The products of several family GT2 P-glycan synthases have dual monosac-charide units and dual linkage types in repeating units in the polysaccharide chain. For example, the membrane bound synthase for the capsular type 3 pneumococcal polysaccharide with a repeating unit GlcP (1,4)GlcAP(1,3) is a 49-kDa protein encoded by the cap3B gene (Arrecubieta et al. 1996 ) and has a dual specificity, synthesizing both (1,3)- and (1,4)-linkages in the polysac-charide. In this system the chain growth can be terminated by withdrawal of the UDPGlcA needed for synthesis. In vivo UDPGlcA levels, controlled by UDPGlcA dehydrogenase activity, may control chain growth (Ventura et al. 2006 ).
Synthesis of the type 37 (1,3;1,2)-P-d-glucan pneumococcal polysaccharide, in which every 1,3-linked glucosyl residue in the backbone is substituted by a (1,2)-P-l inked glucosyl residue, is determined by a single gene (tts) located distant from the cap locus responsible for capsular formation in all other pneumococal types. The tts gene encodes a GT2 glycosyltransferase that is an integral membrane protein with a potentially cleavable signal sequence. Cell-free membrane preparations support the synthesis of the type 37 polysaccharide from UDP[14C]-glucose without the participation of a lipid intermediate. The synthase has a dual specificity, synthesizing both (1,3)-and (1,2)-linkages in this side-chain-branched polymer, a feature shared with the synthases producing the type 3 pneumococcal polysaccharide, hyaluronan in S. pyogenes, the K5 capsular polysaccharide in Escherichia coli, and chondroitin and heparosan from P. multocida. These are different from the NdvB and NdvC synthases for the cyclic (1,3;1,6)-P - glucan.
Similarly, the streptococcal and eukaryotic hyaluronan synthases are transmembrane proteins with large intracellular, central domains that catalyse the incorporation of both GlcpA and GlcpNAc from the respective uridine nucleotides to form a repeating unit. For the S. pyogenes synthase there is both biochemical and genetic evidence (DeAngelis & Weigel 1994; Tlapak-Simmons et al. 1998) that only the hyaluronan synthase gene product is required for hyaluronan synthesis. No primer is needed. Thus, this 48-kDa protein (Kumari & Weigel 1997) binds the two substrates and synthesizes both linkage types in the active site. Whether there are two substrate binding sites and two glycosyl transferase catalytic sites, or whether there is one general binding/catalytic site that alternates in binding and catalytic specificity, remains to be demonstrated.
Finally, the E. coli K5 capsular polysaccharide heparosan, a homologue of heparan, has a repeating ^4)GlcpAP(1,4)GlcpNAca(1^ unit and its syn-thase is a 60-kDa, bifunctional, repetitive glycosyl transferase. There are two transferase catalytic sites in the synthase protein; removal of 139 amino acids from the C-terminus of the protein results in loss of UDPGlcpA activity, but UDPGlcpNAc activity is retained (Griffiths et al. 1998 ).
In each case the synthases are molecules of 48-60 x 106kDa, their action patterns are repetitive and they show high sequence homology with one another. In E. coli K5 heparosan synthase there are two transferase catalytic sites; removal of 139 amino acids from the C-terminus of the protein results in loss of UDPGlcpA activity, but UDPGlcpNAc activity is retained (Griffiths et al. 1998). Mutational studies with P. multocida hylauronan and chondroitin synthases indicate that they also possess independent hexosamine and glu-curonic acid transfer sites (Jing & DeAngelis 2000). Kinetic studies suggest that there are separate receptor binding pockets for each of these glycosyl transferase activities that apparently interact with three or four saccharide units of the nascent hyaluronan chain (Williams et al. 2006 ).
Although GT2 enzymes with dual specificity have not yet been described in higher plants, the examples presented above suggest that their existence in higher plants is possible. The corollary to that possibility is that, under appropriate conditions, plant polysaccharides such as (1,3;1,4)-P-d-glucans might be synthesized by a single enzyme, rather than a multienzyme complex.
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