Biochemical Characterization of a Novel Mammalian Polyphosphate Dependent Glucokinase

Hexokinases are a family of enzymes that catalyse the phosphorylation of glucose by transferring the γ- phosphoryl group from adenosine triphosphate (ATP) to the sixth position hydroxyl group of glucose to generate Glucose 6-phosphate (G6P).

Until now, five isozymes of mammalian hexokinase (HK) have been described: types I, II, III and IV, all of them ATP dependent, and type V which is ADP-dependent. The present thesis describes a novel hexokinase, we have designated as PPGKm. The enzyme is strictly polyphosphate (pp) dependent, is correctly defined as a glucokinase (GK), by virtue of its kinetics and is present in mammalian tisues, at high activity in liver. The enzyme does not use ATP and ADP and indeed appears to be inhibited by them. As far as can be ascertained this is the first description of a mammalian enzyme using inorganic polyphosphate as a phosphoryl donor. Polyphosphate is used as phosphoryl donor in bacterial systems and is quite well characterized, although only two micro-organisms have been shown to be strictly pp-dependent like the one described here.

This novel enzyme (PPGKm) also showed unique features, differing from the others hexokinases in having a longer half-life and can be stored for several months at -20˚C without loss of activity. However, thermal stability was lower than other hexokinases studied.

The enzyme activity is concentrated in the hepatocyte nucleus has a higher molecular weight compared to Hexokinases 1, 2 and 3. The full biological significance of the enzyme is as yet unclear, and attempts to purify and sequence it have been only partly successful. Its specific role in cellular, and especailly nuclear metabolism remains unknown.