Lecture 10 - Bacterial ATP synthesis and how it became the newest target-space for antibiotics Flashcards
Terminal electron acceptors are used by
availability and energy gain e.g. oxygen -> nitrate -> fumarate - as determined by redox potentials
Cytochrome bd oxidase =
Cytochrome bd oxidase = high affinity for oxygen but less PMF
Nitrate =
Nitrate = used by nitrate reductase, redox loop generates PMF
Fumarate =
Fumarate = used by fumigate reductase, no PMF is made (smaller reduction potential difference, does not do the redox loop)
Other roles of PMF
Redox balance, radical detoxification, enzyme protections
In this case, PMF back pressure promotes non proton pumping configurations
Central metabolism is reconfigured in analogous ways
TCA rerouting, reverse TCA, TCA half cycles or no/limited TCA encoded
Fumarate reductase is the ….
least efficient enzyme at generating a proton motive force
Proton motive force
An electrochemical gradient
Pump protons and make our periplasmic space more acidic - pH gradient of protons that has energy stored in it since they want to diffuse back but they are prevented by the membrane
Energy stored in a build up of H+ and other ions in the periplasm
Drives many transporters. Uptake carbon/energy sources
Required for high efficiency ATP synthesis
Even fermenters need a PMF
Some bacteria use sodium ions instead of protons (they need specialised proteins to do this)
Back pressure
Cellular respiration and ATP synthesis occur in concert - tightly coupled
Back pressure = overload of protons in the periplasmic space, can feedback and inhibit proton pumps and this is why bacteria encode some non-protein pumping enzymes but normally under homeostatic conditions we have a fine balance between putting a lot of PMF that is enough to drive ATP synthesis as well as pumping it at the same time
Catabolism:anabolism and proton backpressure govern…
Catabolism:anabolism and proton backpressure govern all steps:
Central carbon metabolism
ETC activity
ATP synthesis
If these links are broke for any reason the cell is called an uncoupled cell
Cell exhausts energy currencies to try and restore PMF
F1F0 ATP synthase
2 major components
F1F0 ATP synthases found in mitochondria, chloroplasts, and bacteria. They are also known as F1F0 ATPases because they can catalyze ATP hydrolysis. The mitochondrial F1 component appears as a spheri- cal structure attached to the mitochondrial inner membrane surface by a stalk. The F0 component is embedded in the membrane. ATP synthase is on the inner surface of the plasma membrane in bacterial cells. F0 participates in proton movement across the membrane. F1 is a large complex in which three a subunits alternate with three b subunits. The catalytic sites for ATP synthesis are located on the b subunits. At the center of F1 is the g subunit. The g subunit extends through F1 and interacts with F0.
ATP synthase as the method from both utilising protons as well as producing the ATP and ATP feedsback into catabolic processes and these catabolic and anabolic processes make NADH…
Chemomechanical coupling
F1 subunit
Cytoplasmic
Binds ADP + Pi and converts to ATP - forms the phosphate high energy bond required to make ATP
Not in the membrane
Soluble fraction - can be knocked off and be soluble in the cytoplasm i.e. can be dissolved
F0 subunit
Membrane bound region
Motor
Able to rotate
Turbine is driven by H+ flow
Rotates central stalk in F1
Flow rotates the subunits and the rotation of the central stalk actually causes conformational changes which is called chemomechanical coupling because there is chemical energy from the proton motive force and then being transduced to mechanical energy in the stalk
Key subunits of ATP synthase
Catalytic subunits …
β - Binds ADP+Phosphate—> ATP
𝛾 - Rotates, induces conformational change
Ion binding subunits …
a-Take H+ and passes to c subunit
c-Rotated and releases H+ in cytoplasm
C-subunit has the proton and rotates it before releasing it into the cytoplasm
Other subunits have roles in regulation or complex stability
Catalytic subunits of ATP synthase
β - Binds ADP+Phosphate—> ATP
𝛾 - Rotates, induces conformational change
Ion binding subunits of ATP synthase
a-Take H+ and passes to c subunit
c-Rotated and releases H+ in cytoplasm
C-subunit has the proton and rotates it before releasing it into the cytoplasm
Other subunits of ATP synthase
Other subunits have roles in regulation or complex stability
Three major conformations of ATP synthase
Three major conformations: Open = Nothing bound Loose = ADP+Pi bound Tight = closed, forms ATP (when the stalk rotates it is closed) Three major steps in rotation Open - loose - tight - open etc 3 x 120 degree steps
Open configuration
Nothing bound
Loose configuration
ADP+Pi bound
Tight configuration
Closed, forms ATP (when the stalk rotates it is closed)
Binding change mechanism
Three major conformations:
Open = Nothing bound
Loose = ADP+Pi bound
Tight = closed, forms ATP (when the stalk rotates it is closed)
Three major steps in rotation
Open - loose - tight - open etc
3 x 120 degree steps
May be 6 or more subsets
May vary between organisms
Conformational change that causes something to happen but what is unique about this is that it is driven by its mechanical process
Textbooks = 3.3 H+ to 1 ATP made Reason = mitochrondrial enzyme has 10 c-subunits = 10H+ (how many protons you can bind per rotation) for full rotation = 3 ATP from 10 H+ However = Different organisms have different amounts of c-subunits. Anywhere from 2.7-5 H+ per ATP
Sodium motive force
Advantageous when passive proton leak/loss is high - some anaerobes
High temperature (makes plasma membrane more fluid and can cause things to passively diffuse more efficiently) or pH (alkaline environment)
e.g. Clostridium paradoxum
Group of organisms that tends to do this and might find that some organisms have this advantage to grow in high temperatures or alkaline environments by switching to a sodium motive force
Used by some pathogens
Fusobacterium nucleate: inhabits gums, periodontal disease
SMF and PMF at same time to get the benefits of both worlds
(Partially) vibrio cholera: cholera, free-living phase -> pH stress
