Kinetics yeast hexokinase

Coe and Bessell and coworkers studied the metabolic fates of 2-deoxy-2-fluoro-D-glucose (2DFG) and related compounds by using yeast hexokinase (as a model for mammalian hexokinase), and determined the kinetic constants K and V ) of the Michaelis-Menten equation D-glucose 0.17 (K in mAf)> 1 00 (relative value, D-glucose taken as 1) 2DG 0.59 0.11, 0.85 2DFG 0.19 0.03, 0.50 2-deoxy-2-fluoro-D-mannose (2DFM) 0.41 0.05, 0.85 2-deoxy-2,2-difluoro-D-nraZ>//Jo-hexose... 

Treatment of 9-(/ -D-ribofuranosyluronic acid)adenine with diphenylphosphoro-chloridate and orthophosphate or tripolyphosphate yields (62) and (63), which, although unstable, inhibit rabbit AMP aminohydrolase and pyruvate kinase, respectively, with behaviour characteristic of active-site-specific reagents.98 Adenylate kinases from several sources are inactivated by iV6-[2- and 4-fluorobenzoyl]-adenosine-5 -triphosphates, with kinetics characteristic of active-site labelling, although these compounds were without effect on yeast hexokinase and rabbit pyruvate kinase.99... 

For this reason, these alternative routes for isotope combination with enzyme-substrate and/or enzyme-product complexes ensures that raising the [A]/[Q] or [B]/[P] pair will not depress either the A< Q or the B< P exchanges. Fromm, Silverstein, and Boyer conducted a thorough analysis of the equilibrium exchange kinetic behavior of yeast hexokinase, and the data shown in Fig. 2 indicate that there is a random mechanism of substrate addition and product release. 

This enzyme plays a key role in the metabolism of glucose and other related sugars. The physical and kinetic properties of yeast hexokinase have been extensively studied. Numerous recent studies have been made of its role in the phosphoryl transfer reaction. 

Kinetic Constants for the Sugar Substrates of Yeast Hexokinase... 

In the last 15 years a large number of publications have been concerned with the determination of the kinetic mechanism of yeast hexokinase. Unfortunately, there has been much disagreement between various authors on the conclusions to be drawn from such studies. 

In a two-substrate reaction similar to that catalyzed by hexokinase, two basic mechanisms may be at work. First, a ping-pong reaction may be occurring in which the enzyme shuttles between a stable enzyme intermediate, such as a phosphorylated enzyme, and a free enzyme. Second, the reaction may be sequential, in which case no reaction occurs until both substrates are on the enzyme. There are two types of sequential mechanisms. If one substrate cannot bind until after the addition of the other substrate the mechanism is said to be ordered. However, if they can combine in any order the mechanism is said to be random. The various kinetic methods for distinguishing between these mechanistic forms have been summarized by Cleland (52). The evidence for and against these possible kinetic schemes will now be summarized for yeast hexokinase. 

Haldane relationships can also be useful in characterizing isozymes or the same enzyme isolated from a different source. Reactions catalyzed by isozymes must have identical equilibrium constants, but the magnitudes of their kinetic parameters are usually different (e.g., the case of yeast and mammalian brain hexokinase ). Note that the Haldane relationship for the ordered Bi Bi mechanism is = Hmax,f p i iq/(f max.r ia b)- This same... 

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