38. BIOSYNTHESIS OF PROTEINS IN BACTERIAL CELLS 411 



in heterotrophs than in prototrophs such as Escherichia coli.*- 5 The existence of an 

 active transport mechanism for the metabolites is not a priori evident in bacteria, 

 since the possibility exists of passive diffusion of organic salts and metabolites across 

 the cell membrane. Gale's early studies with Staphylococcus aureus 6 showed that 

 amino acid transport is energy dependent. From the work of Cohen, Rickenberg 

 ct al? • 8 and of Cohen and Monod with 0-galactosides, 9 and that of Cohen ct al. 10 and 

 Britten et al. 11 with amino acids, it is evident that E. coli contains systems (per- 

 meases) for concentrating these metabolites several hundred-fold within the intra- 

 cellular space. These systems also possess a high degree of specificity for the metabo- 

 lites. 911 Ingenious experiments by Sistrom 12 have demonstrated that /3-galactosides, 

 after their uptake by the permease, exist in a free state inside the cell, and more 

 precisely as shown by Rickenberg, 13 inside the cytoplasmic region limited by the 

 cell membrane. 



The ability of E. coli to actively concentrate a given amino acid can be dissociated 

 from its ability to synthesize protein. 10 ' " Therefore, when amino acid incorporation 

 into protein is measured, the first limiting factor is the system carrying out the spe- 

 cific concentration of this particular amino acid. For example, the valine permease 

 in E. coli becomes saturated when the concentration of valine in the medium reaches 

 2 X 1G" 5 M. 10 If radioactive valine is present at lower concentrations, its rate of in- 

 corporation during growth would give a false measure of the absolute rate of protein 

 synthesis. Similarly, when protein synthesis is measured by incorporation of a radio- 

 active amino acid in a culture of E. coli growing on simple mineral medium, inhibi- 

 tion of amino acid transport by a drug or a competitive inhibitor would be falsely 

 interpreted as an inhibition of protein synthesis. 



Interest in the intracellular pool of metabolites is not restricted to the methodology 

 for studying the rate of protein synthesis. Chemical analysis should lead, in princi- 

 ple, to the recognition of some intermediates in protein and nucleic acid synthesis 

 (either during normal growth or after inhibiting selectively the formation of a macro- 

 molecular component to allow the corresponding intermediates to accumulate). 



In fact, except for glutathione, free peptides are seldom found in large quantities 

 in the bacterial pool. There are a few interesting exceptions: for instance, after treat- 

 ment with one of several drugs (including penicillin) gram-positive bacteria accumu- 

 late considerable quantities of a mucopeptide. 1416 However, this peptide (which 

 contains alanine, glutamic acid, and lysine residues, and is bound to A r -acetylhexosa- 



4 E. F. Gale, Symposia Soc. Exptl. Biol. 242 (1954). 



5 J. Mandelstam, Intern. Rev. Cytol. 5, 51 (1956). 



6 E. F., Gale, J. Gen. Microbiol. 1, 53 (1947). 



7 G. N. Cohen and H. V. Rickenberg, Compt. rend. acad. sci. 240, 466 (1955). 



8 H. V. Rickenberg, G. N. Cohen, G. Buttin, and J. Monod, Ann. inst. Pasteur 91, 

 829 (1956). 



9 G. N. Cohen and J. Monod, Bacteriol. Revs. 21(3), 169 (1957). 



10 G. N. Cohen and H. V. Rickenberg, Compt. rend. acad. sci. 240, 2086 (1955). 



11 R. J. Britten, R. B. Roberts, and E. F. French, Proc. Natl. Acad. Sci. U. S. 41, 

 863 (1955). 



12 W. R. Sistrom, Biochim. et Biophys. Acta 29, 579 (1958). 



13 H. V. Rickenberg, Biochim. et Biophys. Ada 25, 206 (1957). 



14 J. T. Park, 2nd Intern. Congr. Biochem., Paris p. 31 (1952). 



15 J. L. Strominger, J. Biol. Chem. 224, 509 (1957). 



16 J. L. Strominger and R. H. Threm, Biochim. el Biophys. Acta 33, 280, (1959). 



