This section on NDP-sugar biosynthesis begins with a description of an enzyme we identified and characterized that does not obey the general rule of enzyme specificity (at least not as we were accustomed to it with other enzymes we had purified in the past). This enzyme, named SLOPPY, is responsible for the synthesis of at least six different UDP-sugars and is unique to the plant kingdom. No other genes with a high sequence similarity to Sloppy have been identified in other organisms to date.

D-Fructose-6-P, a product of photosynthesis, is a central precursor for all monosaccharide residues in plants (403). Using labeling experiments, however, it was also elegantly shown that plant cells can readily take up other free sugars such as Rha, Glc, Gal, Xyl, GalA, GlcA, Ara, and Fuc, and incorporate them into polysaccharides. It was assumed that a free sugar was first phosphorylated with ATP and then uridylated or guanylated with UTP or GTP to form the corresponding NDP-sugar. Indeed, numerous kinase and pyrophosphorylase activities were isolated in the late 1950s and early 1960s that catalyzed such reactions. The kinases were never purified and it was not explicitly clear ifdifferent kinases catalyzed the C — 1 phosphorylation of each unique sugar or if one kinase phosphorylated several sugars. We discuss the different kinase activities and specificities below. Similarly, it was not clear if the subsequent pyrophosphorylation of each sugar-1-P by pyrophosphorylase was specific or not (411,412). Three independent research groups identified an enzyme in pea (413) and in Arabidopsis (414, 415) that can pyrophosphorylate various sugar-1-phosphates with UTP. The recombinant Arabidopsis protein (At5g52560), termed SLOPPY, has a higher affinity (e. g., lower Km) for GlcA-1-P but it also catalyzes the conversion of Glc-1-P, Gal-1-P, Xyl-1- P, Ara-1-P, and GalA-1-P to their respective UDP-sugars (414). The enzyme is very efficient and specific for the production of UDP-sugars and shows no detectable activity when TTP, GTP, ATP, CTP are substituted for UTP. Although Sloppy has broad sugar-1-P specificity, it cannot accept GalNac-1-P.

The existence of a non-specific pyrophosphorylase raises basic questions. What is the source of free sugars in plants? Do the free sugars contribute significantly to the flux of NDP-sugars for wall biosynthesis pathways? Are free sugars generated inside or outside the cells? If they are imported inside, are they derived from long-distances, cell-to-cell transport, or directly from recycled wall? These are central questions that both need to be addressed and obviously raise more questions. If indeed sugars are recycled from glycan degradation, are there sugar-specific transporters? Recently, a plasma membrane sugar transporter (AtPLT5) was functionally identified in Arabidopsis (416). The transporter is a member of a multigene family and seems to be a “non-specific transporter” since competition assays indicate that AtPLT5 can transport Xyl, Rib, Ara, Glc, and myo-inositol, but not sucrose. Unfortunately, the transport of other sugars such as Gal, Rha, and Fuc was not determined for AtPLT5. But it is likely that other sugars are transported either by this, or other, transporters.

The “recycling” of free sugars into the NDP-sugar pool was termed the “salvage path­way” (403). The relative amount of free sugars released from glycolipids, glycoproteins, wall polysaccharides, and glycosides of small metabolites is hard to quantify. Hence, at this time it is not possible to judge the relative contribution of the salvage pathway to the flux of NDP-sugars versus the carbon flux derived from photosynthesis. In the subsequent subsec­tions we will describe the synthesis of specific NDP-sugars.