DEAN B. COWIK AND RICHARD H. ROBERTS 



15 



A similar comparative experiment was carried out using P'*--!abeled fruc- 

 tose- 1 :()-plu)sphate (2) prepared by a method described by Neuberg, Luslig 

 and Rothenberg (19). A 5-ml suspension of cells in chilled NaCl solution was 

 added to 5 ml of a buffered solution of fructose- 1 :6-phosphate (8.2 mg F-i:6- 

 P per ml). An equal suspension of cells was added to 5 ml radiophosphate 

 solution. Both final solutions were o.i molar with respect to phosphate buffer 

 of pn 7.0. The results are shown in table 11. Approximately equal percentages 

 oi the initial radioactivity of the media were retained by the cells. The two 

 washes revealed that more than 90% of the radioactivity taken up by the cells 

 was readily diffusible into the wash solutions, indicating little metabolic in- 

 corporation. It is concluded that the E. coli cells have a water space volume 

 for glucose- 1 -phosphate and fructose- 1 :6-phosphate equal to that observed for 

 phosphate ions. This water space volume in E. coli (table 6) was 75% of the 



Table 10. Immediate uptake and 



washout of p** labeled phosphate 



and glucose- 1 -phosphate 



% taken up by cells 

 % of total cell radioactivity 

 removed bv first wash 



PO= 



;-7o 



1.4 



Glucose- 

 i-Phos- 

 phate 



5-55 

 73-8 



Table ii. Immediate uptake and 



WASHOUT OF P^ labeled PHOSPHATE 

 AND FRUCTOSE- 1 : 6-PHOSPH ATE 



''(, taken up I)}' cells 

 % of total cell radioactivity 

 removed by 2 washes 



P0= 



7-3 

 95-8 



Fructose- 



i:6-Phos- 



ptiate 



6.2 

 91.9 



total cell volume. Fructose- 1 :6-phosphate is generally postulated to be an 

 early intermediate in glucose metabolism. It fails, however, to support the 

 growth of E. coli cells when supplied as a sole energy source. This failure has 

 often been attributed to the impermeability of the cellular membrane; our 

 measurements indicate that this explanation is not correct. 



Permeability of Torulopsis Utilis to Phosphate. The methods described 

 provide a critical test of an alternative hypothesis for boundary penetration. 

 This hypothesis suggests that certain substances when added to the external 

 environment, first combine specifically with the cell surface and subsequently 

 appear, after some metabolic reaction, unchanged in the internal protoplasm. 

 Such a model has been suggested by Kamen and Spiegleman (13). These 

 investigators conclude that this transport mechanism becomes inactive when 

 sodium azide (2.5 X io~^ m/ 1.) is added to the medium, and that reactions oc- 

 curring on the dell surface are therefore inhibited. If this were actually the case 

 for phosphate transport in T. utilis, the presence of azide in the medium should 

 markedly reduce the apparent water space. 



Table 12 compares the results obtained with cells immersed in a complete 

 medium (21) with and without sodium azide (2.5 X io~^ m/ 1.). Equal volumes 



