24 Samuel Gurin and Roscoe O. Brady 



Brady, R. O., and Gurin, S. (1951). J. biol. Chem., (in press). 

 Brady, R. O., Lukens, F. D. W., and Gurin, S. (1951a). Science (in 



press). 

 Brady, R. O., Lukens, F. D. W., and Gurin, S. (19516). Unpublished. 

 Buchanan, J. M., Sakami, W., and Gurin, S. (1947). J. biol. Chem., 



169, 411. 

 Coon, M. J. (1950). J. biol. Chem., 187, 71. 



JowETT, M., and Quastel, J. H. (1935). Biochem. J., 29, 2181. 

 Little, H. N., and Block, K. (1950). J. biol. Chem., 183, 33. 

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DISCUSSION 



Bloch: We have come to the same conclusion as Dr. Gurin, namely 

 that acetoacetate is an intermediate in the conversion of acetate to 

 cholesterol. Dr. Zabin and I have carried out some experiments with 

 butyrate and isovalerate and have in both cases been able to ascribe 

 the conversion of these compounds to cholesterol to the intermediary 

 formation of acetoacetate. 



I would like to present also some highly speculative material on the 

 mechanism of cholesterol biosynthesis. The evidence that indeed all the 

 carbon atoms of the sterols can be derived from acetic acid is now quite 

 satisfactory. In considering various schemes for the condensation of 

 acetate units, one paper came to our attention which dealt with the 

 biosynthesis of an isoprenoid compound, namely natural rubber. 

 Bonner and Arreguin incubated isolated guayule leaves with various 

 substrates and found that acetate, acetone, acetoacetate, and /S-dimethyl 

 acrylate were potent carbon sources for natural rubber. On the basis 

 of these results Bonner and Arreguin postulated the following mechanism 

 for the formation of an isoprene unit from acetate: condensation of 2 mols 

 of acetate to acetoacetate, decarboxylation to acetone, which then 

 would condense with a further molecule of acetate to j8-dimethyl 

 acrylate, and then finally a conversion to an isoprene unit. 



Note that an isoprene unit if synthesized in this fashion from acetate 

 will contain three methyl carbons of acetate and two carboxyl carbons 

 of acetate. 



There have been many speculations (Channon, Heilbron, Robinson) 

 on the possible role of the isoprenoid hydrocarbon squalene as a precursor 

 of cholesterol. Last year, as a result of some discussions with Dr. 

 Gallagher, Dr. Langdon and I tried again to fit the squalene hypothesis 

 into the existing knowledge on the biosynthesis of cholesterol. This 

 polyisoprenoid hydrocarbon can be arranged as shown in Fig. 1, and a 

 structure is obtained which, by the formation of various cross-linkages, 

 could form the polycyclic steroid structure. Providing that squalene 

 is formed as postulated, namely by the condensation of acetate to 

 acetoacetate, acetone, and so forth, then one should obtain a squalene 

 molecule with the isotope distribution shown in Fig. 1, the circles indi- 

 cating methyl carbons and the crosses carboxyl carbons of acetate. In 



