A retrospective review of the roles of multifunctional glucose-6-phosphatase in blood glucose homeostasis: Genesis of the tuning/retuning hypothesis

Abstract

In a scientific career spanning from 1955 to 2000, my research focused on phosphoenolpyruvate carboxykinase and glucose-6-phosphatase. Grounded in basic enzymology, and initially pursuing the steady-state rate behavior of isolated preparations of these critically important gluconeogenic enzymes, our key findings were confirmed and extended by in situ enzyme rate experiments exploiting isolated liver perfusions. These efforts culminated in the discovery of the liver cytosolic isozyme of carboxykinase, known today as (GTP)PEPCK-C (EC4.1.1.32) and also revealed a biosynthetic function and multicomponent nature of glucose-6-phosphatase (EC3.1.3.9). Discovery that glucose-6-phosphatase possessed an intrinsically biosynthetic activity, now known as carbamyl-P:glucose phosphotransferase - along with a deeper consideration of the enzyme's hydrolytic activity as well as the action of liver glucokinase resulted in the evolution of Tuning/Retuning Hypothesis for blood glucose homeostasis in health and disease. This THEN & NOW review shares with the reader the joy and exhilaration of major scientific discovery and also contrasts the methodologies and approaches on which I relied with those currently in use.

Fig. 1

Some hydrolytic and biosynthetic reactions catalyzed by glucose-6-phosphatase (Nordlie and Arion 1964a; Lueck and Nordlie 1970).

Fig. 2

Kinetic mechanism of multifunctional glucose-6-phosphatase-phosphotransferase. RP is generalized phosphoryl substrate, e.g. PP_i or carbamyl-P; E represents enzyme. Further details are given in the text. (From Nordlie 1974 by permission of Academic Press, New York).

Fig. 3

Structure-function relationships of the glucose-6-phosphatase system. The substrate translocase-catalytic unit model presented here (Foster et al. 1991) is based on earlier concepts of Arion et al. (1975). Depicted is a cross section of the endoplasmic reticulum (E.R. MEMBRANE); SP, stabilizing protein; T₁, T₂, and T₃, substrate/product transporters with the indicated specificity; Catalytic unit, glucose-6-phosphatase (EC 3.1.3.9) enzyme. Two forms of T₂ with differing specificity, termed T₂α and T₂β, have been proposed (Nordlie et al. 1992). (From Foster et al. 1991 by permission of Elsevier Science BV).

Fig. 4

Classic role of hepatic glucose-6-phosphatase in the regulation of blood glucose concentrations. The charged glucose-6-P molecule is impermeable to the plasma membrane. Net rate and direction of flux of glucose between blood and liver is postulated to be determined by the relative rates of glucose phosphorylation (B) and glucose-6-P hydrolysis (A), as first suggested by Cahill et al. (1959), and more recently elaborated upon by others (see Nordlie 1974). (From Nordlie 1980; by permission from University Park Press, Baltimore).

Fig. 5

Schematic depiction of interrelationships among liver, blood, and peripheral tissues with respect to glucose flow and utilization. Net rate and direction of flux of glucose between liver and blood is determined by the relative rates of glucose phosphorylation (B) and glucose-6-P hydrolysis (A), as generally postulated by Cahill et al. (1959). The latter of these two processes (A) is catalyzed by hepatic glucose-6-phosphatase, while the former (B) involves hexokinase, glucokinase, and, we propose, transferase activity of glucose-6-phosphatase-phosphotransferase as well. (From Nordlie, 1974; by permission from Academic Press, New York).

Fig. 6

Activities of several hepatic enzymes, calculated as a function of varied glucose concentrations on the basis of presumed physiologic concentrations of other substrates and further adjusted for inhibition by estimated physiologic concentrations of various inhibitors. These inhibitors include ATP, P_i, HCO₃⁻, and glucose-6-P for carbamyl-P:glucose phosphotransferase; and ATP, P_i, HCO₃⁻ for glucose-6-P phosphohydrolase activity. Hexokinase has been adjusted for inhibition by glucose and glucose-6-P. Details of calculation are given elsewhere (Nordlie 1971). Abbreviations employed are: G-6-P HY., glucose-6-P phosphohydrolase; CPGT, cabamyl-P:glucose phosphotransferase; GK, glucokinase; and HK, hexokinase. Vertical arrows designate crossover points, that is, those concentrations of glucose at which rates of glucose-6-P hydrolysis and glucose phosphorylation by the indicated combinations of enzymes, are equal. (From Nordlie 1974; by permission from Academic Press, New York).

Fig. 7

Net glucose uptake rates in livers from 48-hr fasted (A) and fed (B) rats perfused with various concentrations of D-glucose. Initial perfusate glucose concentrations are indicated on the axes of abscissa either as millimolar concentrations (II) or 10⁻² × mg/100 ml (I). Closed circles indicate net glucose uptake values ± 1 S.D. (vertical bars). Dashed lines indicate summation of glucokinase plus hexokinase activitiy values calculated for the perfused glucose loads. Further details are given in Alvares and Nordlie 1977 . (Reprinted by permission of the American Society for Biological Chemistry and Molecular Biology).

Fig. 8

Effects of varied glucose concentrations on net uptake rates in perfused livers from fed control (X) and glucagon-treated alloxan-diabetic rat livers perfused in the absence (O) and presence (□) of 4 mM 3-mercaptopicolinate. Data for diabetic livers are from Tables I and III of Nordlie et al. 1982. Those for controls, included for reference, are from Alvares and Nordlie 1977. Cross-over points from net glucose production to net glucose utilization were observed at 6 mM, 22 mM, and 4 mM glucose with livers from control, diabetic, and mercatopicolinate-supplemented diabetic preparations, respectively. (Reprinted from Nordlie et al. 1982; by permission from Elsevier SV).

Fig. 9

The author THEN – 1969

Fig. 10

The author NOW at Island Lake – August 2009