ATP-linked sodium transport in Streptococcus faecalis. II. Energy coupling in everted membrane vesicles.

Abstract

Everted membrane vesicles of Streptococcus faecalis accumulated "Na+ to a nominal concentration gradient of about 100 when incubated with ATP and M&+.The characteristics of "Na+ accumulation varied in a complex manner as a function of the pH and of the ionic environment.To illustrate, when the vesicles were prepared in potassium ma1eate:Hepes:sucrose buffer (0.45 N K+) and incubated in the same buffer at pH 7.4, uptake of "Na+ was specifically dependent upon ATP and inhibited by N, N"dicyc1ohexylcarbodiimide (DCCD) and by proton conductors; when the same vesicles were incubated at pH 8, uptake of "Na+ was resistant to the inhibitors.When the vesicles were incubated at pH 7.4 in Tris/maleate:sucrose buffer, "Na' uptake was again unaffected by the inhibitors; moreover, ADP and UTP now supported uptake almost as well as did ATP.The results suggest that there are two modes of '*Na+ uptake, one dependent upon the proton-motive force (interior positive and acid) and the other upon ATP.Independent support comes from studies on revertants of mutant 7683, known to lack Na+ transport in both intact cells and membrane vesicles.Revertants that regained this capacity fell into two classes: in one, Na+ movements appear to depend primarily upon the proton-motive force, in the other upon ATP.The present results, as well as those from intact cells (Heefner, D.L., and Harold, F. M. (1980) J. Biot!.Chem 255, 11396-11402) can be rationalized by the hypothesis that S. faecalis contains two transport systems for sodium: one a primary, ATP-driven sodium pump and the other a secondary porter linked to the proton circulation but activated by ATP.However, we consider it more likely that these are two modes of sodium transport mediated by a single, ATP-linked system.The experiments described in the preceding paper (Heefner and Harold, 1980) led us to conclude that sodium movements in Streptococcus faecalis cannot be ascribed to a secondary Na'/H' antiporter.We proposed instead that the cells establish a circulation of sodium across the membrane: sodium enters the cells down the electrochemical potential gradient by a route of low affinity, probably a relatively nonspecific "leak."Sodiumextrusion is mediated by a distinct transport system that is selective for Na' and requires "ATP" (either