Abstract
A mutant of Streptococcus faecalis has been isolated which has a 20-fold higher growth requirement for l -glutamic acid than the parent strain. The ability to utilize l -glutamine for growth is not affected. The mutant also has a greater dependence than the parent strain on an external supply of vitamin B6. The rate of glutamic acid transport at low extracellular concentrations (0.05 m m ) was greatly reduced in the mutant but was only moderately affected at higher concentrations (5 m m ). In contrast to the parent, in which kinetic studies revealed the operation of a high affinity and a low affinity system for glutamic acid transport, the mutant possessed a single catalytic component with kinetic constants equal to those of the low affinity system. These findings and the greatly elevated growth requirement for glutamic acid indicate that the high affinity system is inactive in the mutant. The transport of aspartic acid and d -glutamic acid by the mutant were similarly adversely affected, whereas essentially identical kinetic constants for glutamine, α-aminoisobutyric acid, and cycloleucine transport were obtained for the mutant and parent strains. The absence of a functional high affinity dicarboxylic amino acid transport system in the mutant allowed a study of the specific properties of the low affinity system. In contrast to the parent strain, the initial rate of uptake of isotopically labeled glutamate in the mutant was not stimulated by glucose at any concentration. However, the accumulation of large amounts of this amino acid on extended incubation was dependent on an energy source. Glutamine competitively inhibited glutamic acid uptake in the mutant. There was no evidence that the mutant produced an inhibitor of the high affinity transport component.
Highlights
A mutant of Sfrepfococcus faecalis has been isolated which has a ZO-fold higher growth requirement for L-glutamic acid than the parent strain
The penicillin treatment was repeated three times and the cells obtained after overnight growth on high glutamate medium were diluted and plated on high glutamate medium solidified with 1.5 y0 washed agar
Half-maximal growth of the mutant in contrast was not attained unless the medium contained approximately 750 pg per ml (5.1 mM) of L-glutamic acid (Fig. 1A)
Summary
The procedures used for maintaining and growing cultures of the parent strain, S. faecalisR (ATCC 8043), designated ATCC, preparing cells for uptake experiments, and carrying out and evaluating the results of transport measurements are described in Reference 1.Mutant strains were obtained by ultraviolet irradiation of a heavily inoculated exponential phase culture grown at 30” for 90 min in the regular synthetic medium. 1.8 ml was used to inoculate 50 ml of a high glutamate medium containing 12.5 mg per ml of n-glutamic acid, 3.5 mg per ml of L-glutamine, and 50 pg per ml of n-aspartic acid. The cells were washed in growth medium lacking glutamic acid, aspartic acid, and glutamine and were used to inoculate (0.05 mg per ml) two flasks of a low glutamate medium containing 20 pg per ml of n-glutamic acid and 1 pg per ml of n-aspartic acid. After washing twice with growth medium lacking glutamate and aspartate, the samples from the penicillin culture were used to inoculate overnight cultures in high glutamate medium. The penicillin treatment was repeated three times and the cells obtained after overnight growth on high glutamate medium were diluted and plated on high glutamate medium solidified with 1.5 y0 washed agar. After 3 days of incubation these plates were replicated to agar medium
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
