VOL. 12 (1953) ENZYMES OF FATTY ACID METABOLISM 303 



CoA have recently become available through chemical synthesis. T. Wieland" has 

 succeeded in synthesizing these compounds by reaction of CoA-SH with the thio- 

 esters of acetoacetic or /3-hydrox3'butyric acid and thiophenol. S-acetoacetyl CoA has 

 also been prepared through the interaction of CoA-SH with diketene^**. 



The absorption spectra of acetoacetyl CoA and crotonyl CoA are essentially the 

 same as those of their thioethanolamine analogues. The main difference is that, in 

 contrast to N-acetyl thioethanolamine, CoA absorbs in the 260 m[x region because of its 

 adenine group. This does not interfere with the enol band of acetoacetyl thioesters but 

 interferes to some extent with the 263 m/x band of the croton}^ derivatives. However, 

 the extinction coefficient of the latter compounds at this wavelength is very high and 

 the interference of the adenine moiety of crotonyl CoA can be eliminated by addition of 

 adenine nucleotide, for example adenylic acid, to the blank cell. 



In contrast to the optical behavior of the acetoacetyl and crotonyl CoA derivatives, 

 j8-hydroxyacyl and saturated acyl CoA compounds show only the thioester band at 233 

 mfi in addition to the adenine band. It is therefore possible in enzymic experiments, to 

 follow the appearance and disappearance of j8-ketoacyl or dehydroacyl CoA compounds 

 if jS-hydroxyacj'l or acyl CoA compounds are involved in the reaction. At pH 8.0 the enol 

 absorption of acetoacetyl thioesters is markedly increased by magnesium ions^'^, 

 probably through formation of a chelate structure. This increase in absorption can 

 conveniently be made use of to increase the sensitivity of the optical enzyme tests. 



ENZYMES OF FATTY ACID METABOLISM 



Activating enzymes. As already stated the main mechanism for the activation of 

 fatty acids in animal tissues is through a reaction of the fatty acid with CoA-SH, in the 

 presence of ATP, to yield the corresponding S-acyl CoA, AMP, and PP. This reaction 

 requires the presence of Mg ionp. The first reaction of this type to be studied in detail 

 was the activation of acetate by an enzyme present in liver, yeast, and other tissues^^. 

 The mechanism of the over-all reaction was established by Lipmann et al^^ with partially 

 purified enzyme preparations from liver and yeast (Reaction i). Similar results were 

 obtained by Hilz and Lynen (unpublished experiments) with a highly purified enzyme 



CH3— COOH + HS— CoA + ATP^ CH3— C— S— CoA+AMP+PP (i) 



from yeast and by Green and collaborators^^'^^ with purified enzymes from heart 

 muscle and liver. The acetate enzyme is active also with propionate. 



Recent experiments^^ suggest that the over-all reaction occurs in three steps as 

 indicated below : 



(a) ATP + enzyme ^ AMP-enzyme + PP 



(b) AMP-enzyme + CoA :^ CoA-enzyme -|- AMP 



(c) CoA-enzyme -{- acetate ^ acetyl-CoA -|- enzyme 



The occurrence of reaction (a) is supported by the incorporation of labelled PP into 

 ATP in the presence of a partially purified enzyme from yeast.This exchange is dependent 

 upon the presence of Mg+^. Reaction (c) is supported by the incorporation of isotopic 

 acetate into acetyl CoA in the presence of the enzyme. 



An enzyme catalyzing the activation of fatty acids from C4 to C^g was isolated 

 from ox liver in D. E. Green's laboratory'^^. The enzyme catalyzes Reaction 2. The same 



References p. 3 13 j 314. 



