EFRAIM RACKER 



The kinetic complexities of metabolic processes, such as 

 glycolysis and the citric acid cycle, which are catalyzed by 

 multienzyme systems, have frequently been pointed out. In 

 order to analyze the individual enzymatic steps, it was necessary 

 to separate the catalysts from each other. Now it slowly becomes 

 apparent that the surface of the single enzyme may mirror some 

 of the complexities encountered in multienzyme systems. The 

 fact that a reaction catalyzed by a single enzyme includes the 

 formation of several enzyme-substrate intermediates, required 

 the introduction of the "Michaelis compound" as the rate- 

 limiting step (4,5). It may be confusing at first to learn that 

 pure crystalline glyceraldehyde-3-phosphate dehydrogenase 

 can catalyze five apparently different reactions {cf. 37). One 

 can explain oxidation-reduction, acyl transfer, phosphorolysis, 

 and perhaps even the phosphatase activity exhibited by this 

 enzyme in terms of the over-all process of aldehyde oxidation 

 coupled to phosphorylation. However, there is at present no 

 plausible explanation for the destruction of DPNH in the pres- 

 ence of the enzyme (36). It is very difficult to obtain accurate 

 kinetic data for the individual steps catalyzed by TDH, since 

 some of the "side reactions" interfere with the measurements. 

 Moreover, small modifications in the protein molecule, such as 

 the blocking of the SH groups, may change the ratios of the 

 different catalytic activities. Since oxidation of SH groups 

 occurs during the purification of TDH, a microheterogeneity is 

 introduced which is more treacherous for kinetic studies than 

 gross impurities with unrelated proteins. This difficulty cannot 

 be overcome by the addition of cysteine or GSH, which not only 

 reduce the enzyme incompletely but also interact with the 

 aldehyde substrate. However, fully reduced enzyme, suitable 

 for kinetic studies, can be isolated from rabbit muscle with the 

 aid of ethylenediamine tetraacetate (23), 



A change in pYL may determine which of a number of 

 possible products is formed by an enzyme. In the case of papain 

 and the cathepsins (15) a decrease in the hydrogen ion con- 

 centration alters the enzyme-catalyzed reaction in such a 



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