Chapter 21 Flashcards
Chemiosmotic Coupling
Nobel prize in chem awarded to Peter Mitchell for contribution to understanding of biological energy transfer through formulation of chemiosmotic theory
- He proposed that electron transport and ATP synthesis are coupled by a proton gradient across the inner mitochondrial membrane
Discovery of structure of the large molecular complex responsible for ATP synthesis is considered as significant as discovery of structure of DNA
What is proton motive force?
Proton gradient generated by oxidation of NADH and FADH2 = proton motive force
The proton motive force ia chemical and electrical gradient, or a charged gradient
The proton motive force powers the synthesis of ATP through complex V
- Complex v = membrane protein located in inner mitochondrial membrane that captures the energy contained within proton motive force to make ATP
What are the two components ATP-synthase is made up of? Describe the components
- Fo ~ embedded in the inner mitochondrial membrane, containing the proton channel
- F1 ~ protrudes into the mitochondrial matrix, containing the active sites for ATP-synthesis
Explain/discuss the the two components of ATP-synthase with regards to the alpha, beta, and gamma subunits
- F1 consists of FIVE types of polypeptide chains (a3, b3, gamma, e, s)
- Gamma subunit has unique shape and interactions w/ the 3-beta subunits ~ each beta subunit interacts w/ a different face if the gamma subunit
- 3 alpha and 3 gamma units make up bulk of F1 (arranged in hexametric ring)
- Each beta subunit is an active site
- Alpha subunits isolate the beta subunits from each other
- Protons flow through Fo unit causing it and gamma subunit to rotate
ATP-synthase formation into dimers
- ATP-synthase units will form dimers and cluster together ~ this contributes to formation of the curvatures of cristae of mitochondria
- The clustering helps to stabilize the rotational force and increases the efficiency of the complex
Describe ATP-Synthase F1 subunit with regards to the 3 combinations of the beta subunits
Proton flow through ATP-synthase leads to the release of tightly bound ATP
The binding change mechanism accounts for the synthesis of ATP in response to proton flow
The 3-catalytic beta subunits of the F1 component can exists in 3 different combinations:
- O form (open) ~ nucleotides can bind to or be released from the beta subunits
- L form (loose) ~ nucleotides are trapped in the beta subunits
- T form (tight) ~ ATP is synthesized from ADP plus orthophosphate
ATP Synthesis & Release – F1 Components
- The binding change mechanism for ATP-synthase is shown on right
- In T-form, ATP is formed but there is no release of ATP
- When gamma subunit rotates, the T-form is converted into O-form, which then allows release of ATP
- New molecules of ADP and Pi can bind to the O-form of the subunit
- With an additional rotation, or a 120 degree rotation, the substrates get trapped in L-form
ATP synthase – a small molecular motor
- To better understand and visualize the ATP-synthase as a small molecular motor, researchers cloned the alpha-3 beta-3 gamma subunits, or the F1 subunit, and attached it to a glass slide
- In addition, actin was attached to the gamma subunit such so that movement could be visualized
- There was no Fo-unit to drive the rotation but instead, providing ATP for hydrolysis resulted in rotation of complex
ATP Synthase – Fo Component
- Proton flow through the F0 component powers ATP synthesis
- The F0 component is made up of an alpha-subunit, and 8-c subunits in vertebrates that are shown
- The a-subunit has 2 channels that reach halfway through the unit. One half channel opens to the inner-membrane space and one-half channel opens to the mitochondrial matrix
- Each c-subunit has a glutamate residue with a negative charge
- Protons will enter from the inner membrane space through the a-subunit reach halfway through the channel and then bind to a glutamate residue on c-subunit
- The glutamate w/ negative charge will be converted into glutamic acid w/ the entry of a positively charged proton
- The c-subunit with the proton with no charge will then move into the non-polar fatty acyl region of the phospholipids of the membrane
- This will allow the next c-subunit w/ glutamate and negative charge to rotate into the space adjacent to the a-subunit where protons traveling through the channel will again come into contact w/ the glutamate residue and form glutamic acid
- This will again allow the c-subunit to move or rotate into the fatty acyl region of the phospholipid of the membrane. This will continue and continue and drive the rotation of the F0 unit
- The rotation of the c-ring powers the movement of the gamma subunit, which in turn alters the conformation of the beta subunits
- the beta subunits facilitate ATP-synthesis
- The number of c-rings determines the # of protons required to synthesize a molecule of ATP
ATP Synthase – Fo Component Pt.2 ~ Intermembrane and Matrix
Intermembrane Space: high proton concentration:
- High probability proton will enter channel
and bind to glutamate
- Binding eliminates negative charge and subunit
can move to hydrophobic membrane region
Matrix: low proton concentration:
- Low/no probability proton will enter channel
and bind to glutamate
- Glutamic acid residue picked up in intermembrane half channel eventually pushed into aqueous space of
matrix half channel and proton leaves - Negative charge and subunit cannot move into hydrophobic membrane region – must move forward to intermembrane channel
Proton Motive Force & ATP Synthase
ATP synthase needs 3 protons to flow through to make ATP
1 spin requires 8 protons for 8 c-ring subunits that makes 3 ATP from 3 beta subunits of F1
8 protons/3 ATP = 2.7 = 3
The ATP that is generated will be generated in the mitochondrial matrix
Recall, ATP that has been generated in mitochondrial matrix cannot be utilized by the cell unless its transported out of the mitochondrial matrix
Describe the steps to transport Across Inner Mitochondrial Membrane
- Inner mitochondrial membrane = impermeable; thus several specific protein transporters will be required for exchange of ions or charged molecules between mitochondria and cytoplasm
- A specific translocator facilitates transport of ADP, Pi, and ATP
- ATP-ADP translocase enzyme = antiporter that translocates 1-molecule of ATP out of mitochondrial matrix in exchange for 1-molecule of ADP into mitochondrial matrix
- The phosphate carrier is another antiporter that translocates 1-moleucle of OH- out of mitochondrial matrix in exchange for 1-moelucle of H2PO4- into mitochondrial matrix
- The ATP–ADP translocase, the phosphate carrier, and the ATP synthase are associated with one another to form a large complex, the ATP synthasome
Transport Across Inner Mitochondrial Membrane: Shuttles overview
The NADH that has been generated in the cytoplasm from glycolysis must be re-oxidized into NAD+ ~ in this oxidation, there is a transfer of electron from NADH into the matrix via 2-different shuttle systems
- Glycerol-3-phosphate
- Prominent in muscle cells
- Generates FADH2 in matrix
- Is independent of matrix concentrations of NADH - Malate-aspartate shuttle
- Prominent in heart and liver cells
- Results in generation of matrix NADH
- Is dependent and inhibited by high concentrations of NADH within matrix
Glycerol 3-Phosphate Shuttle
- Cytoplasmic NADH transfers electrons to DHAP to form glycerol 3-phosphate
- Glycerol 3-phosphate transfers electrons to FAD (making FADH2) in membrane bound mitochondrial glycerol 3-phosphate dehydrogenase
- FADH2 then transfers electrons to Q
- QH2 can deliver electrons to
Complex III ~ Bypass Complex I that pumps proton
Malate-Aspartate Shuttle
- Cytoplasmic NADH electrons transferred to oxaloacetate to form malate
–> Malate enters matrix by an antiport that moves α-ketoglutarate out of the matrix
- Malate transfers electrons to NAD+ (making NADH) in matrix becoming oxaloacetate
- An amine group can be added to oxaloacetate to become aspartate that can leave the matrix
–> Aspartate leaves by antiport that brings glutamate into the matrix
- The cytoplasmic aspartate is deaminated and becomes oxaloacetate
–> α-ketoglutarate receives the amine to become glutamate that can return to the matrix
