26 ENZYMES 



Exercise V 



properties, on the hydrogen ion concentration 

 of the medium. Each enzyme tends to be most 

 active over a narrow range of hydrogen ion 

 concentration, the "pH optimum." Enzymes 

 are rapidly destroyed by boihng, as are proteins 

 generally. 



Another general property of enzymes, as of 

 other proteins, is specificity. Each enzyme cata- 

 lyzes only one or a narrow class of chemical 

 reactions. Hence thousands of different enzymes 

 are needed to catalyze the multitude of chemical 

 reactions carried out by living cells. 



It is one of the triumphs of modern bio- 

 chemistry to extract enzymes and enzyme sys- 

 tems from cells and have them catalyze in the 

 test tube the same reactions and reaction se- 

 quences that we find in living organisms. 

 Indeed a great number of enzymes have been 

 prepared pure and crystalline, and many are 

 now bought and sold commercially like other 

 organic substances. 



In this period we shall work with three enzyme 

 systems, each of which has something special to 

 tell us. Succinic dehydrogenase is an oxidation- 

 reduction (hydrogen-transferring) enzyme, of 

 central importance in cellular respiration. With 

 it we can demonstrate hydrogen transfer, and the 

 mechanism of action of a powerful respir- 

 atory poison. Amylase is a digestive enzyme, 

 which catalyzes an almost irreversible hy- 

 drolysis; with this system we can readily measure 

 the effects of changing enzyme concentration, 

 pH, and temperature on the rate of reaction. 

 Phospliorylase catalyzes the coming to equilib- 

 rium of a reversible system, and so permits us to 

 study synthesis as well as degradation, depending 

 upon how the system is constituted. 



SUCCINIC DEHYDROGENASE 



The citric acid or Krebs cycle is central among 

 the enzyme systems concerned with cellular 

 respiration, the process by which organic mole- 

 cules are burned with molecular oxygen to 

 carbon dioxide, water, and energy in forms use- 

 ful for cellular work. One of the steps of this 



cycle is the oxidation of succinic acid to fumaric 

 acid. This reaction is catalyzed by the enzyme 

 succinic dehydrogenase: 



This reaction can be followed by observing 

 the loss of color of the dye methylene blue 

 (MB) as it is reduced to the colorless form 

 "methylene white" (MB-H2) by accepting the 

 two hydrogen atoms removed from succinic acid. 



We have already spoken of the specificity of 

 enzymes for their substrates. Succinic dehydro- 

 genase, so far as we know, catalyzes only the 

 dehydrogenation of succinic acid, in part be- 

 cause the catalytically active site on the enzyme 

 molecule combines readily with succinic acid to 

 form the enzyme-substrate complex. Sometimes, 

 however, it is possible to fool an enzyme by 

 offering it a molecule that so greatly resembles its 

 normal substrate that the enzyme combines with 

 the impostor instead. Such a molecule in the 

 present instance is malonic acid : 



COOH 



I 

 CH2 



I 

 COOH 



Succinic dehydrogenase, having combined with 

 malonic acid rather than succinic acid, can 

 neither dehydrogenate it nor lose it again. 

 Thus its active site is blocked, and the enzyme 

 is inhibited or poisoned. The inhibition is as 

 specific as the enzyme action and for the same 

 reason. It can be reversed by adding an excess 

 of succinic acid, which competes with malonic 

 acid for the catalytic site. We call malonic acid 

 for this reason a competitive inhibitor. Its action 

 on succinic dehydrogenase makes it about as 

 powerful a poison of cellular respiration as 

 cyanide. 



