Beta-1, 4 - Galactose-based Transfer Cloning And Expression Of The Gene

β-1,4-Galactosyltransferase-I,β-1 4-Gal-Tl, catalyzes the transfer of galactose (Gal) from UDP-Gal to the N-acetylglucosamine (GlcNAc) residue present at the non-reducing terminal end of glycans of glycoproteins and glycolipids, producing (3-1,4 linked galactosylated glycan. hi addition to GlcNAc as an acceptor, the enzyme can also use other sugars, such as N-acyl substituted glucosamines and N-acetyl-D-mannosamine, as acceptors.Although the genes for all β-1,4-Galactosyltransferase family are located at different chromosomal loci, the evolutionary relationship is indicated by a 30-55% amino acid identity. Moreover, they have hi common the localization in the Golgi apparatus, the topology of type II membrane proteins, and the transfer of galactose from UDP-galactose onto the C4-hydroxyl group of GlcNAc residues.The β-1,4-GT is the most extensively studied enzyme among many kinds of glycosyltransferases. β-1,4-GT seems to play a multifunctional role hi normal cell physiology and has been associated with Sperm-egg binding, cell-cell recognition, cell migration on basal lamina and neurite extension, embryonic maturation, rheumatoid arthritis, and cell development.hi this paper,it is studied on expression of theβ-1,4-GT gene. Theβ-1,4-GT gene was got from the plasmid pMGT-239/2615 through PCR techniques, and cloned into the pGEMT vector. Its sequence was determined by DNA sequencing system. It was then subcloned into the pGEX-4T-2 expression vector, and expressed hi E.coli BL(DE3) after transformation and IPTG induction. The constructed plasmid containing GST-GT fused gene gave a high level of expression of the fused gene. Western blot analysis indicated that the fusion protein could specifically bind to anti-GST antibody. The SDS-PAGE analysis showed that the weight of the fusion protein is correct.