On the relationship between the nonlinear dielectric properties and respiratory activity of the obligately aerobic bacterium Micrococcus luteus

We have used our previously-described, four-terminal, nonlinear dielectric spectrometer to study the generation of nonlinear dielectricity by the obligately aerobic, heterotrophic, respiratory bacterium Micrococcus luteus. Intact cells of M. luteus generate harmonica when excited with single sinusoids of appropriate voltage and frequency. Two frequency windows are observed, respectively below (“low frequencies”) and above (“high frequencies”) ca 100 Hz. The low-frequency region shows a voltage window of some +0.4-1.6 V (zero-to-peak, as judged on the outer electrodes). Anaerobic (non-respiring) cells generate odd-numbered harmonics, whilst aerobic (respiring) cells generate even-numbered harmonics. In the high-frequency range, intact cells of M. luteus (and also of Saccharomyces cereuisiae) generate both oddand even-numbered harmonics, whether aerobic or anaerobic. In the low-frequency range, the inhibition of respiration by cyanide or by HQNO (2-n-heptyl-4-hydroxyquinoline-N-oxide) causes the appearance of odd-numbered harmonics in aerobic cell suspensions. Higher concentrations of cyanide (but not of HQNO) cause the disappearance of all harmonics. These facts are consistent with the known effects of these inhibitors on electron transport in the branched respiratory chain of this organism, and suggest that the major source of nonlinear dielectricity within the respiratory chain is either in the branch to cytochrome &ss or proximal to the site at which HQNO binds. Membrane vesicles of M. luteus generate harmonics with patterns similar to those found in intact cells. In the low-frequency region, the addition of NADH to the membrane vesicles causes the replacement of odd-numbered harmonics by even-numbered harmonics, providing extremely clear-cut evidence that the type of harmonics observed depends strongly on whether or not the cells are respiring. In the high-frequency region, both oddand even-numbered harmonics are generated by membrane vesicles, whether or not the membranes are carrying out electron transport. Liposomes consisting of pure phospholipids show similar patterns of harmonics, suggesting that the high-frequency nonlinear dielectricity may be ascribed to the lipid portion of the membranes of cells and vesicles. By contrast, the expression of nonlinear dielectricity at low exciting frequencies is determined by the respiratory system, and whether or not it is active. This phenomenon provides a novel method for the measurement of respiratory activity in situ. * To whom correspondence should be addressed, 0302-4598/91/$03.50