Thus aldolase catalyzing the reversible reaction of F P synt

Thus, aldolase catalyzing the reversible reaction of F1,6-P2 synthesis is involved in the formation of the glyconeogenic or glycolytic complex, dependently on the physiological state of the cell. In the process of glycogen synthesis, the enzyme not only supplies the substrate for FBPase and desensitizes it to AMP inhibition [1], [3], but also partially desensitizes FBPase to action of calcium – the inhibition of the complex is about 50 times weaker than free muscle FBPase (Fig. 7) [14]. Presumably, Ca2+ does not inhibit directly the activity of FBPase associated with aldolase. It seems that calcium firstly destabilizes the complex and then inhibits the activity of free FBPase. Such a mode of the inhibition by calcium and the desensitization to the inhibitor might enable the activity of glyconeogenesis during the moderate muscle activity.
Acknowledgements
Introduction Fructose 1,6-bisphosphatase (d-fructose 1,6-bisphosphate 1-phosphohydrolase, EC 3.1.3.11, FBPase) catalyzes hydrolysis of fructose 1,6-bisphosphate (Fru 1,6P2) to fructose 6-phosphate and inorganic phosphate in the presence of divalent metal ions like magnesium, manganese, cobalt or zinc Benkovic and De Maine, 1982, Tejwani, 1983. Vertebrate FBPases are inhibited competitively by fructose 2,6-bisphosphate and allosterically by AMP Pilkis et al., 1981, Van Schaftingen and Hers, 1981. Liver and muscle isozymes have been found in vertebrate tissues Tejwani, 1983, Gidh-Jain et al., 1994, Tillmann and Eschrich, 1998, Al-Robaiy and Eschrich, 1999, Stein et al., 2001. The liver FBPase is a regulatory enzyme of gluconeogenesis, the muscle isozyme participates in regulation of glycogen synthesis from noncarbohydrate precursors Ryan and Radziuk, 1995, Gleeson, 1996. Tissue distribution of FBPase isozymes has been investigated. The liver isozyme has been found in liver, kidney and lung. In skeletal muscle tissue, only the muscle isozyme is expressed, but in other tissues simultaneous expression of the two isozymes has been observed (Al-Robaiy and Eschrich, 1999). A high degree of similarity of vertebrate FBPases' kinetic properties has been reported. The basic difference between muscle and liver FBPases concerns their sensitivity toward AMP inhibition. I0.5 of muscle FBPase is 10 to 100 times lower than the corresponding value determined for liver isozyme Tejwani, 1983, Skalecki et al., 1995, Skalecki et al., 1999, Rakus et al., 2000, Rakus et al., 2003. A high degree of homology of primary structures of vertebrate FBPases has also been observed. The elucidation of the primary structure of human, mouse, rat and rabbit muscle FBPases revealed more than 95% identity and 98% homology among these proteins. So far only a part of the chicken muscle FBPase primary structure has been determined (166 a.a. residues). This sequence corresponds to the sequence of mammalian muscle FBPase, starting from amino Caspase-1, human recombinant proteinase receptor residue 82. Cellular and subcellular localization of FBPase isozymes has also been investigated. Schmoll et al. (1995) have found FBPase to be an astrocyte-specific enzyme in brain. Gizak et al. (2001) have located FBPase in pneumocyte II. Saez et al. (1996) have reported that in hepatic and renal cells FBPase is located in the perinuclear area. Recently, Gizak et al. (2003) have located the enzyme on both sides of the Z-line of skeletal muscle fibers and, surprisingly, in nuclei of pig cardiomyocytes (Gizak and Dzugaj, 2003). This prompted us to investigate subcellular localization of FBPase in chicken heart muscle cells. The results of immunocytochemical study with electron microscopy, as well as determination of FBPase activity in isolated cardiomyocyte nuclei, revealed that in chicken cardiomyocytes FBPase is also present inside the nuclei. The physiological meaning of this finding is discussed.
Materials and methods
Results
Discussion Our investigation revealed that, like in mammals, nuclei of chicken heart muscle cells also contain FBPase. Determination of I0.5 and, the partial primary structure of FBPase from isolated chicken cardiomyocyte nuclei, as well as from cytosol indicates, that the same muscle isozyme is present in both compartments. Compared to mammals, chicken has a higher glucose concentration in biological fluids (10–12 mM; Belo et al., 1976), and a higher rate of carbohydrate metabolism is observed. FBPase activity in chicken pectoral muscle is much higher than that in mammalian skeletal muscle. Nevertheless, a similar mechanism of FBPase regulation in mammalian and chicken muscles might be expected. Recently we reported that, like mammalian muscle FBPase, the chicken muscle isozyme is desensitized to AMP inhibition by muscle aldolase (Dziewulska-Szwajkowska et al., 2004). The presence of muscle FBPase in chicken cardiomyocyte nuclei raises a question concerning its physiological role in this compartment. Several enzymes of carbohydrate metabolism have been found in nuclei: glucokinase (Miwa et al., 1990), aldolase Saez and Slebe, 2000, Mamczur and Dzugaj, 2004, glyceraldehyde 3-phosphate dehydrogenase, phosphoglycerate kinase (for review, see Ronai, 1993), glycogen synthase (Ferrer et al., 1997) and FBPase (Gizak and Dzugaj, 2003). The physiological role of the nuclear enzymes is rather an enigma. Unlike glycogen synthase, whose function is supposedly the same in the cytosol and in the nucleus, the physiological roles of the other enzymes seem to vary from the cytosol to the nucleus. It has been hypothesized that phosphoglycerate kinase may participate in DNA synthesis and cell-cycle progression (Popanda et al., 1998). Glyceraldehyde 3-phosphate dehydrogenase recognizes the sequence and structural features of the RNA and is involved in transcription (Sirover, 1997); lactate dehydrogenase is a recognized stabilizing factor and it participates in DNA reparation (Popanda et al., 1998).