AMINO ACID TRANSPORT IN MICROORGANISMS 589 
Candida utilis also is not readily understood in terms of a single free pool. Indeed, 
this applies to all instances of pool heterogeneity. It should be noted however, that 
much of the evidence for pool heterogeneity comes from experiments with exponen- 
tially growing cells and that the properties of these accumulation systems closely 
resemble those we have found using early exponential phase cells of L. avabinosus. 
It is clear from our studies that cells in different phases of growth contain pools with 
different elution properties. This raises the possibility that the heterogeneity of 
pools cited by CowlE, for example, may be a heterogeneity in the cell population and 
indicative of a difference in the type of pool a given cell accumulates rather than a 
heterogeneity of pools within each of the cells in the population. 
It may also be appropriate to note that considerable evidence exists in addition to 
the osmotic observations which is apparently inconsistent with the binding site 
theory. Thus, the evidence for a bound amino acid pool in EF. coli is strongest when 
properties of small pools are considered. Large pools, on the other hand, are osmoti- 
cally sensitive and show a remarkable lack of specificity, properties which are not 
easily accommodated by the binding site theory. In L. avabinosus loss of as much 
as 25%, of the internal RNA and a substantial amount of protein can occur with 
little or no diminution of glutamate accumulation capacity*®”. Even prior to this loss 
the number of amino acid molecules accumulated exceeds the total number of nu- 
cleotide residues contained in the cell. Except for the adsorption of amino acids to 
cell wall® 1°, 76 there is no direct evidence in broken cell studies for an association of 
sizable amounts of pool amino acids and cell fractions. It is clear that in their purest 
form both theories can be challenged, that much more evidence is required and that 
each case must be examined individually. In view of these inconsistencies, it would 
come as no surprise if a composite mechanism operated in most organisms, 7.e., that 
amino acids are actively transported against real gradients, but that a sizable portion 
of the accumulated “pool” exists in association with labile internal binding sites, or 
as some other sequestered form. 
Nature of the accumulation catalysts 
Most workers, regardless of experimental material utilized observe that accumulation 
involves interaction of the amino acid with some cell component probably having 
catalytic properties. Our extensive studies on the effects of nutritional deficiencies, 
some of which have been described briefly above, indicate that vitamin Bg, biotin 
and pantothenic acid do not fulfill a direct catalytic function in amino acid accumu- 
lation. This is based largely on the observation that in no case is there a significant 
change in the initial rate of accumulation when cells are depleted of these vitamins. 
While all these deficiencies markedly influence the amount of amino acid which can 
be accumulated, these appear to be secondary effects arising from unfavorable 
structural changes in the cell. While one may argue, as we have elsewhere*’, that 
such negative evidence has limited value in excluding these substances from conside- 
ration as integral parts of a catalytic system, the exclusion of vitamin B, as a carrier 
seems most secure now that we have conclusively demonstrated the completely in- 
direct nature of its effect on uptake capacity and shown that normal accumulation 
can be achieved by cells possessing as few as 40 molecules of the vitamin. Although 
evidence continues to appear which suggests some relation between vitamin B, and 
References p. 592/594 
