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Alpha 1,4-galactosyltransferase and dna encoding thereofAlpha 1,4-galactosyltransferase and dna encoding thereof description/claimsThe Patent Description & Claims data below is from USPTO Patent Application 20090269802, Alpha 1,4-galactosyltransferase and dna encoding thereof. Brief Patent Description - Full Patent Description - Patent Application Claims
1. Field of the Invention The present invention relates to the production of a polypeptide using a recombinant DNA, or to a tool useful for diagnosis or treatment of diseases, or more specifically to α1,4-galactosyltransferase and DNA encoding thereof, a recombination vector containing the DNA, and a transformed cell transfected with the DNA or by the recombination vector, and to a method for producing Gb3/CD77 or globo-series glycolipids by using the transformed cell. 2. Description of the Related Art Glycosphingolipids are amphipathic molecules(ref.1) that are synthesized by sequential actions of glycosyltransferases(ref.2). Addition of one of three different sugars onto lactosylceramide (which may be termed “LacCer” hereinafter) results in the synthesis of either one of three major glycolipid series, i.e., ganglioside-series (a 2,3-sialic acid), lacto/neolacto-series (β1,4-N-acetylglucosamine) and globo-series (α1,4-galactose). Although a number of genes coding for enzymes responsible for the synthesis of the carbohydrate moiety of glycosphingolipids have been recently isolated(ref.3), no glycosyltransferase genes specific for the synthesis of globo-series glycolipids have been isolated to date. Globotriaosylceramide (hereinafter sometimes referred to as “Gb3”) is synthesized by α1,4-galactosyltransferase (α1,4 Gal-T) from LacCer(ref.4). This glycolipid has been characterized on red blood cells as the Pk antigen of the P blood group system(ref.5). Since Wiels et al(ref.6) reported that Gb3 was a Burkitt\'s lymphoma associated antigen, the expression and biological significance of Gb3 have been vigorously studied(ref.7, 8 and 9). Since Gb3 was clustered as CD77, this antigen will be referred to as Gb3/CD77. Gb3/CD77 was reported to be expressed in high amounts on Burkitt\'s lymphoma cells. However, it is now considered to be a differentiation antigen expressed on B cells, and can also be found in some malignant tumors of B cell lineage(ref.7). Among normal leukocytes, it is only expressed on a subset of tonsillar B cells in the germinal centers (GC)(ref.9). Interestingly, GC B lymphocytes expressing Gb3/CD77 undergo rapid and spontaneous apoptosis when isolated and cultured in vitro(ref.11). Furthermore, Burkitt\'s lymphoma cells with Gb3/CD77 antigen were also easily induced to enter apoptosis upon culture at low serum concentration or cross-linking by anti-immunoglobulin M antibodies(ref.12). Gb3/CD77 has been recognized as a receptor for verotoxins (VTs), the Shiga-like toxin from E. coli 0157 strain that can trigger serious cytotoxic effects(ref.13 and 14). VT B-subunit specifically binds to Gb3/CD77, then A subunit is incorporated into cells, resulting in the degradation of 28 S ribosomal RNA and cell death(ref.15). However, only B-subunit is also able to induce apoptosis when cross-linked(ref.16). These results indicate that Gb3/CD77 is a critical molecule in mediating apoptosis signals, although the precise mechanisms remain to be investigated. As stated above, it was revealed that Gb3/CD77, a globo-series glycolipid, is synthesized by addition of α1,4-galactose to LacCer(ref.4), but the glycosyltransferase specific for this synethetic reaction has not been isolated yet. An object of the present invention is to isolate a Gb3/CD77 synthase, that is, a 1,4-galactosyltransferase, and DNA encoding thereof, and to provide the use thereof. The present inventors have studied hard to achieve the above objects, and succeeded in isolating DNA encoding α1,4-galactosyltransferase, revealing its nucleotide sequence, and confirming that the DNA is responsible for the expression of active α1,4-galactosyltransferase. Thus, they achieved the present invention. The present invention provides a polypeptide of (a) or (b) below (hereinafter sometimes referred to as “the polypeptide of the present invention”): (a) a polypeptide consisting of an amino acid sequence represented by the amino acid Nos. 46-353 in SEQ ID NO: 2; or (b) a polypeptide which comprises an amino acid sequence including substitution, deletion, insertion or transposition of one or few amino acids in the amino acid sequence of (a) and which has an enzymatic activity to transfer a galactose residue from a galactose donor to C4 position of galactose residue of lactosylceramide or galactosylceramide which serves as an acceptor. The polypeptide of the present invention also includes a polypeptide of (a′) or (b′) below: (a′) a polypeptide consisting of an amino acid sequence represented by the amino acid Nos. 20-353 in SEQ ID NO:2; or (b′) a polypeptide which comprises an amino acid sequence including substitution, deletion, insertion or transposition of one or few amino acids in the amino acid sequence of (a′) and which has an enzymatic activity to transfer a galactose residue from a galactose donor to C4 position of galactose residue of lactosylceramide or galactosylceramide which serves as an acceptor. The polypeptide of the present invention also includes a polypeptide of (a″) or (b″) below: (a″) a polypeptide consisting of an amino acid sequence represented by SEQ ID NO:2; or (b″) a polypeptide which comprises an amino acid sequence including substitution, deletion, insertion or transposition of one or few amino acids in the amino acid sequence of (a″) and which has an enzymatic activity to transfer a galactose residue from a galactose donor to C4 position of galactose residue of lactosylceramide or galactosylceramide which serves as an acceptor. The present invention also provides a DNA encoding the polypeptides according to any one of above polypeptides (hereinafter sometimes referred to as “the DNA of the present invention”). The DNA of the present invention includes a DNA represented by (a) or (b) below:
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