Synthesis of Pyruvate Dehydrogenase in Staphylococcus aureus Is Stimulated by Osmotic Stress

Abstract

The pyruvate dehydrogenase multienzyme complex (PDHC) was found to be upregulated by osmotic stress in the osmotolerant pathogen Staphylococcus aureus. Upregulation was detectable in the levels of both activity and protein and was judged to be about fourfold when sodium chloride was used to adjust the water activity (a w ) of the growth medium to 0.94. The upregulation of the PDHC was also found to be humectant dependent and was greatest when impermeant, nonmetabolizable humectants were used to adjust a w . Further experiments provided evidence that in addition to osmotic upregulation, the PDHC complex is also subject to catabolite repression, thus providing a possible explanation for the observation that high concentrations of carbohydrates are generally more inhibitory to the growth of this bacterial pathogen than are high concentrations of salts.Staphylococcus aureus, an important human pathogen, is considered a highly osmotolerant bacterium and is able to grow at a water activity (a w ) of as low as 0.86 (33). Its osmotolerance may be an important determinant of its ability to grow in foods and on human skin, from where it can spread and/or form toxins and cause disease. Although osmoadaptation of S. aureus has been studied fairly extensively in the past (8,9,29,30,38,42,43,46), it remains unclear why this organism is considerably more osmotolerant than other bacterial pathogens. In fact, thus far, studies have revealed an osmoadaptive process apparently typical for bacteria in general: namely, the accumulation of glycine betaine, proline, or other common compatible solutes through transport from the growth medium (25,29,39) and increased synthesis of stress proteins such as chaperones (31) and alkyl hydroperoxide reductase C (8). However, recent studies have given indications that the regulation of stress gene expression may be somewhat unusual in the staphylococci. For example, expression of the alternative sigma factor B is repressed during osmotic stress in S. aureus (13) while it is strongly induced in Bacillus subtilis under those conditions (11,45). Although a few studies have attempted to identify osmotically upregulated proteins in S. aureus (8,31,44), this field still remains relatively unexplored.A major goal of the present study was to investigate changes in protein composition of this osmotolerant organism in response to osmotic stress and to identify proteins important for its osmoadaptation. Herein, we present evidence that the pyruvate dehydrogenase multienzyme complex (PDHC) of S. aureus, an enzyme of central importance in this organism's primary metabolism, is upregulated by osmotic stress. The implications of this finding with respect to the osmoadaptation of S. aureus and the regulation of its metabolic activity are discussed. MATERIALS AND METHODSStrains and media. S. aureus strain ATCC 12600 was obtained from the American Type Culture Collection (Manassas, Va.) and grown in Trypticase soy broth (TSB; Difco Laboratories, Detroit, Mich.) at 37°C. The a w values of media were measured in d...

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