Traditionally, glycogen synthase (GS) has been considered to catalyze the key step of glycogen synthesis and to exercise most of the control over this metabolic pathway. However, recent advances have shown that other factors must be considered. Moreover, the control of glycogen deposition does not follow identical mechanisms in muscle and liver. Glucose must be phosphorylated to promote activation of GS. Glucose-6-phosphate (Glc-6-P) binds to GS, causing the allosteric activation of the enzyme probably through a conformational rearrangement that simultaneously converts it into a better substrate for protein phosphatases, which can then lead to the covalent activation of GS. The potency of Glc-6-P for activation of liver GS is determined by its source, since Glc-6-P arising from the catalytic action of glucokinase (GK) is much more effective in mediating the activation of the enzyme than the same metabolite produced by hexokinase I (HK I). As a result, hepatic glycogen deposition from glucose is subject to a system of control in which the 'controller', GS, is in turn controlled by GK. In contrast, in skeletal muscle, the control of glycogen synthesis is shared between glucose transport and GS. The characteristics of the two pairs of isoenzymes, liver GS/GK and muscle GS/HK I, and the relationships that they establish are tailored to suit specific metabolic roles of the tissues in which they are expressed. The key enzymes in glycogen metabolism change their intracellular localization in response to glucose. The changes in the intracellular distribution of liver GS and GK triggered by glucose correlate with stimulation of glycogen synthesis. The translocation of GS, which constitutes an additional mechanism of control, causes the orderly deposition of hepatic glycogen and probably represents a functional advantage in the metabolism of the polysaccharide.

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The ABO system is one of the most important blood group systems in transfusion/transplantation medicine. However, the evolutionary significance of the ABO gene and its polymorphism remained unknown. We took an integrative approach to gain insights into the significance of the evolutionary process of ABO genes, including those related not only phylogenetically but also functionally. We experimentally created a code table correlating amino acid sequence motifs of the ABO gene-encoded glycosyltransferases with GalNAc (A)/galactose (B) specificity, and assigned A/B specificity to individual ABO genes from various species thus going beyond the simple sequence comparison. Together with genome information and phylogenetic analyses, this assignment revealed early appearance of A and B gene sequences in evolution and potentially non-allelic presence of both gene sequences in some animal species. We argue: Evolution may have suppressed the establishment of two independent, functional A and B genes in most vertebrates and promoted A/B conversion through amino acid substitutions and/or recombination; A/B allelism should have existed in common ancestors of primates; and bacterial ABO genes evolved through horizontal and vertical gene transmission into 2 separate groups encoding glycosyltransferases with distinct sugar specificities.T he human histo-blood group ABO system is crucial in safe blood transfusion and cell/tissue/organ transplantation 1,2 . This system consists in A and B oligosaccharide antigens expressed on red blood cells (RBCs) as glycoproteins and glycolipids and antibodies against those antigens in serum. A and B antigens are also expressed by epithelial and endothelial cells, and in secretor type individuals they are also expressed on mucins secreted by exocrine glands. The immuno-dominant structures of A and B antigens are GalNAca1-.3(Fuca1-.2)Gal-and Gala1-.3(Fuca1-.2)Gal-, respectively. A and B alleles of the ABO genetic locus encode A and B transferases, which respectively transfer an N-acetyl-D-galactosamine (GalNAc) or a D-galactose (Gal) to H substances with an a1,3-glycosidic linkage. H substances with the Fuca1-.2Gal-structure are synthesized by fucosylation catalyzed by a1,2-fucosyltransferases (a1,2-FTs) encoded by FUT1/FUT2/SEC1 genes. FUT1-encoded a1,2-FTs and FUT2/SEC1-encoded a1,2-FTs exhibit distinct acceptor substrate specificity, and are differentially expressed amongst tissues. In humans SEC1 is a pseudogene and FUT2 gene presents frequent null alleles so that about 20% of individuals are incapable of expressing either H, A, or B antigens in secretions (nonsecretor type). In the absence of a1,2-FTs no H antigens are produced. Therefore, A/B transferases function only when at least one active a1,2-FT is simultaneously present.In 1990 we correlated the nucleotide sequences of A, B, and O allelic cDNAs and the expression of A and B antigens, and elucidated the molecular genetic basis of human histo-blood group ABO system 3,4 . Four amino acid substitutions (Arg176Gly, Gly235Ser, Leu266Met, an...

The detailed catalytic mechanism by which glycosyltransferases catalyze the transfer of a glycosyl residue from a donor sugar to an acceptor is not known. Through the multiple alignment of all known eukaryotic glycogen synthases we have found an invariant 17-amino acid stretch enclosed within the most conserved region of the members of this family. This peptide includes an E-X 7 -E motif, which is highly conserved in four families of retaining glycosyltransferases. Site-directed mutagenesis was performed in human muscle glycogen synthase to analyze the roles of the two conserved Glu residues (Glu-510 and Glu-518) of the motif. Proteins were transiently expressed in COS-1 cells as fusions to green fluorescence protein. The E510A and E518A mutant proteins retained the ability to translocate from the nucleus to the cytosol in response to glucose and to bind to intracellular glycogen. Although the E518A variant had approximately 6% of the catalytic activity shown by the green fluorescence protein-human muscle glycogen synthase fusion protein, the E510A mutation inactivated the enzyme. These results led us to conclude that the E-X 7 -E motif is part of the active site of eukaryotic glycogen synthases and that both conserved Glu residues are involved in catalysis. We propose that Glu-510 may function as the nucleophile and Glu-518 as the general acid/base catalyst.Glycosyltransferases and glycosidases catalyze the transfer of glycosyl residues from a donor sugar to an acceptor. The acceptor in glycosidases is water, the end result being hydrolysis of the glycoconjugate. For transferases the acceptor molecule is in most cases a growing carbohydrate chain, but it can also be a protein, a lipid, or a range of other compounds such as steroids, bilirubin, flavonones, carotenoids, etc