Mammals and birds are sensitive either to

Mammals and birds are sensitive either to hypo or hyper glycemia. In contrary some frogs when freezing/thawing experience very high glycemia (Storey and Storey, 1986). Nevertheless when the effect of temperature on Fru-2,6P2 concentration was investigated, the constant glucose level in frogs kept at 5 and 25 centigrade was observed (Dziewulska-Szwajkowska et al., 1997). Our investigation revealed that the frog muscle enzyme is sensitive to inhibition by calcium ions like the mammalian muscle FBPase. Recently we have presented evidence that the loop 52–72 is critical for the inhibition of the muscle FBPase by calcium and that the Glu69 is essential for the high affinity of the enzyme toward calcium ions (Zarzycki et al., 2007). The primary structure of the loop 52–72 of the P. esculentus muscle FBPase presented in Fig. 3 indicates high homology with corresponding structures of the mammalian and frog muscle FBPases. In all known primary structures of muscle FBPases the Glu69 is present in comparison with Gln69 in the liver isozyme. The crystallographic analysis and site-directed mutagenesis (Villeret et al., 1995, Choe et al., 2000) revealed three metal Pyridoxal isonicotinoyl hydrazone of the liver FBPase. It has been reported (Zhang et al., 1994) that magnesium binds to the first and to the second metal binding site and that potassium binds to the third site only. The ionic radius of potassium is nearly twice larger than the corresponding radius of magnesium. The ionic radius of calcium is nearly 40% larger than that of magnesium. Hypothetically calcium should bind to site three rather than to site one or two, particularly that Asp68 has been localized in the third site in the liver FBPase. In the muscle enzyme the presence of two acidic residues (Asp68 and Glu69) interacting with calcium might stabilize the loop 52–72 in the disengaged conformation. The shape of the inhibition curve of frog muscle FBPase by calcium ions indicates the cooperativity in the interaction of calcium ions with the enzyme. Supposedly interaction of calcium ions with P. esculentus muscle FBPase also results in “disengaged” conformation. It is feasible that in frog muscle glyconeogenesis is regulated by Fru-2,6P2 and additionally by calcium ions. Hypothetically, during the FBPase evolution, the substitution of Gln69/Glu resulted in a change in the kinetic properties of the new isozyme making it sensitive toward inhibition by calcium and thus enabling the regulation of glyconeogenesis in muscle cells by calcium ions. Glu69 is the decisive but not the only amino acid residue involved in FBPase calcium interaction, additional mutations were probably necessary to make mammalian FBPase very sensitive towards calcium inhibition.
Introduction Type 2 diabetes mellitus (T2DM), which accounts for more than 90% of all diabetics, is characterized by fasting hyperglycemia and an excessive rise in the plasma glucose concentration after glucose or meal ingestion. The global incidence of this disease is predicted to rise to more than 366 million by the year 2030. T2DM usually leads to complications such as retinopathy, nephropathy, and neuropathy. Clinical studies have suggested that fasting hyperglycemia in T2DM is associated with excessive glucose production through gluconeogenesis. Thus, the inhibition of gluconeogenesis is a useful approach in reducing increased blood glucose levels in patients with T2DM. Fructose-1,6-bisphosphatase (FBPase) is one of the rate-limiting enzymes of hepatic gluconeogenesis. FBPase is predominantly expressed in the liver and the kidney and catalyzes the hydrolysis of fructose-1,6-bisphosphate to fructose-6-phosphate. FBPase inhibitors would lower blood glucose levels by reducing hepatic glucose output and are expected to be a novel class of drugs for the treatment of T2DM. There are several small-molecule inhibitors of FBPase,4, 5, 6, 7, 8, 9 in which AMP mimetic MB05032 (1) exhibited high inhibitory activity (Fig. 1). The prodrug of MB05032 (CS-917, 2) lowered blood glucose levels in animal models and was in clinical development.