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Christian Korner - One of the best experts on this subject based on the ideXlab platform.
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deficiency of gdp man glcnac2 pp dolichol Mannosyltransferase causes congenital disorder of glycosylation type ik
American Journal of Human Genetics, 2004Co-Authors: Markus Schwarz, Christian Thiel, Ludwig Lehle, Kurt Von Figura, Jurgen Lubbehusen, Bert Dorland, Tom J De Koning, Christian KornerAbstract:The molecular nature of a severe multisystemic disorder with a recurrent nonimmune hydrops fetalis was identified as deficiency of GDP-Man:GlcNAc2-PP-dolichol Mannosyltransferase, the human orthologue of the yeast ALG1 gene (MIM 605907). The disease belongs to the group of congenital disorders of glycosylation (CDG) and is designated as subtype CDG-Ik. In patient-derived serum, the total amount of the glycoprotein transferrin was reduced. Moreover, a partial loss of N-glycan chains was observed, a characteristic feature of CDG type I forms. Metabolic labeling with [6-3H]glucosamine revealed an accumulation of GlcNAc2-PP-dolichol and GlcNAc1-PP-dolichol in skin fibroblasts of the patient. Incubation of fibroblast extracts with [14C]GlcNAc2-PP-dolichol and GDP-mannose indicated a severely reduced activity of the β1,4-Mannosyltransferase, elongating GlcNAc2-PP-dolichol to Man1GlcNAc2-PP-dolichol at the cytosolic side of the endoplasmic reticulum. Genetic analysis of the patient’s hALG1 gene identified a homozygous mutation leading to the exchange of a serine residue to leucine at position 258 in the hALG1 protein. The disease-causing nature of the hALG1 mutation for the glycosylation defect was verified by a retroviral complementation approach in patient-derived primary fibroblasts and was confirmed by the expression of wild-type and mutant hALG1 in the Saccharomyces cerevisiae alg1-1 strain.
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deficiency of dolichyl p man man7glcnac2 pp dolichyl Mannosyltransferase causes congenital disorder of glycosylation type ig
Biochemical Journal, 2002Co-Authors: Christian Thiel, Markus Schwarz, Martin Hasilik, Ulrike Grieben, F Hanefeld, Ludwig Lehle, Kurt Von Figura, Christian KornerAbstract:Deficiency of the endoplasmic reticulum enzyme dolichyl-phosphate mannose (Dol-P-Man):Man 7 GlcNAc 2 -PP-dolichyl Mannosyltransferase leads to a new type of congenital disorder of glycosylation, designated type Ig. The patient I presented with a multisystemic disorder with microcephaly, developmental retardation, convulsions and dysmorphic signs. The isoelectric focusing pattern of the patient's serum transferrin showed the partial loss of complete N-glycan side chains. In skin fibroblasts from the patient, the activity of Dol-P-Man : Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase was severely reduced leading to the accumulation of Man 7 GlcNAc 2 -PP-Dol, which was transferred to newly synthesized glycoproteins. Sequencing of the Dol-P-Man:Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase cDNA revealed a compound heterozygosity for two point mutations, leading to the exchange of leucine 1 5 8 for a proline residue and a premature translation stop with loss of the C-terminal 74 amino acids. The parents were heterozygous for one of the two mutations. Retroviral expression of the wild-type Dol-P-Man: Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase cDNA in patient's fibroblasts normalized the Mannosyltransferase activity.
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carbohydrate deficient glycoprotein syndrome type iv deficiency of dolichyl p man man5 glcnac2 pp dolichyl Mannosyltransferase
The EMBO Journal, 1999Co-Authors: Christian Korner, Ludwig Lehle, Roland Knauer, Ulrich Stephani, Thorsten Marquardt, Kurt Von FiguraAbstract:Type IV of the carbohydrate deficient glycoprotein syndromes (CDGS) is characterized by microcephaly, severe epilepsy, minimal psychomotor development and partial deficiency of sialic acids in serum glycoproteins. Here we show that the molecular defect in the index patient is a missense mutation in the gene encoding the Mannosyltransferase that transfers mannose from dolichyl-phosphate mannose on to the lipid-linked oligosaccharide (LLO) intermediate Man(5)GlcNAc(2)-PP-dolichol. The defect results in the accumulation of the LLO intermediate and, due to its leaky nature, a residual formation of full-length LLOs. N-glycosylation is abnormal because of the transfer of truncated oligosaccharides in addition to that of full-length oligosaccharides and because of the incomplete utilization of N-glycosylation sites. The Mannosyltransferase is the structural and functional orthologue of the Saccharomyces cerevisiae ALG3 gene.
Kurt Von Figura - One of the best experts on this subject based on the ideXlab platform.
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deficiency of gdp man glcnac2 pp dolichol Mannosyltransferase causes congenital disorder of glycosylation type ik
American Journal of Human Genetics, 2004Co-Authors: Markus Schwarz, Christian Thiel, Ludwig Lehle, Kurt Von Figura, Jurgen Lubbehusen, Bert Dorland, Tom J De Koning, Christian KornerAbstract:The molecular nature of a severe multisystemic disorder with a recurrent nonimmune hydrops fetalis was identified as deficiency of GDP-Man:GlcNAc2-PP-dolichol Mannosyltransferase, the human orthologue of the yeast ALG1 gene (MIM 605907). The disease belongs to the group of congenital disorders of glycosylation (CDG) and is designated as subtype CDG-Ik. In patient-derived serum, the total amount of the glycoprotein transferrin was reduced. Moreover, a partial loss of N-glycan chains was observed, a characteristic feature of CDG type I forms. Metabolic labeling with [6-3H]glucosamine revealed an accumulation of GlcNAc2-PP-dolichol and GlcNAc1-PP-dolichol in skin fibroblasts of the patient. Incubation of fibroblast extracts with [14C]GlcNAc2-PP-dolichol and GDP-mannose indicated a severely reduced activity of the β1,4-Mannosyltransferase, elongating GlcNAc2-PP-dolichol to Man1GlcNAc2-PP-dolichol at the cytosolic side of the endoplasmic reticulum. Genetic analysis of the patient’s hALG1 gene identified a homozygous mutation leading to the exchange of a serine residue to leucine at position 258 in the hALG1 protein. The disease-causing nature of the hALG1 mutation for the glycosylation defect was verified by a retroviral complementation approach in patient-derived primary fibroblasts and was confirmed by the expression of wild-type and mutant hALG1 in the Saccharomyces cerevisiae alg1-1 strain.
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deficiency of dolichyl p man man7glcnac2 pp dolichyl Mannosyltransferase causes congenital disorder of glycosylation type ig
Biochemical Journal, 2002Co-Authors: Christian Thiel, Markus Schwarz, Martin Hasilik, Ulrike Grieben, F Hanefeld, Ludwig Lehle, Kurt Von Figura, Christian KornerAbstract:Deficiency of the endoplasmic reticulum enzyme dolichyl-phosphate mannose (Dol-P-Man):Man 7 GlcNAc 2 -PP-dolichyl Mannosyltransferase leads to a new type of congenital disorder of glycosylation, designated type Ig. The patient I presented with a multisystemic disorder with microcephaly, developmental retardation, convulsions and dysmorphic signs. The isoelectric focusing pattern of the patient's serum transferrin showed the partial loss of complete N-glycan side chains. In skin fibroblasts from the patient, the activity of Dol-P-Man : Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase was severely reduced leading to the accumulation of Man 7 GlcNAc 2 -PP-Dol, which was transferred to newly synthesized glycoproteins. Sequencing of the Dol-P-Man:Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase cDNA revealed a compound heterozygosity for two point mutations, leading to the exchange of leucine 1 5 8 for a proline residue and a premature translation stop with loss of the C-terminal 74 amino acids. The parents were heterozygous for one of the two mutations. Retroviral expression of the wild-type Dol-P-Man: Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase cDNA in patient's fibroblasts normalized the Mannosyltransferase activity.
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carbohydrate deficient glycoprotein syndrome type iv deficiency of dolichyl p man man5 glcnac2 pp dolichyl Mannosyltransferase
The EMBO Journal, 1999Co-Authors: Christian Korner, Ludwig Lehle, Roland Knauer, Ulrich Stephani, Thorsten Marquardt, Kurt Von FiguraAbstract:Type IV of the carbohydrate deficient glycoprotein syndromes (CDGS) is characterized by microcephaly, severe epilepsy, minimal psychomotor development and partial deficiency of sialic acids in serum glycoproteins. Here we show that the molecular defect in the index patient is a missense mutation in the gene encoding the Mannosyltransferase that transfers mannose from dolichyl-phosphate mannose on to the lipid-linked oligosaccharide (LLO) intermediate Man(5)GlcNAc(2)-PP-dolichol. The defect results in the accumulation of the LLO intermediate and, due to its leaky nature, a residual formation of full-length LLOs. N-glycosylation is abnormal because of the transfer of truncated oligosaccharides in addition to that of full-length oligosaccharides and because of the incomplete utilization of N-glycosylation sites. The Mannosyltransferase is the structural and functional orthologue of the Saccharomyces cerevisiae ALG3 gene.
Widmar Tanner - One of the best experts on this subject based on the ideXlab platform.
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Protein-O-glycosylation in yeast: protein-specific Mannosyltransferases
Glycobiology, 1997Co-Authors: Martina Gentzsch, Widmar TannerAbstract:S. cerevisiae contains at least six genes (PMT1-6) for dolicholphosphate-D-mannose: protein-O-D-Mannosyltransferases. The in vivo mannosylation of seven O-mannosylated yeast proteins has been analyzed in a number of pmt mutants. The results clearly indicate that the various protein O-Mannosyltransferases have different specificities for protein substrates. Five of the proteins tested (chitinase, a-agglutinin, Kre9p, Bar1p, Pir2p/hsp 150) are mainly underglycosylated in pmt1 and pmt2 mutants, whereby qualitative differences exist among the various proteins. Two of the O-mannosylated proteins (Ggp1p and Kex2p) are not at all affected in pmt1 and pmt2 mutants but are clearly underglycosylated when PMT4 is mutated. Although the PMT4 gene product is shown to be responsible for O-mannosylating a Ser-rich region of Ggp1p in vivo, a penta-seryl-peptide is not an in vitro substrate for this transferase. A PMT3 mutation does affect O-mannosylation of chitinase only in the genetic background of a pmt1pmt2 double mutation, indicating that PMT1 and PMT2 can compensate for a deleted PMT3 gene.
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Protein O-glycosylation in Saccharomyces cerevisiae: the protein O-Mannosyltransferases Pmt1p and Pmt2p function as heterodimer
FEBS Letters, 1995Co-Authors: Martina Gentzsch, Thomas Immervoll, Widmar TannerAbstract:The protein O-Mannosyltransferases Pmt1p and Pmt2p are catalyzing the O-glycosylation of serine and threonine residues in the endoplasmic reticulum of yeast. Deletion of each of these proteins by disruption of the corresponding gene leads to a dramatic decrease of Mannosyltransferase activity in vitro. With an anti-Pmt1p immunoaffinity column a complex of Pmt1p and a second protein was purified; this protein turned out to be Pmt2p. Overexpression of Pmt1p or Pmt2p, respectively, does not increase Mannosyltransferase activity in vitro. Overexpression of both Mannosyltransferases together, however, raises in vitro activity threefold. These data indicate that Pmt1p and Pmt2p function as a complex catalyzing protein O-glycosylation in yeast.
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protein o glycosylation in yeast the pmt2 gene specifies a second protein o Mannosyltransferase that functions in addition to the pmt1 encoded activity
Journal of Biological Chemistry, 1995Co-Authors: Marc Lussier, Martina Gentzsch, Anne Marie Sdicu, Howard Bussey, Widmar TannerAbstract:Abstract The PMT2 gene from Saccharomyces cerevisiae was identified as FUN25, a transcribed open reading frame on the left arm of chromosome I (Ouellette, B. F. F., Clark, M. W. C., Keng, T., Storms, R. G., Zhong, W., Zeng, B., Fortin, N., Delaney, S., Barton, A., Kaback, D. B., and Bussey, H.(1993) Genome 36, 32-42). The product encoded by the PMT2 gene shows significant similarity with the dolichyl phosphate-D-mannose:protein O-D-Mannosyltransferase, Pmt1p (EC 2.4.1.109), which is required for initiating the assembly of O-linked oligosaccharides in S. cerevisiae (Strahl-Bolsinger, S., Immervoll, T., Deutzmann, R., and Tanner, W.(1993) Proc. Natl. Acad. Sci. U. S. A. 90, 8164-8168). The PMT2 gene encodes a new protein O-D-Mannosyltransferase. Yeast cells carrying a PMT2 disruption show a diminished in vitro and in vivo O-mannosylation activity and resemble mutants with a nonfunctional PMT1 gene. Strains bearing a pmt1 pmt2 double disruption show a severe growth defect but retain residual O-mannosylation activity indicating the presence of at least one more protein-O-Mannosyltransferase.
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A new Dol-P-Man:protein O-D-Mannosyltransferase activity from Saccharomyces cerevisiae
Glycobiology, 1995Co-Authors: Martina Gentzsch, Sabine Strahl-bolsinger, Widmar TannerAbstract:The deletion of the protein Mannosyltransferase 1 gene (PMT1) of Saccharomyces cerevisiae results in viable cells. O-Mannosylation of proteins is reduced to about half of the value in comparison to wild-type cells. In order to distinguish between the the PMT1 gene product (= Pmt1p) and residual transferase activity, an in vitro assay to measure Dol-P-Man:protein Mannosyltransferase activity in cells deleted for PMT1 has been developed. The transferase activity of these cells exhibits a pH optimum of 6.5 as compared to pH 7.5 for Pmt1p. The Km value of the residual enzyme activity for the hexapeptide YNPTSV is 7 times higher than that of Pmt1p and shows a clear preference for the seryl residue. Differences in substrate affinities as well as in seryl/threonyl depend on the specific sequence of the peptides used in the enzyme assay. The new enzyme activity shows a significantly lower thermal stability as compared to Pmt1p.
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protein o glycosylation in saccharomyces cerevisiae purification and characterization of the dolichyl phosphate d mannose protein o d Mannosyltransferase
FEBS Journal, 1991Co-Authors: Sabine Strahlbolsinger, Widmar TannerAbstract:The enzyme dolichyl-phosphate-D-mannose: protein O-D-Mannosyltransferase has been solubilized from Saccharomyces cerevisiae membranes and its Mannosyltransferase activity demonstrated using short peptides. The specific activity of the protein was enriched 130-fold before it was further purified by native and SDS gel chromatography. A 92-kDa band correlated well with the enzyme activity; an antibody raised against this protein precipitated the Mannosyltransferase. The 92-kDa band was hydrolysed to 84 kDa after treatment with endoglycosidase F, indicating that the protein is a glycoprotein which may contain four carbohydrate chains. The purified mannosyltrasferase is distinctly influenced in transfer specificity by amino acids next to serine and threonine within the acceptor peptides. Thus acidic amino acids strongly inhibit acceptor activity as do glycine and proline residues as amino-terminal and carboxy-terminal neighbours, respectively.
Vladislav M Panin - One of the best experts on this subject based on the ideXlab platform.
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drosophila dystroglycan is a target of o Mannosyltransferase activity of two protein o Mannosyltransferases rotated abdomen and twisted
Glycobiology, 2010Co-Authors: Naosuke Nakamura, Dmitry Lyalin, Stephanie H Stalnaker, Olga Lavrova, Lance Wells, Vladislav M PaninAbstract:Recent studies highlighted an emerging possibility of using Drosophila as a model system for investigating the mechanisms of human congenital muscular dystrophies, called dystroglycanopathies, resulting from the abnormal glycosylation of α-dystroglycan. Several of these diseases are associated with defects in O-mannosylation, one of the most prominent types of α-dystroglycan glycosylation mediated by two protein O-Mannosyltransferases. Drosophila appears to possess homologs of all essential components of the mammalian dystroglycan-mediated pathway; however, the glycosylation of Drosophila Dystroglycan (DG) has not yet been explored. In this study, we characterized the glycosylation of Drosophila DG using a combination of glycosidase treatments, lectin blots, trypsin digestion, and mass spectrometry analyses. Our results demonstrated that DG extracellular domain is O-mannosylated in vivo. We found that the concurrent in vivo activity of the two Drosophila protein O-Mannosyltransferases, Rotated Abdomen and Twisted, is required for O-mannosylation of DG. While our experiments unambiguously determined some O-mannose sites far outside of the mucin-type domain of DG, they also provided evidence that DG bears a significant amount of O-mannosylation within its central region including the mucin-type domain, and that O-mannose can compete with O-GalNAc glycosylation of DG. We found that Rotated Abdomen and Twisted could potentiate in vivo the dominant-negative effect of DG extracellular domain expression on crossvein development, which suggests that O-mannosylation can modulate the ligand-binding activity of DG. Taken together these results demonstrated that O-mannosylation of Dystroglycan is an evolutionarily ancient mechanism conserved between Drosophila and humans, suggesting that Drosophila can be a suitable model system for studying molecular and genetic mechanisms underlying human dystroglycanopathies.
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the twisted gene encodes drosophila protein o Mannosyltransferase 2 and genetically interacts with the rotated abdomen gene encoding drosophila protein o Mannosyltransferase 1
Genetics, 2006Co-Authors: Dmitry Lyalin, Kate Koles, Sigrid D Roosendaal, Elena Repnikova, Laura Van Wechel, Vladislav M PaninAbstract:The family of mammalian O-Mannosyltransferases includes two enzymes, POMT1 and POMT2, which are thought to be essential for muscle and neural development. Similar to mammalian organisms, Drosophila has two O-Mannosyltransferase genes, rotated abdomen (rt) and DmPOMT2, encoding proteins with high homology to their mammalian counterparts. The previously reported mutant phenotype of the rt gene includes a clockwise rotation of the abdomen and defects in embryonic muscle development. No mutants have been described so far for the DmPOMT2 locus. In this study, we determined that the mutation in the twisted (tw) locus, tw1, corresponds to a DmPOMT2 mutant. The twisted alleles represent a complementation group of recessive mutations that, similar to the rt mutants, exhibit a clockwise abdomen rotation phenotype. Several tw alleles were isolated in the past; however, none of them was molecularly characterized. We used an expression rescue approach to confirm that tw locus represents DmPOMT2 gene. We found that the tw1 allele represents an amino acid substitution within the conserved PMT domain of DmPOMT2 (TW) protein. Immunostaining experiments revealed that the protein products of both rt and tw genes colocalize within Drosophila cells where they reside in the ER subcellular compartment. In situ hybridization analysis showed that both genes have essentially overlapping patterns of expression throughout most of embryogenesis (stages 8–17), while only the rt transcript is present at early embryonic stages (5 and 6), suggesting its maternal origin. Finally, we analyzed the genetic interactions between rt and tw using several mutant alleles, RNAi, and ectopic expression approaches. Our data suggest that the two Drosophila O-Mannosyltransferase genes, rt and tw, have nonredundant functions within the same developmental cascade and that their activities are required simultaneously for possibly the same biochemical process. Our results establish the possibility of using Drosophila as a model system for studying molecular and genetic mechanisms of protein O-mannosylation during development.
Ludwig Lehle - One of the best experts on this subject based on the ideXlab platform.
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deficiency of gdp man glcnac2 pp dolichol Mannosyltransferase causes congenital disorder of glycosylation type ik
American Journal of Human Genetics, 2004Co-Authors: Markus Schwarz, Christian Thiel, Ludwig Lehle, Kurt Von Figura, Jurgen Lubbehusen, Bert Dorland, Tom J De Koning, Christian KornerAbstract:The molecular nature of a severe multisystemic disorder with a recurrent nonimmune hydrops fetalis was identified as deficiency of GDP-Man:GlcNAc2-PP-dolichol Mannosyltransferase, the human orthologue of the yeast ALG1 gene (MIM 605907). The disease belongs to the group of congenital disorders of glycosylation (CDG) and is designated as subtype CDG-Ik. In patient-derived serum, the total amount of the glycoprotein transferrin was reduced. Moreover, a partial loss of N-glycan chains was observed, a characteristic feature of CDG type I forms. Metabolic labeling with [6-3H]glucosamine revealed an accumulation of GlcNAc2-PP-dolichol and GlcNAc1-PP-dolichol in skin fibroblasts of the patient. Incubation of fibroblast extracts with [14C]GlcNAc2-PP-dolichol and GDP-mannose indicated a severely reduced activity of the β1,4-Mannosyltransferase, elongating GlcNAc2-PP-dolichol to Man1GlcNAc2-PP-dolichol at the cytosolic side of the endoplasmic reticulum. Genetic analysis of the patient’s hALG1 gene identified a homozygous mutation leading to the exchange of a serine residue to leucine at position 258 in the hALG1 protein. The disease-causing nature of the hALG1 mutation for the glycosylation defect was verified by a retroviral complementation approach in patient-derived primary fibroblasts and was confirmed by the expression of wild-type and mutant hALG1 in the Saccharomyces cerevisiae alg1-1 strain.
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deficiency of dolichyl p man man7glcnac2 pp dolichyl Mannosyltransferase causes congenital disorder of glycosylation type ig
Biochemical Journal, 2002Co-Authors: Christian Thiel, Markus Schwarz, Martin Hasilik, Ulrike Grieben, F Hanefeld, Ludwig Lehle, Kurt Von Figura, Christian KornerAbstract:Deficiency of the endoplasmic reticulum enzyme dolichyl-phosphate mannose (Dol-P-Man):Man 7 GlcNAc 2 -PP-dolichyl Mannosyltransferase leads to a new type of congenital disorder of glycosylation, designated type Ig. The patient I presented with a multisystemic disorder with microcephaly, developmental retardation, convulsions and dysmorphic signs. The isoelectric focusing pattern of the patient's serum transferrin showed the partial loss of complete N-glycan side chains. In skin fibroblasts from the patient, the activity of Dol-P-Man : Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase was severely reduced leading to the accumulation of Man 7 GlcNAc 2 -PP-Dol, which was transferred to newly synthesized glycoproteins. Sequencing of the Dol-P-Man:Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase cDNA revealed a compound heterozygosity for two point mutations, leading to the exchange of leucine 1 5 8 for a proline residue and a premature translation stop with loss of the C-terminal 74 amino acids. The parents were heterozygous for one of the two mutations. Retroviral expression of the wild-type Dol-P-Man: Man 7 GlcNAc 2 -PP-Dol Mannosyltransferase cDNA in patient's fibroblasts normalized the Mannosyltransferase activity.
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carbohydrate deficient glycoprotein syndrome type iv deficiency of dolichyl p man man5 glcnac2 pp dolichyl Mannosyltransferase
The EMBO Journal, 1999Co-Authors: Christian Korner, Ludwig Lehle, Roland Knauer, Ulrich Stephani, Thorsten Marquardt, Kurt Von FiguraAbstract:Type IV of the carbohydrate deficient glycoprotein syndromes (CDGS) is characterized by microcephaly, severe epilepsy, minimal psychomotor development and partial deficiency of sialic acids in serum glycoproteins. Here we show that the molecular defect in the index patient is a missense mutation in the gene encoding the Mannosyltransferase that transfers mannose from dolichyl-phosphate mannose on to the lipid-linked oligosaccharide (LLO) intermediate Man(5)GlcNAc(2)-PP-dolichol. The defect results in the accumulation of the LLO intermediate and, due to its leaky nature, a residual formation of full-length LLOs. N-glycosylation is abnormal because of the transfer of truncated oligosaccharides in addition to that of full-length oligosaccharides and because of the incomplete utilization of N-glycosylation sites. The Mannosyltransferase is the structural and functional orthologue of the Saccharomyces cerevisiae ALG3 gene.