AMINO ACID TRANSPORT IN MICROORGANISMS 579 
bound state would not exclude active transport as the mode of entry, so long as the 
concentration of that portion of the pool in the free state still exceeded the extra- 
cellular concentration. 
GALE adopted a cautious attitude when interpreting the results of his extensive 
studies recognizing that the data available did not exclude either of the above- 
mentioned mechanisms*!. Recently he has proposed that the process involves 
active transport** 34. The suggestion that EF. coli is freely permeable to most in- 
organic ions and the usual cell metabolites! led to considerable controversy which 
has greatly increased our understanding of the permeability properties of bacterial 
cell membranes and stimulated a critical examination of the intracellular state of 
accumulated solutes including amino acids. In the original hypothesis amino acids 
in E. coli were believed to be retained on specific adsorption sites located within the 
cell which appeared from space studies to be freely penetrated by a wide variety of 
low molecular weight solutes. Contradicting these findings, MITCHELL observed initially 
that most of the internal volume of S. aureus was not accessible to inorganic phos- 
phate ion’’. It should be noted that while high concentrations of phosphate were 
excluded from the cell in the absence of an energy source, a one for one exchange of 
intra- and extracellular phosphate did occur. This relative aspect of “impermeability” 
is not always appreciated. Inhibition studies showed that this exchange involved 
catalytic rather than stoichiometric adsorption sites. These studies were extended 
by MitcHELL AND MoyLe to show that most of the cell interior of this organism in 
the resting state also was not accessible to various amino acids and inorganic ions, 
thus indicating the occurrence of a permeability barrier near the cell surface*?: 8°. In 
their hands the cell water of F. coli likewise was inaccessible to NaCl and phosphate 
salts*4. Subsequently, a large number of other investigators also have observed 
impermeability of intact bacterial cells, protoplasts and spheroplasts to various 
solutes thereby establishing the widespread existence of a functional permeability 
batter mear the: bactenial:cellssurfacess:39371,-8182, 84, 85; 245,178) 
As indicated above, the state of the intracellular solutes is not established by the 
demonstration of a permeability barrier. It is conceivable that an important fraction 
of the intracellular solutes might still be retained on adsorption sites. Therefore, as 
evidence grew for the existence of effective cytoplasmic membranes in bacteria, 
considerable effort was made to determine whether the intracellular constituents are 
osmotically active, since this would be one way of deciding whether or not they are 
free. The ingenious experiment of MITCHELL AND Moy te**: 8° in which cell pastes 
were exposed to atmospheres above a series of sucrose solutions of graded concen- 
tration and subsequently weighed to determine the variation in water content and 
thus the relative osmotic strength of the cell constituents and the sucrose solution 
indicated that the intracellular osmotic pressure in S. aureus approximately equalled 
that predicted if the total internal solutes extracted by cold TCA were in free solution. 
Unfortunately, this experiment was not sufficiently precise to show the extent to 
which the relatively small portion of the total solute pool made up by amino acids 
was contributing to the observed osmotic activity. 
Another approach has involved measurement of changes in light scattering (in- 
dicative of changes in volume) when cells or protoplasts are exposed to penetrating 
solutes. MITCHELL AND Moy te®?; 84; 85, 89° Avi-Dor ef¢ al.§: 8, GILBY AND FEW? and 
others have observed swelling with some substances although generally not with 
References p. 592/594 
