This hypothesis is in line

This hypothesis is in line with our phylogenetic analysis of FBPase sequences which revealed that vertebrate FBP2 evolved significantly faster than FBP1 (Fig. 2). An average distance between FBP2 and ancestral vertebrate FBPase was higher than that for FBP1. Moreover, preliminary study on amphibia revealed that FBP2 from Pelophylax esculentus is inhibited by AMP like the mammalian muscle isozyme (I0.5=0.11μM) but relatively weakly, similarly to fish FBP2, affected by Ca (I0.5=91μM) (Dziewulska-Szwajkowska and Dzugaj, 2010). This indicates that the extremely high sensitivity of FBP2 to AMP arose earlier in vertebrate evolution than the sensitivity to calcium. The kinetic properties of piscine FBPases raise a question about the sensitivity to AMP and Ca of the ancestral vertebrate/chordate FBPase. Is the moderate sensitivity of fish FBP2 to the inhibitors a primal feature of vertebrates FBPase or is it a reflection of the fast specialization of FBP2 upon gene duplication? To find the answer, we tested the susceptibility of partially purified FBPase from fruit fly, Drosophila melanogaster, to inhibition by AMP and calcium. The activity of FBPase isolated from D. melanogaster was 1.1U/g of wet weight and the ratio of the activity measured at pH 9.3 to the activity at pH 7.5 was 0.3, indicating lack of the enzyme proteolysis. The inhibition of fruit fly FBPase by calcium was relatively strong (I0.5=150μM and Hill constant ~2.4) and similar to that of piscine and amphibian FBP2. On the other hand, AMP inhibited the Drosophila enzyme to the same degree (I0.5=5.0μM and Hill constant ~1.7) as it inhibits the all vertebrates FBP1. The low sensitivity of FBP1 to AMP and the lack of inhibition by the physiological [Ca] might guarantee the insensitivity of liver gluconeogenesis to the concentrations of metabolites and ions occurring within hepatocytes. In fact, it is commonly believed that the vertebrate liver gluconeogenesis is regulated mainly hormonally—by glucagon. On the other hand, the high sensitivity of FBP2 to AMP and Ca allows for the precise regulation of muscle glyconeogenesis by, respectively, gdc-0980 status of muscle fibers and muscle contraction, and for adequate acceleration of glycolysis. However, although Ca inhibits piscine FBP2 much weaker than the mammalian and avian isozyme, it is still able to disrupt the interaction of FBP2 with the Z-line (Fig. 3), as it was previously observed in the warm-blooded vertebrate muscles (Gizak et al., 2004, Mamczur et al., 2005). The association of FBP2 with the Z-line and with aldolase is presumed to be indispensable for glycogen synthesis from non-carbohydrates in warm-blooded vertebrates. From this, it is clear that the mammalian-like, very strong binding of Ca to FBP2 is not necessary for regulation of muscle glyconeogenesis. Thus, it might be supposed that the extremely high sensitivity of mammalian and avian FBP2 toward Ca is not related to the regulation of FBP2 activity and the stability of the glyconeogenic complex. It was demonstrated that mammalian FBP2 is involved in an adaptation to calcium-related stress signaling in cardiomyocytes (Gizak et al., 2012). We have shown that calcium-induced binding of FBP2 to mammalian mitochondria protects them against swelling and depletion of ATP. It is also well documented that mammalian muscles, in contrast to these of ectothermal vertebrates, display a close mitochondrion–calcium release units association (Franzini-Armstrong and Boncompagni, 2011). Additionally, we suggest that the high sensitivity of mammalian and avian FBP2 to Ca is not related to regulation of stability of the glyconeogenic complex, instead it ensures the precise regulation of mitochondrial metabolism during prolonged calcium elevation in contracting muscle fibers.
Introduction Fructose 1,6-bisphosphatase (FBPase; EC 3.1.3.11) catalyzes the hydrolysis of fructose 1,6-bisphosphate to fructose 6-phosphate and inorganic phosphate in the presence of divalent metal cations like magnesium, manganese, cobalt or zinc (Benkovic and De Maine, 1982, Tejwani, 1983). It seems that in all terrestrial vertebrates (mammals, birds, reptiles and amphibia) two FBPase genes, and therefore two isozymes, are present (Tillmann and Eschrich, 1998, Al-Robaiy and Eschrich, 1999, Tillmann et al., 2002). The liver isozyme, involved in regulation of gluconeogenesis, has been found primarily in gluconeogenic tissues like liver, kidney and lung (Mizunuma and Tashima, 1978, Tejwani, 1983, Skalecki et al., 1999). The muscle FBPase, participating in the glycogen synthesis from lactate (Ryan and Radziuk, 1995, Gleeson, 1996), is the sole isozyme expressed in skeletal muscle tissue. In other tissues simultaneous expression of the two isozymes has been observed (Al-Robaiy and Eschrich, 1999). Both isozymes are activated by monovalent cations like potassium or ammonium, inhibited competitively by fructose 2,6-bisphosphate and allosterically by AMP. The basic difference between the liver and muscle isozymes concerns their sensitivity to AMP inhibition. In the case of the muscle enzyme, I0.5 value for AMP is about 0.1μM, and it is 10–100 times lower than the same value determined for the liver isozyme (Tejwani, 1983, Skalecki et al., 1999).