Expression cloning of mouse cDNA of CMP-NeuAc:lactosylceramide α2,3- sialyltransferase, an enzyme that initiates the synthesis of gangliosides

Satoshi Fukumoto, Hiroshi Miyazaki, George Goto, Takeshi Urano, Keiko Furukawa, Koichi Furukawa

Research output: Contribution to journalArticle

60 Citations (Scopus)

Abstract

Expression cloning of a cDNA for the α2,3-sialyltransferase (GM3 synthase) (EC 2.4.99.-) gene was performed using a GM3-lacking mouse fibroblast line L cell and anti-GM3 monoclonal antibody. Plasmids from a cDNA library generated with poly(A)+ RNA of a mouse fibrosarcoma line CMS5j and pdl3027 (polyoma T antigen) were co-transfected into L cells. The isolated cDNA clone pM3T-7 predicted a type II membrane protein with 13 amino acids of cytoplasmic domain, 17 amino acids of transmembrane region, and a large catalytic domain with 329 amino acids. Introduction of the cDNA clone into L cells resulted in the neo-synthesis of GM3 and high activity of α2,3- sialyltransferase. Among glycosphingolipids, only lactosylceramide showed significant activity as an acceptor, indicating that this gene product is a sialyltransferase specific for the synthesis of GM3. An amino acid sequence deduced from the cloned cDNA showed the typical sialyl motif with common features among α2,3-sialyltransferases. Among various mouse tissues, brain, liver, and testis showed relatively high expression of a 2.3-kilobase mRNA, whereas all tissues, more or less, expressed this gene.

Original languageEnglish
Pages (from-to)9271-9276
Number of pages6
JournalJournal of Biological Chemistry
Volume274
Issue number14
DOIs
Publication statusPublished - Apr 2 1999
Externally publishedYes

Fingerprint

Sialyltransferases
Cytidine Monophosphate
Gangliosides
Cloning
Organism Cloning
Complementary DNA
Amino Acids
Genes
Enzymes
Clone Cells
Tissue
Glycosphingolipids
Messenger RNA
Fibrosarcoma
Viral Tumor Antigens
Fibroblasts
Gene Library
Liver
Testis
Amino Acid Sequence

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Expression cloning of mouse cDNA of CMP-NeuAc:lactosylceramide α2,3- sialyltransferase, an enzyme that initiates the synthesis of gangliosides. / Fukumoto, Satoshi; Miyazaki, Hiroshi; Goto, George; Urano, Takeshi; Furukawa, Keiko; Furukawa, Koichi.

In: Journal of Biological Chemistry, Vol. 274, No. 14, 02.04.1999, p. 9271-9276.

Research output: Contribution to journalArticle

Fukumoto, Satoshi ; Miyazaki, Hiroshi ; Goto, George ; Urano, Takeshi ; Furukawa, Keiko ; Furukawa, Koichi. / Expression cloning of mouse cDNA of CMP-NeuAc:lactosylceramide α2,3- sialyltransferase, an enzyme that initiates the synthesis of gangliosides. In: Journal of Biological Chemistry. 1999 ; Vol. 274, No. 14. pp. 9271-9276.
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