610 
INVITED DISCUSSION 
SOME PROPERTIES AND POTENTIAL USES OF BACTERIAL 
MUTANTS DEFECTIVE IN AMINO ACID TRANSPORT 
MARTIN LUBIN 
Department of Pharmacology, Harvard Medical School, Cambridge, Mass. (U.S.A.) 
The study of active transport is vexing. In spite of considerable work on transport 
of sugars and amino acids in a variety of cell types, there remains in doubt no less 
than the entire sequence of biochemical reactions. The availability of suitable 
bacterial mutants has enlarged our information on /-galactoside transport, and the 
recent isolation of a series of mutants defective in amino acid transport provides 
hope for a fresh approach to the problem of amino acid accumulation. 
Of the (E.coli) mutants we have isolated!, defective in the transport of either 
histidine, glycine, or proline, only the last has as yet received extensive study in our 
laboratory. The transport-defective mutant (Tr-,,.) was derived from a parent strain 
blocked in the biosynthesis of proline, and has the property of requiring a high proline 
supplement in the medium (250-500 ug/ml) for rapid growth. Using [!C|proline, 
KessEL? has found that in the Tr-,,, strain, the free pool of proline equilibrates 
with the medium without evidence of a concentrating mechanism. The relative 
rates of leak of proline from parent strain (Tr+,,.) and mutant strain (Tr,,.) have 
been measured both at 37° and o°, and found to be similar, at a given temperature, 
for both strains. This leads to the unequivocal conclusion that the defect in the 
mutant lies in the system responsible for active uptake. 
A surprising observation was made when exchange of proline at 0° was studied. 
It should be recalled that Botton et al.? reported very rapid exchange of labeled 
for unlabeled proline at 0° in the absence of net transport. KESSEL has confirmed 
this for the parent (Tr*,,,) strain, but finds this exchange is nearly absent in the 
Tro mutant, although exchange of histidine or glycine is unimpaired. This 
result is consistent with the notion that the mutant has lost a catalytic site or 
a shuttle-system involved both in exchange and active transport, the latter pro- 
cess requiring coupling to an energy source. It will be of interest to determine, 
by examining other transport-defective mutants, including those of the /-galacto- 
side system, how often loss of transport by mutation is accompanied by loss of ex- 
change. 
The second feature of interest which has emerged from our study concerns the 
physiological localization of the transport process. The result was not surprising, 
but the experiment had not previously been done. Back-mutants with the ability to 
synthesize proline were selected on minimal plates. The Tr+,,, and Tr-,,. properties 
were identified by the extreme difference in the ability to concentrate [14C]proline. 
Both back-mutants, however, grew at nearly identical rates in minimal medium. The 
conclusion we have reached is that the defect in this Tr-,,, mutant is not accom- 
References p. 611 
