis of the association of these glycosylation changes with disease activity revealed that Galactosylation, independent of sialylation, is associated with improvement of RA during pregnancy. Functional studies in human cell systems should be performed to obtain more insight into this matter.
-
high throughput igg fc n glycosylation profiling by mass spectrometry of glycopeptides
Journal of Proteome Research, 2013Co-Authors: Maja Pucic Bakovic, Maurice H J Selman, Andre M Deelder, Gordan Lauc, Harry Campbell, Marcus Hoffmann, Igor Rudan, Manfred WuhrerAbstract:Age and sex dependence of subclass specific immunoglobulin G (IgG) Fc N-glycosylation was evaluated for 1709 individuals from two isolated human populations. IgGs were obtained from plasma by affinity purification using 96-well protein G monolithic plates and digested with trypsin. Fc N-glycopeptides were purified and analyzed by negative-mode MALDI-TOF-MS with 4-chloro-α-cyanocinnamic acid (Cl-CCA) matrix. Age-associated glycosylation changes were more pronounced in younger individuals ( 57 years) and in females than in males. Galactosylation and sialylation decreased with increasing age and showed significant sex dependence. Interestingly, the most prominent drop in the levels of galactosylated and sialylated glycoforms in females was observed around the age of 45 to 60 years when females usually enter menopause. The incidence of bisecting N-acetylglucosamine increased in younger individuals and reached a plateau at older age. Furthermore, we compared the results to the total IgG N-glycosylation of the same populations recently analyzed by hydrophilic interaction liquid chromatography (HILIC). Significant differences were observed in the levels of Galactosylation, bisecting N-acetylglucosamine and particularly sialylation, which were shown to be higher in HILIC analysis. Age and sex association of glycosylation features was, to a large extent, comparable between MALDI-TOF-MS and HILIC IgG glycosylation profiling.
-
immunoglobulin g Galactosylation and sialylation are associated with pregnancy induced improvement of rheumatoid arthritis and the postpartum flare results from a large prospective cohort study
Arthritis Research & Therapy, 2009Co-Authors: Fleur E Van De Geijn, Manfred Wuhrer, Maurice H J Selman, Andre M Deelder, Johanna M W Hazes, Sten P Willemsen, Yael A De Man, Radboud J E M DolhainAbstract:Introduction Improvement of rheumatoid arthritis (RA) during pregnancy has been causatively associated with increased Galactosylation of immunoglobulin G (IgG) N-glycans. Since previous studies were small, did not include the postpartum flare and did not study sialylation, these issues were addressed in the present study.
-
glycosylation profiling of immunoglobulin g igg subclasses from human serum
Proteomics, 2007Co-Authors: Manfred Wuhrer, Carolien A M Koeleman, Radboud J E M Dolhain, Jord C Stam, Fleur E Van De Geijn, Theo C Verrips, Cornelis H Hokke, Andre M DeelderAbstract:All four subclasses of human serum IgG contain a single N-glycosylation site in the constant region of their heavy chain, which is occupied by biantennary, largely core-fucosylated and partially truncated oligosaccharides, that may carry a bisecting N-acetylglucosamine and sialic acid residues. IgG glycosylation has been shown to be altered under various physiological and pathological circumstances. IgG N-glycan profiles vary with age, and Galactosylation for example is enhanced during pregnancy. Several diseases including rheumatoid arthritis are associated with a reduction in Galactosylation of the IgG N-glycans. Here, we describe a robust method for the isolation of IgG subclasses using protein A (binds IgG1, IgG2, and IgG4) and protein G (binds additionally IgG3) at the 96-well plate level, which is suitable for automation. Isolated IgGs were digested with trypsin, and obtained glycopeptides were analyzed by nano-LC-MS. Glycopeptides were characterized by CID as well as electron transfer dissociation (ETD). The method provided glycosylation profiles for IgG1, IgG2, IgG3, and IgG4 and revealed distinct differences in N-glycosylation between the four IgG subclasses. The changes in Galactosylation associated with rheumatoid arthritis could readily be monitored. This method is suitable for the subclass-specific analysis of IgG glycosylation from clinical samples.
Radboud J E M Dolhain - One of the best experts on this subject based on the ideXlab platform.
-
acpa igg Galactosylation associates with disease activity in pregnant patients with rheumatoid arthritis
Annals of the Rheumatic Diseases, 2018Co-Authors: Albert Bondt, Manfred Wuhrer, Lise Hafkenscheid, David Falck, Martijn T Kuijper, Yoann Rombouts, Johanna M W Hazes, Radboud J E M DolhainAbstract:Objectives Patients with autoantibody-positive rheumatoid arthritis (RA) are less likely to experience pregnancy-induced improvement of RA disease activity (DAS28-C reactive protein (CRP)) compared with patients with autoantibody-negative RA. Anti-citrullinated protein antibodies (ACPAs) are the most specific autoantibodies for RA. We previously demonstrated that disease improvement is associated with changes in total IgG glycosylation, which regulate antibody effector function. Therefore, we sought to analyse the ACPA-IgG glycosylation profile during pregnancy with the aim to understand the lower change of pregnancy-induced improvement of the disease in patients with autoantibody-positive RA. Methods ACPA-IgGs were purified from ACPA-positive patient sera (n=112) of the Pregnancy-induced Amelioration of Rheumatoid Arthritis cohort, a prospective study designed to investigate pregnancy-associated improvement of RA. The fragment crystallisable (Fc)glycosylation profile of ACPA-IgGs was characterised by mass spectrometry and compared with that of total IgG derived from the same patients or from ACPA-negative patients. Results All ACPA-IgG subclasses display significant changes in the level of Galactosylation and sialylation during pregnancy, although less pronounced than in total IgG. The pregnancy-induced increase in ACPA-IgG Galactosylation, but not sialylation, associates with lower DAS28-CRP. In ACPA-positive patients, no such association was found with changes in the Galactosylation of total IgG, whereas in ACPA-negative patients changes in disease activity correlated well with changes in the Galactosylation of total IgG. Conclusions In ACPA-positive RA, the pregnancy-induced change in Galactosylation of ACPA-IgG, and not that of total IgG, associates with changes in disease activity. These data may indicate that in ACPA-positive patients the Galactosylation of ACPA-IgG is of more pathogenic relevance than that of total IgG.
-
association between Galactosylation of immunoglobulin g and improvement of rheumatoid arthritis during pregnancy is independent of sialylation
Journal of Proteome Research, 2013Co-Authors: Albert Bondt, Manfred Wuhrer, Maurice H J Selman, Andre M Deelder, Johanna M W Hazes, Sten P Willemsen, Radboud J E M DolhainAbstract:Rheumatoid arthritis (RA) is known to improve during pregnancy and to flare after delivery. Changes in the glycosylation of immunoglobulin G (IgG)'s fragment crystallizable (Fc) have been suggested to play a role herein. Recent animal studies indicate that not Galactosylation but mainly sialylation is important in this respect. We aim to find new associations between IgG-Fc N-glycosylation and improvement of RA during pregnancy and the flare after delivery. Sera of RA patients (n = 251 pregnancies) and healthy controls (n = 32), all participating in a prospective cohort study on RA and pregnancy (PARA study), were collected before conception, during pregnancy, and after delivery. Using a recently developed fast and robust nanoRP-HPLC-sheath-flow-ESI-MS method the glycosylation of IgG Fc-glycopeptides was measured in a subclass specific manner, with relative standard deviations of <4% for the 8 most abundant IgG Fc glycopeptides during the entire measurement period of over 3 weeks. In patients and controls, several glycosylation changes were observed during pregnancy. In depth analysis of the association of these glycosylation changes with disease activity revealed that Galactosylation, independent of sialylation, is associated with improvement of RA during pregnancy. Functional studies in human cell systems should be performed to obtain more insight into this matter.
-
immunoglobulin g Galactosylation and sialylation are associated with pregnancy induced improvement of rheumatoid arthritis and the postpartum flare results from a large prospective cohort study
Arthritis Research & Therapy, 2009Co-Authors: Fleur E Van De Geijn, Manfred Wuhrer, Maurice H J Selman, Andre M Deelder, Johanna M W Hazes, Sten P Willemsen, Yael A De Man, Radboud J E M DolhainAbstract:Introduction Improvement of rheumatoid arthritis (RA) during pregnancy has been causatively associated with increased Galactosylation of immunoglobulin G (IgG) N-glycans. Since previous studies were small, did not include the postpartum flare and did not study sialylation, these issues were addressed in the present study.
-
glycosylation profiling of immunoglobulin g igg subclasses from human serum
Proteomics, 2007Co-Authors: Manfred Wuhrer, Carolien A M Koeleman, Radboud J E M Dolhain, Jord C Stam, Fleur E Van De Geijn, Theo C Verrips, Cornelis H Hokke, Andre M DeelderAbstract:All four subclasses of human serum IgG contain a single N-glycosylation site in the constant region of their heavy chain, which is occupied by biantennary, largely core-fucosylated and partially truncated oligosaccharides, that may carry a bisecting N-acetylglucosamine and sialic acid residues. IgG glycosylation has been shown to be altered under various physiological and pathological circumstances. IgG N-glycan profiles vary with age, and Galactosylation for example is enhanced during pregnancy. Several diseases including rheumatoid arthritis are associated with a reduction in Galactosylation of the IgG N-glycans. Here, we describe a robust method for the isolation of IgG subclasses using protein A (binds IgG1, IgG2, and IgG4) and protein G (binds additionally IgG3) at the 96-well plate level, which is suitable for automation. Isolated IgGs were digested with trypsin, and obtained glycopeptides were analyzed by nano-LC-MS. Glycopeptides were characterized by CID as well as electron transfer dissociation (ETD). The method provided glycosylation profiles for IgG1, IgG2, IgG3, and IgG4 and revealed distinct differences in N-glycosylation between the four IgG subclasses. The changes in Galactosylation associated with rheumatoid arthritis could readily be monitored. This method is suitable for the subclass-specific analysis of IgG glycosylation from clinical samples.
Kaoru Takegawa - One of the best experts on this subject based on the ideXlab platform.
-
identification of novel α1 3 galactosyltransferase and elimination of α galactose containing glycans by disruption of multiple α galactosyltransferase genes in schizosaccharomyces pombe
Journal of Biological Chemistry, 2012Co-Authors: Takao Ohashi, Kazuhito Fujiyama, Kaoru TakegawaAbstract:The oligosaccharides from fission yeast Schizosaccharomyces pombe contain large amounts of d-galactose (Gal) in addition to d-mannose (Man), in contrast to the budding yeast Saccharomyces cerevisiae. Detailed structural analysis has revealed that the Gal residues are attached to the N- and O-linked oligosaccharides via α1,2- or α1,3-linkages. Previously we constructed and characterized a septuple α-galactosyltransferase disruptant (7GalTΔ) anticipating a complete lack of α-Gal residues. However, the 7GalTΔ strain still contained oligosaccharides consisting of α1,3-linked Gal residues, indicating the presence of at least one more additional unidentified α1,3-galactosyltransferase. In this study we searched for unidentified putative glycosyltransferases in the S. pombe genome sequence and identified three novel genes, named otg1+–otg3+ (α one, three-galactosyltransferase), that belong to glycosyltransferase gene family 8 in the Carbohydrate Active EnZymes (CAZY) database. Gal-recognizing lectin blotting and HPLC analyses of pyridylaminated oligosaccharides after deletion of these three additional genes from 7GalTΔ strain demonstrated that the resultant disruptant missing 10 α-galactosyltransferase genes, 10GalTΔ, exhibited a complete loss of Galactosylation. In an in vitro Galactosylation assay, the otg2+ gene product had Gal transfer activity toward a pyridylaminated Man9GlcNAc2 oligosaccharide and pyridylaminated Manα1,2-Manα1,2-Man oligosaccharide. In addition, the otg3+ gene product exhibited Gal transfer activity toward the pyridylaminated Man9GlcNAc2 oligosaccharide. Generation of an α1,3-linkage was confirmed by HPLC analysis, α-galactosidase digestion analysis, 1H NMR spectroscopy, and LC-MS/MS analysis. These results indicate that Otg2p and Otg3p are involved in α1,3-Galactosylation of S. pombe oligosaccharides.
-
characterization of two different types of udp glucose galactose 4 epimerase involved in Galactosylation in fission yeast
Microbiology, 2010Co-Authors: Shotaro Suzuki, Tomohiko Matsuzawa, Yayoi Nukigi, Kaoru Takegawa, Naotaka TanakaAbstract:Schizosaccharomyces species are currently the only known organisms with two types of genes encoding UDP-glucose/-galactose 4-epimerase, uge1 + and gal10 +. A strain deleted for uge1+ exhibited a severe Galactosylation defect and a decrease in activity and in UDP-galactose content when grown in glucose-rich medium (2 % glucose), indicating that Uge1p is a major UDP-glucose/-galactose 4-epimerase under these growth conditions. In contrast, gal10 + was efficiently expressed and involved in Galactosylation of cell-surface proteins in low-glucose medium (0.1 % glucose and 2 % glycerol), but not in galactose-containing medium. In a uge1Δgal10Δ strain, the Galactosylation defect was suppressed and UDP-galactose content restored to wild-type levels in galactose-containing medium. Disruption of gal7 +, encoding galactose-1-phosphate uridylyltransferase, in the uge1Δgal10Δ strain reversed suppression of the Galactosylation defect and reduced levels of UDP-galactose, indicating that galactose is transported from the medium to the cytosol and is converted into UDP-galactose via galactose 1-phosphate by Gal7p in Sch. pombe.
-
the schizosaccharomyces pombe gms1 gene encodes an udp galactose transporter homologue required for protein Galactosylation
Biochemical and Biophysical Research Communications, 1997Co-Authors: Mitsuaki Tabuchi, Naotaka Tanaka, Shojiro Iwahara, Kaoru TakegawaAbstract:Abstract In a previous study, we isolated aSchizosaccharomyces pombemutant defective in protein Galactosylation (Takegawa, K., Tanaka, N., Tabuchi, M. and Iwahara, S. (1996)Biosci. Biochem. Biotech.60, 1156–1159). From anS. pombegenomic library, we cloned thegms1+gene which restored the Galactosylation of cell wall glycoproteins. Gms1 protein shares significant sequence similarity with human UDP-galactose and murine CMP-sialic acid transporters. The fission yeast strains deleted for thegms1+gene lacked galactose residues in sell surface glycoproteins and were significantly decreased in UDP-galactose transport activity. These results showed that thegms1+encodes an UDP-galactose transporter, and this protein appears to be an essential role for the incorporation of UDP-galactose into the lumen of Golgi inS. pombe.
