Information on EC 2.4.1.232 - initiation-specific alpha-1,6-mannosyltransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.4.1.232
-
RECOMMENDED NAME
GeneOntology No.
initiation-specific alpha-1,6-mannosyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
Transfers an alpha-D-mannosyl residue from GDP-mannose into lipid-linked oligosaccharide, forming an alpha-(1->6)-D-mannosyl-D-mannose linkage
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycosyl group transfer
-
hexosyl group transfer
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Various types of N-glycan biosynthesis
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Metabolic pathways
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-
SYSTEMATIC NAME
IUBMB Comments
GDP-mannose:oligosaccharide 6-alpha-D-mannosyltransferase
Requires Mn2+. In Saccharomyces cerevisiae, this enzyme catalyses an essential step in the outer chain elongation of N-linked oligosaccharides. Man8GlcNAc and Man9GlcNAc are equally good substrates.
CAS REGISTRY NUMBER
COMMENTARY hide
135622-87-6
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346003-17-6
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346003-17-6
Schizosaccharomyces pombe gene och1+
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain J-1012 and B-792
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-
Manually annotated by BRENDA team
NIH B-792 serotype B strain
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-
Manually annotated by BRENDA team
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SwissProt
Manually annotated by BRENDA team
gene ALG12
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-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
4-methylumbelliferyl-alpha-D-Man + GDP-mannose
4-methylumbelliferyl-alpha-D-Man-(1->6)-D-Man + GDP
show the reaction diagram
alpha-D-mannobiose + GDP-mannose
alpha-D-mannotriose + GDP
show the reaction diagram
alpha-D-mannose + Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine
?
show the reaction diagram
-
recombinant enzyme shows 28.8% of the activity with Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GLcNAcbeta(1,4)GlcNAc-pyridylamine
-
-
?
GDP-mannose + alpha-1,6-D-mannobiose
?
show the reaction diagram
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two distinct Mnn9p-containing multi-protein complexes add multiple mannosyl residues forming a series of poly-mannose structures in vitro, no substrate of Och1p
formation of a series of poly-mannose structures, which contain mainly alpha-1,6- and some additional alpha-1,2-linked mannoses
?
GDP-mannose + Man8GlcNAc
GDP + Man9GlcNAc
show the reaction diagram
-
Man8GlcNAc from thyroglobulin Man9GlcNAc by treatment with yeast specific mannosidase, specificity, enzyme catalyzes addition of mannose to the alpha-1,3-mannose of the substrate, initiates outer chain formation
-
?
GDP-mannose + Man8GlcNAc2-PA
GDP + Man9GlcNAc2-PA
show the reaction diagram
GDP-mannose + Man9GlcNAc
GDP + Man10GlcNAc
show the reaction diagram
-
Man9GlcNAc from thyroglobulin, specificity, enzyme catalyzes addition of mannose to the alpha-1,3-mannose of the substrate, initiates outer chain formation
-
?
GDP-mannose + Man9GlcNAc2-PA
GDP + Man10GlcNAc2-PA
show the reaction diagram
GDP-mannose + Manalpha(1,2)Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine
?
show the reaction diagram
-
recombinant enzyme shows 74.2% of the activity with Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GLcNAcbeta(1,4)GlcNAc-pyridylamine. Och1p has the catalytic potential to transfer two molecules of mannose to Manalpha(1,2)Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine. The second mannose is attached with alpha1,6-linkage to either the alpha1,6-linked or the alpha1,3-linked mannose that is attached to alpha1,6-linked mannose
-
-
?
GDP-mannose + Manalpha(1,2)Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine
?
show the reaction diagram
-
recombinant enzyme shows 58.5% of the activity with Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GLcNAcbeta(1,4)GlcNAc-pyridylamine
-
-
?
GDP-mannose + Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine
?
show the reaction diagram
-
-
-
-
?
GDP-mannose + Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine
?
show the reaction diagram
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recombinant enzyme shows 66.1% of the activity with Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GLcNAcbeta(1,4)GlcNAc-pyridylamine
-
-
?
GDP-mannose + Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine
?
show the reaction diagram
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recombinant enzyme shows 28.8% of the activity with Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GLcNAcbeta(1,4)GlcNAc-pyridylamine
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-
?
GDP-mannose + Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine
?
show the reaction diagram
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recombinant enzyme shows 14.4% of the activity with Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GLcNAcbeta(1,4)GlcNAc-pyridylamine
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-
?
GDP-mannose + Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GlcNAcbeta(1,4)GlcNAc-pyridylamine
?
show the reaction diagram
-
recombinant enzyme shows 14.4% of the activity with Manalpha(1,2)Manalpha(1,6)[Manalpha(1,3)]Manalpha(1,6)[Manalpha(1,2)Manalpha(1,2)Manalpha(1,3)]Manbeta(1,4)GLcNAcbeta(1,4)GlcNAc-pyridylamine
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-
?
GDP-mannose + Manalpha(1-2)Manalpha(1-6)[Manalpha(1-2)Manalpha(1-3)]Manalpha(1-6)[Manalpha(1-2)Manalpha(1-2)Manalpha(1-3)]Manbeta(1-4)GlcNAc
GDP + Manalpha(1-2)[Manalpha(1-6)]Manalpha(1-6)[Manalpha(1-2)Manalpha(1-3)]Manalpha(1-6)[Manalpha(1-2)Manalpha(1-2)Manalpha(1-3)]Manbeta(1-4)GlcNAc
show the reaction diagram
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Man9GlcNAc from rat liver glycoproteins and thyroglobulin, enzyme initiates outer chain synthesis, the mannose residue added is alpha-1,6-linked to the alpha-1,6-mannose residue of the substrate, removal of the alpha-1,2-linked mannose residue from Man9GlcNAc is not essential for enzyme activity
Man10GlcNAc
?
GDP-mannose + Manalpha(1-2)Manalpha(1-6)[Manalpha(1-3)]Manalpha(1-6)[Manalpha(1-2)Manalpha(1-2)Manalpha(1-3)]Manbeta(1-4)GlcNAc
GDP + Manalpha(1-2)[Manalpha(1-6)]Manalpha(1-6)[Manalpha(1-3)]Manalpha(1-6)[Manalpha(1-2)Manalpha(1-2)Manalpha(1-3)]Manbeta(1-4)GlcNAc
show the reaction diagram
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Man8GlcNAc from rat liver glycoproteins and thyroglobulin, enzyme initiates outer chain synthesis, the mannose residue added is alpha-1,6-linked to the alpha-1,6-mannose residue of the substrate
Man9GlcNAc
?
GDP-mannose + Manalpha(1-3)Manalpha(1-2)Manalpha(1-3)Manalpha(1-2)Man
GDP + Manalpha(1-3)[Manalpha(1-6)]Manalpha(1-2)Manalpha(1-2)Manalpha(1-2)Man
show the reaction diagram
GDP-mannose + oligosaccharide acceptor
?
show the reaction diagram
-
Mnn9p is involved in protein mannosylation, incorporates alpha-1,6-linked mannose residues, the elongation of the N-glycan outer chain, initiated by Och1p, is continued by two mannosyltransferase complexes: V-complex composed of Mnn9p and Van1p and A-complex composed of Mnn9p, Anp1p, Hoc1p, Mnn10p and Mnn11p
-
?
GDP-mannose + oligosaccharide acceptor
GDP + alpha-mannosyl-1,6-oligosaccharide
show the reaction diagram
GDP-mannose + pyridylaminated Man5GlcNAc2
GDP + pyridylaminated Man6GlcNAc2
show the reaction diagram
incorporates alpha-1,6-linked mannose, poor substrate
-
?
GDP-mannose + pyridylaminated Man6GlcNAc2
GDP + pyridylaminated Man7GlcNAc2
show the reaction diagram
incorporates alpha-1,6-linked mannose
-
?
GDP-mannose + pyridylaminated Man7GlcNAc2
GDP + pyridylaminated Man8GlcNAc2
show the reaction diagram
incorporates alpha-1,6-linked mannose
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?
GDP-mannose + pyridylaminated Man8GlcNAc2
GDP + pyridylaminated Man9GlcNAc2
show the reaction diagram
incorporates alpha-1,6-linked mannose
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?
GDP-mannose + pyridylaminated Man9GlcNAc2
GDP + pyridylaminated Man10GlcNAc2
show the reaction diagram
incorporates alpha-1,6-linked mannose
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?
GDP-mannose + pyridylaminated oligosaccharide acceptor
?
show the reaction diagram
GDP-mannose + pyridylaminated oligosaccharide acceptor
GDP + pyridylaminated alpha-mannosyl-1,6-oligosaccharide
show the reaction diagram
incorporates alpha-1,6-linked mannose, substrate specificity
-
?
GDP-mannose + pyridylamino-Manalpha1-3Man-alpha1-2Manalpha1-2Manalpha1-2Man
?
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
GDP-mannose + oligosaccharide acceptor
?
show the reaction diagram
-
Mnn9p is involved in protein mannosylation, incorporates alpha-1,6-linked mannose residues, the elongation of the N-glycan outer chain, initiated by Och1p, is continued by two mannosyltransferase complexes: V-complex composed of Mnn9p and Van1p and A-complex composed of Mnn9p, Anp1p, Hoc1p, Mnn10p and Mnn11p
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?
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
Ni2+, Mg2+, Ca2+, and Zn2+ cannot substitute for Mn2+
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Triton X-100
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requirement, maximum activity at 0.5-2%
additional information
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och1+ gene expression is induced by NaCl and KCl through the transcription factor Atf1p
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.35
Manalpha(1-2)Manalpha(1-6)[Manalpha(1-2)Manalpha(1-3)]Manalpha(1-6)[Manalpha(1-2)Manalpha(1-2)Manalpha(1-3)]Manbeta(1-4)GlcNAc
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-
0.39
Manalpha(1-2)Manalpha(1-6)[Manalpha(1-3)]Manalpha(1-6)[Manalpha(1-2)Manalpha(1-2)Manalpha(1-3)]Manbeta(1-4)GlcNAc
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-
additional information
additional information
steady-state kinetics of wild-type ScMnn9 enzyme
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
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activity assay
7.1 - 7.6
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in Tris maleate buffer
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2 - 7
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relative activity at pH 2 28%, at pH 7 100%, strain J-1012 alpha-1,6-MT
6.5 - 8.5
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the recombinant Och1p in Tris/malate buffer exhibits above 80% of maximum activity between pH 6.5 and pH 8.5
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
27 - 37
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strain J-1012 alpha-1,6-MT
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
sequence of Anl1p contains a putative transmembrane domain, enzyme interacts with YlMnn9p
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant wild-type and mutant enzymes, sitting drop vapour diffusion, mixing of 0.0005 ml of protein solution with 0.0005 ml of preciputation solution containing 0.1 M HEPES, pH 7.5, and 2 M ammonium sulfate, crystals are soaked in 50% Na-malonate, pH 7.5, containing mersalyl acid for 16 h at 20C, crystals of ScMnn9-D236N mutant are transferred to 50% Na-malonate, pH 7.5, and soaked with 3 mM GDP and 10 mM MnCl2 for 10 min at 20C, X-ray diffraction structure determination and analysis at 2.0-2.2 A resolution, modeling
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
particulate enzyme fractions are obtained by centrifugation of Candida albicans cell homogenates
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recombinant enzyme
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recombinant His6-tagged wild-type and mutant enzymes from Escherichia coli BL21(DE3) pLysS by nickel affinity chromatography
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ANL1 and OCH1 gene
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as a fusion with the with green fluorescent protein, MNN9-OCH1 chimeric genes are constructed and cloned; as a fusion with the with red fluorescent protein, MNN9-OCH1 chimeric genes are constructed and cloned
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into the pGEMT-Easy vector
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MNN9 gene encoding Mnn9p is cloned and sequenced
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OCH1 gene encodes Och1p, an alpha-1,6-mannosyltransferase
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och1+ gene encodes alpha-1,6-mannosyltransferase, recombinant expression and cloning, och1+ gene knockout fission yeast
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och1+ gene, sequencing
recombinant expression of His6-tagged wild-type and mutant enzymes in Escherichia coli BL21(DE3) pLysS, expression of mutant enzymes in Saccharomyces cerevisiae strain BY4741 DELTAMNN9
recombinant Och1p is expressed in Pichia pastoris as a secreted protein, after truncation of its transmembrane region and fusion with myc and histidine tags at the C-terminus
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transcription of the OCH1 gene encoding alpha-1,6-mannosyltransferase is regulated by 2 transcription factors, Swi4 and Skn7, it is controlled by the ubiquitin-dependent degradation pathway
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transcriptional control of the och1+ gene, gene expression is not regulated during the cell cycle, but is induced by NaCl and KCl through the transcription factor Atf1p
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D236N
site-directed mutagenesis, D236 is the first aspartic acid in the canonical GT-A DxD catalytic motif, and mutation to the isosteric asparagine results in the loss of activity
D280N
site-directed mutagenesis, inactive mutant
H389A
site-directed mutagenesis, inactive mutant
M16/O16
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chimeric Mnn9p-Och1p protein, a peptide from the first to the 16th amino acids of Mnn9p is fused with a polypeptide from the 16th to the 480th amino acids of Och1p
M33/O37
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chimeric Mnn9p-Och1p protein, a peptide from the first to the 33th amino acids of Mnn9p is fused with a polypeptide from the 37th to the 480th amino acids of Och1p
M93/O80
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chimeric Mnn9p-Och1p protein, a peptide from the first to the 93th amino acids of Mnn9p is fused with a polypeptide from the 80th to the 480th amino acids of Och1p
R209A
site-directed mutagenesis, inactive mutant
D236N
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site-directed mutagenesis, D236 is the first aspartic acid in the canonical GT-A DxD catalytic motif, and mutation to the isosteric asparagine results in the loss of activity
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D280N
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site-directed mutagenesis, inactive mutant
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H389A
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site-directed mutagenesis, inactive mutant
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R209A
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site-directed mutagenesis, inactive mutant
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W266stop
inactive mutant strain 7X4 with a nonsense mutation resulting in a truncated Och1p, temperature- and hygromycin B-sensitive, no synthesis of outer chains on acid phosphatase
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
drug development
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along with the possibility of a lack of the immunogenic terminal alpha-1,3-mannose linkages in Yarrowia lipolytica, a Yloch1delta mutant strain can be exploited as a platform strain for developing another potential yeast system that can produce recombinant glycoproteins with human-compatible oligosaccharides