DEAN B. COWIF. AND RICHARD B. ROBERTS 



13 



tinguish between metabolic and nonmetabolic phosphorus within the cell. 

 Some metabolic binding of phosphate can occur without growth even at low 

 temperatures, and in the absence of exogenous glucose and ammonium salts. 

 Nevertheless, the criteria and methods employed in the study of sulfate pene- 

 tration are also applicable to the investigation of phosphate transport. 



Cells grown overnight in low-glucose synthetic medium, after harvesting 

 and washing in saline, were added to chilled culture tubes containing glucose 

 and various quantities of P^--labeled sodium phosphate buflfer. The P'- solu- 



A. Immediate Uptake] of P^ 



B. Uptake of P'^ after 40 Minutes' ImmersionX 



* Calculated on the basis of 75% of total cell volume for water space. 

 t Cultures maintained at 4°C. 

 X Cultures maintained at 37°C. 



tion was hydrolyzed for 30 minutes with i N HCl at ioo°C to insure that all 

 the phosphorus was phosphate (14). One set of tubes also contained ammonium 

 salts. The cells were immediately centrifuged and washed. Aliquots of the 

 washed cells and wash solutions were measured for radioactivity. 



From the results shown in table 8A it is apparent that the radioactivity 

 removed from the cells by washing is proportional to the phosphate concen- 

 tration of the original medium. In contrast, the quantity of phosphorus bound 

 depends less on the phosphorus concentration of the medium. Furthermore, 

 this wash loss is equal to the radioactivity which would be expected to dififuse 

 into the water space of the cells, assuming complete permeability of the cellular 

 membrane to phosphate ions. This water space volume is observed to be the 

 same for phosphate and sulfate ions in E. coli. No effect of the NH4 ions was 

 observed. 



