Molecular cloning of a human cDNA encoding β-1,4-galactosyltransferase with 37% identity to mammalian UDP-Gal:GlcNAc β-1,4-galactosyltransferase

• DOI: 10.1073/pnas.95.2.472
• Publication Date: 2002-07-26T14:40:11Z
• Authors: Takeshi Sato; Kiyoshi Furukawa; Hans Bakker; Dirk H. Van den E...; Irma Van Die
• Keywords: Galactosyltransferase; Northern blot; BETA (programming language)

A cDNA encoding a β-1,4-galactosyltransferase named β-1,4-GalT II was cloned from library of the human breast tumor cell line, MRK-nu-1. Initially, 860-bp PCR fragment obtained MRK-nu-1 mRNA by 3′-rapid amplification ends using two nested degenerate oligonucleotide primers based on highly conserved amino acid sequence found in catalytic domain mammalian β-1,4-galactosyltransferases and Lymnaea stagnalis β-1,4- N -acetylglucosaminyltransferase (β-1,4-GlcNAcT), both which utilize same sugar acceptor. This subsequently used as probe to isolate 4.7-kb that contained an ORF 1,164 bp predicting polypeptide 388 aa. Its deduced shows identity 37% with previously characterized (referred I) 28% L. β-1,4-GlcNAcT. Study properties fused protein expressed soluble form COS-7 cells revealed it is genuine but has no lactose synthetase activity presence α-lactalbumin. Northern blot analysis 24 tissues showed they all express transcript, although levels varied. These results indicate contain another β-1,4-GalT.