188 ROLLIN D. HOTCHKISS AND AuDREY H. EvANS 



There would seem to remain four principal classes of hypo- 

 theses which might rationalize this family of three sulphon- 

 amide resistance levels and their various combinations: (l)that 

 there are several independent PAB-using enzyme systems 

 essential for cell growth and three of these can become re- 

 sistant to inhibition; (2) that the affinity of a single enzyme for 

 sulphonamide relative to PAB is being cumulatively reduced 

 by each unit of mutation; (3) that the permeability of the cells 

 to sulphonamide is quantitatively altered as the result of muta- 

 tions; or (4) that an alternative metabolic pathway, which is 

 PAB-sparing, becomes available in each resistant strain. 



Although a priori any one mutant might arise as above, 

 the behaviour of the series of mutants makes some of the 

 hypotheses improbable. It is difficult to understand the 

 independence of the factors a, b, and d if they determine 

 separate and essential systems (hypothesis 1) — how, for ex- 

 ample, could an enzyme Ea, the one altered to give mutant a 

 its resistance to SA, be unchanged in both the comparatively 

 resistant mutant d and the sensitive wild-type strain? Such 

 an independence would seem more in keeping with a series of 

 independent enzymes which provided alternative pathways 

 to the same end-products (hypothesis 4). On the other hand, 

 the cumulative effects of the marker pairs suggest that in 

 their phenotypic effects they are even more co-operative than 

 any independent determinants would be. For example, a 

 (giving a resistance to 10 (xg. SA/ml.) and d (40 (jig./ml.), when 

 recombined by transformation give a strain ad resistant to 

 300 (jLg. SA/ml. It would seem most likely that the co-operat- 

 ing factors a and d are acting upon the same enzyme (hypo- 

 thesis 2) or the same permeability-determining system 

 (hypothesis 3) since they potentiate each other so notably. 



Little is known about factors controlling permeability, but 

 it seems clear that a time rate cannot be the limiting one, 

 since near-infinite time is available as an inhibited cell slows 

 down and stops growing. Furthermore, if a concentration 

 rate is the limiting one for permeability, the properties of a 

 permeability-determining substance which mutates to states 



