bovis BCG and M smegmatis is blocked in the ATP hydrolysis mode

bovis BCG and M. smegmatis is blocked in the ATP hydrolysis mode and is not able to generate a PMF by hydrolyzing ATP. The essentiality of ATP synthase is thus based on a function in the synthesis direction, GSK J4 research buy most likely either for the production of ATP, pH homeostasis, or for contributing to the NAD+/NADH redox balance. The task of PMF maintenance under low oxygen tensions is most probably fulfilled by other membrane–protein complexes, such as by nitrate reductase or by fumarate reductase acting in reverse (Schnorpfeil et al., 2001; Wayne & Sohaskey, 2001). In order to gain an insight into the mechanism of ATP hydrolysis blockage

in mycobacteria, we tested the effect of four different treatments reported to activate ‘latent’ ATP hydrolysis activity in bacteria. Limited trypsin proteolysis is reported to cleave the inhibitory intrinsic subunit ɛ and in this way activate ATP hydrolysis (Bogin et al., 1970; Keis et al., 2006), while the addition of methanol is thought to compromise hydrophobic interactions within ATP synthase (Hisabori et al., 1997). Moreover, oxy-anions,

for example sulfite, are reported to remove inhibitory ADP and to uncouple ATP synthase function (Bakels et al., 1994; Cappellini et al., 1997; Pacheco-Moisés et al., 2002). Finally, membrane energization is known to relieve ADP inhibition and to switch the conformation of subunit ɛ to a noninhibitory Selleck Trichostatin A state (Suzuki et al., 2003). The ATP hydrolysis activity of IMVs of M. smegmatis was indeed activated >30-fold by trypsin (Table 2), indicating that subunit ɛ is an important determinant for ATP hydrolysis blockage in this fast-grower. However, in the case of M. bovis BCG, trypsin treatment did not lead to significant activation (Table 2). This lack of activation can be explained either by

inaccessibility of the trypsin cleavage site or by the presence of alternative inhibitory mechanisms. To further investigate ATP hydrolysis in M. bovis BCG, we tested the effect of methanol, sodium sulfite and PMF activation. Whereas sulfite slightly activated ATP hydrolysis activity, both addition of methanol and membrane energization by succinate led to more significant activation for M. bovis BCG, with the resulting activity ∼10-fold higher than the ATP synthesis activity (Table 2). Dipeptidyl peptidase The results suggest that ATP hydrolysis in both slow-growing as well as fast-growing mycobacteria is regulated in a PMF-dependent manner, preventing excess ATP consumption under low oxygen tensions. Suppression of activity appears to be more pronounced in the slow-grower, which may be an adaptation to environments with a low energy supply and/or decreased oxygen tensions, for example in remote parts of the mammalian lungs. mycobacteria, requiring oxygen for growth, but able to persist under anaerobic conditions, thus utilize a similar mechanism of ATP hydrolysis inhibition as reported for the obligate aerobic bacteria P.

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