PROLINE TRANSPORT IN STAPHYLOCOCCUS-AUREUS - A HIGH-AFFINITY SYSTEM AND A LOW-AFFINITY SYSTEM INVOLVED IN OSMOREGULATION

L-Proline enhanced the growth of Staphylococcus aureus in high-osmotic-strength medium, i.e., it acted as an osmoprotectant. Study of the kinetics of L-[C-14]proline uptake by S. aureus NCTC 8325 revealed high-affinity (K(m) = 1.7-mu-M; maximum rate of transport [V(max)] = 1.1 nmol/min/mg [dry weight]) and low-affinity (K(m) = 132-mu-M; V(max) = 22 nmol/min/mg [dry weight]) transport systems. Both systems were present in a proline prototrophic variant grown in the absence of proline, although the V(max) of the high-affinity system was three to five times higher than that of the high-affinity system in strain 8325. Both systems were dependent on Na+ for activity, and the high-affinity system was stimulated by lower concentrations of Na+ more than the low-affinity system. The proline transport activity of the low-affinity system was stimulated by increased osmotic strength. The high-affinity system was highly specific for L-proline, whereas the low-affinity system showed a broader substrate specificity. Glycine betaine did not compete with proline for uptake through either system. Inhibitor studies confirmed that proline uptake occurred via Na+ -dependent systems and suggested the involvement of the proton motive force in creating an Na+ gradient. Hyperosmotic stress (upshock) of growing cultures led to a rapid and large uptake of L-[C-14]proline that was not dependent on new protein synthesis. It is suggested that the low-affinity system is involved in adjusting to increased environmental osmolarity and that the high-affinity system may be involved in scavenging low concentrations of proline.