Literature summary for 2.4.1.143 extracted from

Miyazaki, T.; Miyashita, R.; Mori, S.; Kato, T.; Park, E.Y.
Expression and characterization of silkworm Bombyx mori beta-1,2-N-acetylglucosaminyltransferase II, a key enzyme for complex-type N-glycan biosynthesis (2019), J. Biosci. Bioeng., 127, 273-280 .

Search for more literature for 2.4.1.143

Cloned(Commentary)

Cloned (Comment)	Organism
gene GnTII, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic tree, functional expression of the enzyme using a silkworm-based Bombyx mori nucleopolyhedrovirus bacmid expression system, the enzyme is secreted. The recombinant enzyme exhibits similar pH and temperature dependency and the same substrate specificity as human GnTII expressed by the same system, but deglycosylation with peptide:N-glycanase F does not affect its enzymatic activity. Construction of a vector including the N-terminally FLAG (DYKDDDDK)-tagged lumenal region (Leu27eAla498) including the stem domain and the catalytic domain of BmGnTII together with a signal peptide sequence from bombyxin and transformed into Escherichia coli BmDH10Bac-CP-/Chi- competent cells, which contain the cysteine protease- and chitinase-deficient BmNPV bacmid. RT-PCR expression analysis. The cDNA templates are synthesized from total RNAs extracted from whole bodies of first to fifth-instar larvae and pupa and tissues of fifth-instar larvae. Tissues used are as follows: fat body, midgut, silk gland, epidermis, and Malpighian tubule. To produce the soluble form of BmGnTII n hemolymph, chitosan/BmNPV bacmid nanocomplexes are prepared and then injected into fifth-instar silkworm larvae. The bacmid-injected larvae are reared on an artificial diet	Bombyx mori

Cloned (Comment)

Organism

gene GnTII, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic tree, functional expression of the enzyme using a silkworm-based Bombyx mori nucleopolyhedrovirus bacmid expression system, the enzyme is secreted. The recombinant enzyme exhibits similar pH and temperature dependency and the same substrate specificity as human GnTII expressed by the same system, but deglycosylation with peptide:N-glycanase F does not affect its enzymatic activity. Construction of a vector including the N-terminally FLAG (DYKDDDDK)-tagged lumenal region (Leu27eAla498) including the stem domain and the catalytic domain of BmGnTII together with a signal peptide sequence from bombyxin and transformed into Escherichia coli BmDH10Bac-CP-/Chi- competent cells, which contain the cysteine protease- and chitinase-deficient BmNPV bacmid. RT-PCR expression analysis. The cDNA templates are synthesized from total RNAs extracted from whole bodies of first to fifth-instar larvae and pupa and tissues of fifth-instar larvae. Tissues used are as follows: fat body, midgut, silk gland, epidermis, and Malpighian tubule. To produce the soluble form of BmGnTII n hemolymph, chitosan/BmNPV bacmid nanocomplexes are prepared and then injected into fifth-instar silkworm larvae. The bacmid-injected larvae are reared on an artificial diet

Bombyx mori

Protein Variants

Protein Variants	Comment	Organism
additional information	construction of a vector including the N-terminally FLAG (DYKDDDDK)-tagged lumenal region (Leu27eAla498) including the stem domain and the catalytic domain of BmGnTII together with a signal peptide sequence from bombyxin and transformed into Escherichia coli BmDH10Bac-CP--Chi- competent cells, which contain the cysteine protease- and chitinase-deficient BmNPV bacmid. RT-PCR expression analysis. The cDNA templates are synthesized from total RNAs extracted from whole bodies of first to fifth-instar larvae and pupa and tissues of fifth-instar larvae. Tissues used are as follows: fat body, midgut, silk gland, epidermis, and Malpighian tubule. To produce the soluble form of BmGnTII n hemolymph, chitosan/BmNPV bacmid nanocomplexes are prepared and then injected into fifth-instar silkworm larvae. The bacmid-injected larvae are reared on an artificial diet	Bombyx mori

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mn2+	required	Bombyx mori

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
alpha-D-Man-(1->6)-[beta-D-GlcNAc(1->2)-alpha-D-Man(1->3)]-beta-D-Man-(1->4)-beta-D-GlcNAc-(1->4)-D-GlcNAc + UDP-GlcNAc	Bombyx mori	i.e. pyridylaminated MGn glycan, an intermediate during N-glycan processing	?	-	?

Organism

Organism	UniProt	Comment	Textmining
Bombyx mori	A0A451FEC8	-	-

Source Tissue

Source Tissue	Comment	Organism	Textmining
epidermis	-	Bombyx mori	-
fat body	-	Bombyx mori	-
malpighian tubule	-	Bombyx mori	-
midgut	-	Bombyx mori	-
silk gland	-	Bombyx mori	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
alpha-D-Man-(1->6)-[beta-D-GlcNAc(1->2)-alpha-D-Man(1->3)]-beta-D-Man-(1->4)-beta-D-GlcNAc-(1->4)-D-GlcNAc + UDP-GlcNAc	i.e. pyridylaminated MGn glycan, an intermediate during N-glycan processing	Bombyx mori	?	-	?

Synonyms

Synonyms	Comment	Organism
beta-1,2-N-acetylglucosaminyltransferase II	-	Bombyx mori
BmGnTII	-	Bombyx mori
GnTII	-	Bombyx mori
Mgat2	UniProt	Bombyx mori

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Bombyx mori

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
6.5	-	assay at	Bombyx mori

General Information

General Information	Comment	Organism
evolution	compared with the structure of human GnTII, the amino acid residues involved in catalytic activity and substrate recognition are almost fully conserved in Bombyx mori GnTII, which is consistent with its enzymatic properties	Bombyx mori
metabolism	beta-1,2-N-acetylglucosaminyltransferase II is a key enzyme for complex-type N-glycan biosynthesis. Both insect and mammalian cells produce Man(alpha1->6)[GlcNAc(beta1->2)Man(alpha1->3)]Man(beta1->4)GlcNAc(beta1->4)GlcNAc (MGn) glycan as an intermediate during N-glycan processing. In insect cells, beta-N-acetylglucosaminidase (fused lobes, FDL) removes a GlcNAc residue of the alpha1-3 arm of MGn glycan to produce Man(alpha1->6)[Man(alpha1->3)]ManGlcNAc2 (MM), a core structure from paucimannose-type N-glycans	Bombyx mori
physiological function	beta-1,2-N-acetylglucosaminyltransferase II (GnTII) catalyzes the transfer of GlcNAc from a UDP-GlcNAc donor to the alpha1-6 arm of MGn glycan to produce biantennary complex-type glycans in mammalian cells	Bombyx mori