Abstract
Human β-1,4-galactosyltransferase (β-1,4-GalT) V was shown to be involved in the biosynthesis of N-glycans, O-glycans and lactosylceramide (Lac-Cer) by in vitro studies. To determine its substrate specificity, enzymatic activity and its products were analyzed using mouse embryonic fibroblast (MEF) cells from β-1,4-GalT V (B4galt5)-mutant mice. Analysis of expression levels of the β-1,4-GalT I-VI genes revealed that the expression of the β-1,4-GalT V gene in B4galt5 +/- - and B4galt5 -/- -derived MEF cells are a half and null when compared to that of B4galt5 +/+ -derived MEF cells without altering the expression levels of other β-1,4-GalT genes. These MEF cells showed no apparent difference in their growth. When β-1,4-GalT activities were determined towards GlcNAcβ-S-pNP, no significant difference in its specific activity was obtained among B4galt5 +/+ -, B4galt5 +/- - and B4galt5 -/- -derived MEF cells. No significant differences were obtained in structures and amounts of N-glycans and lectin bindings to membrane glycoproteins among B4galt5 +/+ -, B4galt5 +/- - and B4galt5 -/- -derived MEF cells. However, when cell homogenates were incubated with glucosylceramide in the presence of UDP-[3H]Gal, Lac-Cer synthase activity in B4galt5 +/- - and B4galt5 -/- -derived MEF cells decreased to 41% and 11% of that of B4galt5 +/+ -derived MEF cells. Consistent with this, amounts of Lac-Cer and its derivative GM3 in B4galt5 -/- -derived MEF cells decreased remarkably when compared with those of B4galt5 +/+ -derived MEF cells. These results indicate that murine β-1,4-GalT V is involved in Lac-Cer biosynthesis.
| Original language | English |
|---|---|
| Pages (from-to) | 685-695 |
| Number of pages | 11 |
| Journal | Glycoconjugate Journal |
| Volume | 27 |
| Issue number | 7-9 |
| DOIs | |
| Publication status | Published - 2010 Oct |
| Externally published | Yes |
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Keywords
- β-1,4-GalT V
- B4galt5 mice
- Lactosylceramide
- MEF cells
ASJC Scopus subject areas
- Biochemistry
- Cell Biology
- Molecular Biology
Cite this
Involvement of murine β-1,4-galactosyltransferase v in lactosylceramide biosynthesis. / Kumagai, Tadahiro; Sato, Takeshi; Natsuka, Shunji; Kobayashi, Yukito; Zhou, Dapeng; Shinkai, Tadashi; Hayakawa, Satoru; Furukawa, Kiyoshi.
In: Glycoconjugate Journal, Vol. 27, No. 7-9, 10.2010, p. 685-695.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Involvement of murine β-1,4-galactosyltransferase v in lactosylceramide biosynthesis
AU - Kumagai, Tadahiro
AU - Sato, Takeshi
AU - Natsuka, Shunji
AU - Kobayashi, Yukito
AU - Zhou, Dapeng
AU - Shinkai, Tadashi
AU - Hayakawa, Satoru
AU - Furukawa, Kiyoshi
PY - 2010/10
Y1 - 2010/10
N2 - Human β-1,4-galactosyltransferase (β-1,4-GalT) V was shown to be involved in the biosynthesis of N-glycans, O-glycans and lactosylceramide (Lac-Cer) by in vitro studies. To determine its substrate specificity, enzymatic activity and its products were analyzed using mouse embryonic fibroblast (MEF) cells from β-1,4-GalT V (B4galt5)-mutant mice. Analysis of expression levels of the β-1,4-GalT I-VI genes revealed that the expression of the β-1,4-GalT V gene in B4galt5 +/- - and B4galt5 -/- -derived MEF cells are a half and null when compared to that of B4galt5 +/+ -derived MEF cells without altering the expression levels of other β-1,4-GalT genes. These MEF cells showed no apparent difference in their growth. When β-1,4-GalT activities were determined towards GlcNAcβ-S-pNP, no significant difference in its specific activity was obtained among B4galt5 +/+ -, B4galt5 +/- - and B4galt5 -/- -derived MEF cells. No significant differences were obtained in structures and amounts of N-glycans and lectin bindings to membrane glycoproteins among B4galt5 +/+ -, B4galt5 +/- - and B4galt5 -/- -derived MEF cells. However, when cell homogenates were incubated with glucosylceramide in the presence of UDP-[3H]Gal, Lac-Cer synthase activity in B4galt5 +/- - and B4galt5 -/- -derived MEF cells decreased to 41% and 11% of that of B4galt5 +/+ -derived MEF cells. Consistent with this, amounts of Lac-Cer and its derivative GM3 in B4galt5 -/- -derived MEF cells decreased remarkably when compared with those of B4galt5 +/+ -derived MEF cells. These results indicate that murine β-1,4-GalT V is involved in Lac-Cer biosynthesis.
AB - Human β-1,4-galactosyltransferase (β-1,4-GalT) V was shown to be involved in the biosynthesis of N-glycans, O-glycans and lactosylceramide (Lac-Cer) by in vitro studies. To determine its substrate specificity, enzymatic activity and its products were analyzed using mouse embryonic fibroblast (MEF) cells from β-1,4-GalT V (B4galt5)-mutant mice. Analysis of expression levels of the β-1,4-GalT I-VI genes revealed that the expression of the β-1,4-GalT V gene in B4galt5 +/- - and B4galt5 -/- -derived MEF cells are a half and null when compared to that of B4galt5 +/+ -derived MEF cells without altering the expression levels of other β-1,4-GalT genes. These MEF cells showed no apparent difference in their growth. When β-1,4-GalT activities were determined towards GlcNAcβ-S-pNP, no significant difference in its specific activity was obtained among B4galt5 +/+ -, B4galt5 +/- - and B4galt5 -/- -derived MEF cells. No significant differences were obtained in structures and amounts of N-glycans and lectin bindings to membrane glycoproteins among B4galt5 +/+ -, B4galt5 +/- - and B4galt5 -/- -derived MEF cells. However, when cell homogenates were incubated with glucosylceramide in the presence of UDP-[3H]Gal, Lac-Cer synthase activity in B4galt5 +/- - and B4galt5 -/- -derived MEF cells decreased to 41% and 11% of that of B4galt5 +/+ -derived MEF cells. Consistent with this, amounts of Lac-Cer and its derivative GM3 in B4galt5 -/- -derived MEF cells decreased remarkably when compared with those of B4galt5 +/+ -derived MEF cells. These results indicate that murine β-1,4-GalT V is involved in Lac-Cer biosynthesis.
KW - β-1,4-GalT V
KW - B4galt5 mice
KW - Lactosylceramide
KW - MEF cells
UR - http://www.scopus.com/inward/record.url?scp=78651234929&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=78651234929&partnerID=8YFLogxK
U2 - 10.1007/s10719-010-9313-2
DO - 10.1007/s10719-010-9313-2
M3 - Article
VL - 27
SP - 685
EP - 695
JO - Glycoconjugate Journal
JF - Glycoconjugate Journal
SN - 0282-0080
IS - 7-9
ER -