68 INTERMEDIARY METABOLISM AND GROWTH I 



these substances while other mutants can utiHze anthraniUc or indole for growth 

 (Yanofsky, 1955a, 1956). The indole-anthranilic pathway is also consistent with 

 the results of isotope experiments. 



Glucose is a more direct precursor of the protein tryptophane of Aerobacter 

 aerogenes than is acetate. Glucose-3-''*C is converted to the carboxyl carbon of 

 tryptophane and to several ring carbon atoms (Rafelson, 1955a, b). 



In Neurospora, the amino-nitrogen is retained and the carboxyl group is lost 

 during the conversion of anthranilic acid to indole (Yanofsky, 1955a). Isotope 

 experiments with labelled glycerol or labelled glucose suggested that carbons two 

 and three of the indole ring were derived respectively from carbons one and two 

 of a ribose derivative. On the basis of studies with E. coli extracts, 5-phosphoribosyl- 

 I -pyrophosphate (PRPP) was implicated as the actual donor of the two carbon 

 atoms (Yanofsky, 1955b). An E. coli enzyme was obtained which catalyzed the 

 condensation of the former substance with anthranilic acid to give indole-3-glyc- 

 erol phosphate. The latter substance was isolated from the incubation medium 

 (Yanofsky, 1956). Indole-3-glycerol is accumulated by several tryptophane 

 requiring strains oi E. coli. A second enzyme converted indole-3-glycerol phosphate 

 to indole and triose phosphate. 



The final step in tryptophane synthesis is the pyridoxal phosphate catalyzed 

 condensation of indole and serine (Tatum and Shemin, 1954). By using serine 

 labelled with ^''C in the beta carbon, ^^N in the amino group, and deuterium on 

 the alpha and beta carbons, it was shown that in Neurospora the condensation 

 involves an intramolecular dehydration of serine and a condensation of the alpha 

 aminoacrylic acid-pyridoxal phosphate intermediate thereby formed with indole. 

 This follows from the fact that one deuterium atom is lost in the condensation 

 reaction. Had the condensation resulted from an intermolecular dehydration 

 reaction, no deuterium would have been lost. 



{e) Alanine, valine, leucine, and isoleucine 



Alanine. Alanine is formed by transamination from the glycolytic intermediate, 

 pyruvate. The conversion of labelled pyruvate, lactate, glucose, or glycerol to 

 free alanine has been demonstrated both in vivo and in vitro in a variety of animal 

 tissues (Busch et al., 1956; Kit and Greenberg, 1951; Kit and Graham, 1956a). 

 Neoplastic tissues are particularly active in converting labelled substrates to 

 alanine. 



Valine and isoleucine. Isotope competition experiments with E. coli and Neurospora 

 suggested that a metabolic relationship exists between valine and leucine and that 

 both amino acids belong to the "pyruvate family" (Abelson, 1954; Abelson and 

 Vogel, 1955). Thus, the incorporation of glucose-* ''C into protein valine and 

 leucine is reduced by non-labelled pyruvate, oi-ketoisovalerate and valine. Evidence 

 as to the intermediates involved was derived initially from nutritional experiments 

 with microorganisms. A Neurospora mutant was isolated which reqviired both 

 valine and isoleucine for growth. This mutant accumulated a, p-dihydroxy-^- 

 methyl valeric acid and a, [^-dihydroxyisovaleric acid in the culture medium. The 

 mutant was able to grow, however, if a-ketoisovaleric acid and a-keto-^-methyl 



