Related Experiment Videos
Interaction of vanadate with membrane-bound ATPase from Mycobacterium phlei
Abstract:
Vanadate inhibited the formation of proton gradient and membrane potential as well as Ca2+ transport by everted membrane vesicles from Mycobacterium phlei, with half-maximal inhibition occurring at 5 to 14 microM. That this is due to the inhibition of the proton-translocating ATPase was suggested by the observation that the inhibition described above occurred only when the processes were driven by the hydrolysis of ATP but not when energized by the oxidation of succinate and NADH. Furthermore, vanadate did indeed inhibit ATP hydrolysis by these membrane vesicles. Although the inhibition of ATP hydrolysis could be demonstrated only in the presence of high concentrations (e.g. 11 mM) of Mg2+, this was presumably due to the fact that we were measuring the sum of ATP hydrolysis by both coupled and partially uncoupled enzymes. This is the first reported effect of vanadate on bacterial proton-translocating ATPase.
Insights
Vanadate inhibits proton-translocating ATPase in Mycobacterium phlei, affecting proton gradients, membrane potential, and calcium transport. This study is the first to report vanadate
Area of Science:
- Biochemistry
- Microbiology
- Cellular Physiology
Background:
- Proton gradients and membrane potential are crucial for cellular energy production.
- Bacterial ATPases play a vital role in maintaining these gradients.
- Understanding inhibitors of these processes is key to studying bacterial energy metabolism.
Purpose of the Study:
- To investigate the effect of vanadate on membrane-associated processes in Mycobacterium phlei.
- To determine if vanadate inhibits the proton-translocating ATPase of Mycobacterium phlei.
- To characterize the mechanism of vanadate inhibition.
Main Methods:
- Utilized everted membrane vesicles from Mycobacterium phlei.
- Measured proton gradient and membrane potential formation.
- Assayed Ca2+ transport and ATP hydrolysis.
- Tested energy coupling with ATP hydrolysis versus succinate/NADH oxidation.
Main Results:
- Vanadate inhibited proton gradient, membrane potential, and Ca2+ transport.
- Inhibition occurred specifically when processes were ATP-driven, not succinate/NADH-driven.
- Vanadate inhibited ATP hydrolysis by the membrane vesicles.
- Half-maximal inhibition occurred at 5-14 microM vanadate.
Conclusions:
- Vanadate inhibits the proton-translocating ATPase in Mycobacterium phlei.
- This inhibition affects key energy-dependent membrane functions.
- This is the first report of vanadate impacting bacterial proton-translocating ATPase.