Effects of Ca2+ and a protonophore on growth of an Escherichia coli L-form

The influence of Ca2+ ions on the growth of an L-form (NC7) derived from Escherichia coli K12 was investigated. In a medium containing NaCl as osmotic stabilizer 1 mM-Ca2+ was required for optimal growth of the L-form, while with KCl as osmotic stabilizer, in a medium containing 0.1 or 1.0 mM-Ca2+, optimium growth was observed at 32 and 37 degrees C, respectively. When the L-form, growing exponentially at 32 degrees C in medium containing KCl and 0.1 mM-Ca2+, was shifted to 37 degrees C growth was strikingly suppressed. In contrast, the suppression of growth in the presence of 1.0 mM-Ca2+ at 32 degrees C was relieved when the culture was shifted to 37 degrees C. When the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP), at a final concentration of 10 microM, was added to a medium containing NaCl and sucrose as osmotic stabilizers, together with 10 mM-glucose, the parent strain could grow exponentially. In contrast, growth of the L-form was completely stopped by 10 microM-CCCP under the same conditions. In the presence of 20 microM-CCCP, the L-form accumulated more than twice as much 45Ca as in the absence of the protonophore. Thus, it is suggested that growth of the L-form NC7 is coupled to the protonmotive force. Possible mechanisms for the coupling of calcium to growth of the L-form are discussed.