The distribution and synthesis of hepatic glucokinase.

Glucokinase is an ATP-glucose 6-phosphotransferase given a separate classification, EC 2.7.1.2, from the several isoenzyme forms of hexokinase (EC 2.7.1.1); this is justified by the difference in molecular weight, its relatively high K , value for glucose and the fact that it is not significantly inhibited by the product of the reaction, glucose 6-phosphate (Trayer, 198 I). Many observations on hepatic glucose metabolism had suggested that this tissue contains a glucose-phosphorylating enzyme with these kinetic properties, and the presence of glucokinase was demonstrated in several laboratories in the early 1960's (see Weinhouse, 1976, for review). Although earlier studies had indicated a K , of 10-20m~-glucose, a more detailed kinetic study on highly purified enzyme from rat liver showed some kinetic co-operativity towards glucose and a Kt of 5m~-glucose (Niemeyer el al., 1 9 7 5 ~ ; Storer & CornishBowden, 1976). Sigmoidicity in the response to glucose concentration has subsequently been demonstrated for metabolic events in isolated hepatocytes and in cell-free liver extracts from the rat (Bontemps et al., 1978). One of the best early demonstrations of the positive role of glucokinase in glycogen synthesis from glucose was the species-distribution study by Ballard (1965); incorporation of ['4Clglucose into glycogen essentially paralleled glucokinase activity. More recent studies have, however, cast uncertainties re the precise role of glucokinase in hepatic carbohydrate metabolism. Although glycogen synthesis from glucose when the latter is at a concentration of greater than about 30mM has been repeatedly demonstrated in vitro and also in vivo after the administration of large glucose loads (see Stalmans, 1976, for review), such experiments are performed under extreme conditions and care is required in their interpretation in case the incorporation of I4C into glycogen is due to exchange rather than net synthesis. The availability in recent years of suspensions of freshly isolated rat hepatocytes has permitted the clear demonstration (Katz et al., 1976) that glycogen synthesis occurs best from gluconeogenic precursors and in the presence of lower (and more physiological) glucose concentrations (approx 10 mM). Similar conclusions were made from studies with perfused liver (Hems el al., 1972; Whitton & Hems, 1977) and in vivo in mice (Baker, 1977). The formation of fatty acids from glucose in rat liver is a minor pathway both in vivo (Hems et al., 1975) and in vitro (Clark el al., 1974; Salmon el al., 1974; Bloxham ef al., 1977; Katz et al., 1977). Nevertheless, it remains clear that, for the utilization of glucose when it is available at a substantial concentration, the presence of glucokinase is required (J. Katz et al., 1979). The recycling between glucose and glucose 6-phosphate that can be demonstrated by using isotopic tracers (Katz & Rognstad, 1976; Katz et al., 1978) requires the simultaneous action of both glucokinase and glucose 6-phosphatase. When the detritiation of [2-3Hlglucose is used an an estimate of the rate of formation of glucose 6-phosphate, the latter is at a maximum when glucokinase activity is high and glucose 6-phosphatase activity comparatively low (Katz et al., 1975). The effect of the relative fluxes, determined by the relative concentrations of the substrates glucose and glucose 6-phosphate respectively, through the two enzymes upon the intracellular concentration of glucose 6-phosphate is considered to be of likely regulatory significance. Possible roles of this cycling have been discussed (Newsholme & Start, 1973; Hue & Hers, 1974; Katz et al., 1978). The sensitivity of the rate of glucose 6-phosphate formation to a small change in glucose concentration will be increased by the sigmoidal property of glucokinase noted above and may be part of a more complex circuitory yet to be discovered. The tissue localization of glucokinase again points to a unique role for the enzyme. Although controversy continues as to whether or not hepatic parenchymal cells contain low hexokinase activity (see, e.g., Walker, 1966; Weinhouse, 1976; Bontemps et al., 1978; Wakelam & Walker. 1980~). glucokinase is confined to the parenchymal cells and is absent from other liver cell types (Van Berkel, 1979). On the basis of micro-dissection studies, Katz et al. (1977) consider that the glucokinase activity may be higher in the perivenous regions of liver tissue compared with periportal areas. A number of claims for the presence of glucokinase activity in non-hepatic tissues have been made, some of which were noted by Weinhouse (1976). Davagnino & Ureta (1980) and Allen el al. (1980) have shown by various means that many of those and other reports of non-hepatic 'glucokinase' can be attributed to N-acetyl-Dglucosamine kinase activity. This latter enzyme can phosphorylate glucose, although the K , is very much higher than that of the glucokinase (Allen 8c Walker, 1980a,b). Although not all the tissues have been positively shown not to possess glucokinase activity, the need to consider this possibility is apparent and the means to do so are provided in those references. Hepatic glucokinase is also a unique glucose-phosphorylating enzyme in that it is adaptive (see reviews by Niemeyer et al., 19756; Weinhouse, 1976). This behaviour has been examined in most detail in the rat, in which species activity slowly decreases during starvation or the feeding of carbohydrate-free diet and increases again more rapidly upon re-feeding carbohydrate to such animals. Activity falls to very low values as a result of alloxanor streptozotocin-diabetes and is restored by treatment with insulin. This type of adaptive behaviour is seen in a number of other species (e.g. Ureta el al., 1971; Willms el al., 1970). Diabetes associated with ketosis, where there is an absolute insulin deficiency, seems to be invariably accompanied by very low glucokinase activity. The fall in glucokinase activity with starvation does not occur in some species, e.g. guinea pig, dog and some mouse strains; it is not known whether circulatory insulin concentrations change in these examples as it does in the rat. In forms of diabetes associated with both hyperglycaemia and hyperinsulinaemia (e.g. the spontaneous, non-ketotic forms in spiny mice and certain obese mouse strains and in human maturity-onset diabetes), elevated glucokinase activities have been recorded (Willms et al., 1970). The above facts lend comprehensive support for an involvement of insulin in the synthesis of glucokinase, which is confirmed by the marked fall in glucokinase activity aRer the injection of anti-insulin serum (Niemeyer et al., 1967; Ruderman & Lauris, 1968). Because ingestion of glucose results in release of insulin in normal animals, it has not been possible to decide whether the substrate glucose plays a direct role in the synthesis of glucokinase or whether all its effects are via insulin (Weinhouse, 1976). The high amounts of glucose in the insulin-deficient diabetic condition obviously cannot induce glucokinase synthesis alone. Low glucokinase activities appear for the first time in the weaning rat from about 16 days of age onwards, even if the animal is not weaned (Walker el al., 1974) or is weaned on to a carbohydrate-free diet (Walker & Holland, 1965). Other hormones may also be involved in the complex regulation of glucokinase activity in vivo. These have been discussed by Weinhouse (1976); a role for glucocorticoids is very debatable. Hormones such as adrenaline and glucagon, which operate by elevating the tissue cyclic AMP concentration, prevent glucokinase synthesis in vivo (Ureta el a/., 1970; Pilkis, 1970). A particularly interesting feature reported by Coleman (1977) was that glucokinase activity appeared to be under genetic