Chapter 5 - Part 1 PDF

Why Mitochondrion? July 9, 2020 Impaired mitochondrial recycling drives neurons death in Parkinson’s study (Sep 4,2020) 69 Mitochondrial Structure :How do they look like? Mitochondria: characteristic morphologies despite variable appearance. Typical mitochondria are bean-shaped organelles but m...

Why Mitochondrion? July 9, 2020 Impaired mitochondrial recycling drives neurons death in Parkinson’s study (Sep 4,2020) 69 Mitochondrial Structure :How do they look like? Mitochondria: characteristic morphologies despite variable appearance. Typical mitochondria are bean-shaped organelles but may be round or threadlike. Size and number of mitochondria reflect the energy requirements of the cell. Fig 5.1 Elongated mitochondria of fibroblast Transmission electron micrograph Mitochondria in the sperm mid-piece (50-75 mitochondria in a sperm cell)-why so many- needs 70 energy to reach the egg Mitochondrial Structure • Mitochondria can fuse with one another, or split in two. Fig 5.2 b 3D model of contacts between ER and mitochondria – The balance between fusion and fission is likely a major determinant of mitochondrial number, length, and degree of interconnection. 71 Mitochondrial Structure: Mitochondrial Membranes • The outer boundary of a mitochondrion contains two membranes: – The outer mitochondrial membrane and – The inner mitochondrial membrane • The outer mitochondrial (50 % protein) membrane serves as its outer boundary Fig 5.3a-Scanning electron micrograph of a macerated mitochondrion The outer membrane contains a large pore-forming protein called porin. The inner mitochondrial membrane (75 % protein) is divided into two major domains that have different proteins and carry out distinct functions : Inner boundary membrane Cristae: where the machinery for ATP is located The inner membrane contains cardiolipin but not cholesterol (just like 72 bacterial membranes) Mitochondrial Structure: Mitochondrial Membranes – The inner boundary membrane domain, along with the outer membrane, forms a double-membrane outer envelope • The other domain lies in the interior of the organelle as a series of invaginated membranous sheets (large surface area), called cristae, which houses the machinery needed for aerobic respiration and ATP formation • The inner boundary membrane and internal cristal membranes are joined to one another by cristae junctions Mitochondria contain 37 genes and all are essential for normal mitochondrial function while 13 genes are involved in oxidative phosphorylation Fig 5.3c. Schematic diagrams showing the 3D internal structure and a thin section of a mitochondrion from bovine heart tissue 73 Mitochondrial Structure: Mitochondrial Membranes • The membranes of the mitochondrion divide the organelle into two aqueous compartments: • One within the interior of the mitochondrion, called the matrix, with gel-like consistency Image taken from :https://www.quia.com/files/quia/users/lmcgee/Biology-Learning-Center-upload-10-2105/cell_processes/cellular_respiration/assets/graphics/cells_and_parts/mitochondrialabeled1.gif • A second between the outer and inner membrane, called the intermembrane space • The inner membrane is impermeable to even small molecules, virtually all molecules and ions require special membrane transporters to gain entrance to the matrix. 74 Oxidative Metabolism in the Mitochondrion An overview Glycolysis • The first steps in oxidative metabolism are carried out in glycolysis in the cytosol; it is an anaerobic process (9/10 steps not use oxygen) • Glycolysis produces two pyruvate, two NADH, and two (net) molecules of ATP per glucose – Most of the energy remains in pyruvate! • Aerobic organisms use O2 to extract more ATPs from pyruvate and NADH • The ATP molecules are produced via substrate-level phosphorylation here 75 Oxidative Metabolism in the Mitochondrion An overview Glycolysis Remember only main things no structure To produce two Pyruvate molecules: ATP produced= 2x2=4 ATP used=2 Net gain of ATP=4-2=2 76 An overview of Glycolysis Key Steps Step 1: Phosphorylation of glucose – Used 1 ATP Step 2: Isomerization of Glucose-6-phosphate – converting an aldose into a ketose sugar Step 3: Phosphorylation of fructose- 6-phosphate – Used 1 ATP Step 4: Cleavage of fructose 1,6 diphosphate - form: glyceraldehyde 3-phosphate (an aldose) and dihydroxyacetone phosphate (a ketose) Step 5: Isomerization of dihydroxyacetone phosphate (DHAP) into glyceraldehyde 3-phosphate Step 6: Oxidative phosphorylation of glyceraldehyde 3-phosphate molecules to form 1,3- bisphosphoglycerate – used inorganic phosphate and NAD+ is reduced to coenzyme NADH by the H+ ions from glyceraldehyde 3-phosphate Step 7: Transfer of phosphate from 1,3- bisphosphoglycerate to ADP- ATP formed Step 8: Isomerization of 3-phosphoglycerate into 2-phosphoglycerate (shift of phosphoryl group from C3 to C2) 77 Step 9: Dehydration of 2-phosphoglcerate to form phosphoenolpyruvate Acetyl CoA Formation from Pyruvate What did you notice? First CO2 is removed from the original glucose molecule Pyruvate is transported across the inner membrane (into the matrix) and decarboxylated to form acetyl CoA, which enters the next stage Thioester bond Joining to CoA results in acetyl CoA by thioester link. It is a high-energy bond and very unstable. 78 Q) How Acetyl CoA forms from Pyruvate? Pyruvate formed is transported into the into matrix by oxidative decarboxylation of pyruvate and form acetyl CoA (electrons are transferred to NAD+ to NADH)- used later to generate ATP Q) What is the end product of glycolysis? 2 pyruvate, 2 NADH and 2 ATP molecules 79 Oxidative Metabolism in the Mitochondrion: The TCA Cycle Remember only main things no structure like NADH and ATP no produced For two pyruvate molecules: CO2 produced=6 NADH produced= 8x3=24 FADH2 produced=2x2=4 GTP= 2 Total= 30 ATP For one pyruvate molecule: CO2 produced=3 NADH produced= 4 FADH2 produced=1 GTP= 1 8.Dehydrogenation 1.Condensation 7.Hydration 2.Dehydration/Rehydration 5 paors of elctrons (from hydrogen atoms of substrate) to be used In ATP production 3. Oxidative decarboxylation 6.Dehydration 5. Substrate level phosphorylation 4.Oxidative decarboxylation 80 Citric acid cycle (Krebs cycle or TCA cycle) succinate dehydrogenase, which is bound to the inner membrane, all the enzymes of the TCA cycle reside in the soluble phase of the matrix Four reactions in the cycle transfer a pair of electrons to NAD+ to form NADH, or to FAD to form FADH2. Some steps are combine in the next slide 81 Q) How many enzymes catalyzes the TCA cycle? Can you name them? 1.Citrate synthase 2.Aconitase 3.Isocitrate dehydrogenase 4. -ketoglutarate dehydrogenase 5.Succinyl-CoA synthetase 6.Succinate dehydrogenase 7.Fumarase 8.Malate dehydrogenase 83 Q) How many total carbons from an original glucose molecule will enter into the TCA cycle? a) 2 b) 3 c) 4 d) 6 Q) How many total carbons from an original glucose molecule will enter into the TCA cycle in the absence of oxygen? a) 2 b) 3 c) 4 d) 0 84 Recall or Summary succinyl-CoA is hydrolyzed with release of free energy. That energy is conserved in the substrate phosphorylation of GDP with phosphate and form GTP Why GTP ? 5. Substrate level phosphorylation Where do you see GTP hydrolysis and ATP hydrolysis? GTP hydrolysis is common in signal transduction, whereas ATP hydrolysis is used for force generation or other energy requiring process. What happened with the lactate produced in the body? Under limited supply of oxygen body temporarily converts pyruvate into a substance called lactate that convert into pyruvate and produce glucose by a process called gluconeogenesis (formation of glucose from non-carbohydrate sources is called gluconeogenesis) 85 Oxidative Metabolism in the Mitochondrion What happens to NADH molecules produced during glycolysis? How NADH imports into the mitochondrial matrix from cytosol? • Mitochondria are not able to import the NADH formed in the cytosol during glycolysis ( 2 NADH/glucose molecule) • There are two main ways that cells can make use of the NADH molecules produced in the cytosol: 1. Malate-Aspartate Shuttle 2. Glycerol Phosphate Shuttle 86 Summary Slide 1. Malate-Aspartate Shuttle MAD Commute Malate in. Alpha-ketoglutarate and D(Aspartate) out. 2. Glycerol Phosphate Shuttle GLYCOLSIS- Step 5: Isomerization of dihydroxyacetone phosphate (DHAP) into glyceraldehyde 3phpsphate https://www.ncbi.nlm.nih.gov/books/NBK22470/ 87 Oxidative Metabolism in the Mitochondrion Malate-aspartame shuttle Malate dehydrogenase is present in two forms in the shuttle system: mitochondrial malate dehydrogenase and cytosolic malate dehydrogenase. 1. In cytosol, malate dehydrogenase catalyzes the reaction of oxaloacetate and produces malate (require 2 electrons and H+ to form malate) – NADH is oxidized to NAD+ in cytosol and now can enter into the mitochondrial matrix. 2. Now antiporter imports the malate from cytosol and exports alpha-ketoglutarate from the matrix into the cytosol simultaneously. 3. Once Malate reaches the mitochondrial matrix, Mitochondrial malate dehydrogenase catalyzes the reaction into oxaloacetate (NAD+ is reduced with two electrons to form NADH). 4. Oxaloacetate is then transformed into aspartate by mitochondrial aspartame aminotransferase and transport into the cytosol. 5. Aspartame needs an amino radical to form Is it symport or antiporter? 88 Oxidative Metabolism in the Mitochondrion Malate-aspartame shuttle (summary) OUT Oxaloacetate NADH Glutama te A D Aspartate or Alpha ketoglutarate NAD+ IN M NAD+ Malate NADH Oxaloacetate 89 Oxidative Metabolism in the Mitochondrion The Glycerol phosphate shuttle • The reduced coenzymes FADH2 and NADH are the primary products of the TCA cycle • Mitochondria are not able to import the NADH formed in the cytosol during glycolysis ( 2 NADH/glucose molecule) • It can be done by cytosolic glycerol 3-phosphate dehydrogenase and mitochondrial 3- phosphate dehydrogenase enzymes. 1. Electrons are transferred from NADH to dihydroxyacetone phosphate (DHAP) to form glycerol 3-phosphate (G3P). – DHAP + 2e- + H+ G3P – G3P moves into intermembrane space and gets oxidized by G3P dehydrogenase, reverting back to DHAP – G3P dehydrogenase reduces FAD into FADH2 by taking two electrons 2. These electrons are indirectly fed into the mitochondrial NADH formedchain during glycolysis enters the electron-transport and used for ATP formation mitochondria via the glycerol phosphate shuttle Fig 5.9: Electron transfer from NADH to DHAP to form glycerol 3phosphate, then to FAD to form FADH2 90 Oxidative Metabolism in the Mitochondrion Summary of oxidative phosphorylation The Importance of Reduced Coenzyme in the Formation of ATP in the Electron Transport Chain • Step 1:High-energy electrons from NADH and FADH2 are passed to electron carriers of ETC, energy is released, which is coupled to energy-required conformational changes, leading to H+ being pumped out across the inner membrane • Step 2: ATP is formed by the controlled movement of H+ back across the membrane through the ATP-synthesizing enzyme Inner membrane Inter membrane space – Coupling of H+ translocation to ATP synthesis is called chemiosmosis • Tally: – 3 molecules of ATP are formed from each pair of electrons donated by NADH – 2 molecules of ATP are formed from each pair of electrons donated by FADH2. –https://www.youtube.com/watch?v=VER6xW_r1vc Per glucose molecule, the net gain is about 36 ATP molecules Fig 5:10- Two step process of oxidative phosphorylation: Formation and harnessing of the proton gradient 91 Oxidative Metabolism in the Mitochondrion Summary of oxidative phosphorylation Glycolysis To produce two Pyruvate molecules: ATP produced= 2x2=4 NADH produced= 2x3=6 ATP used=2 Total= 8 TCA cycle For two pyruvate molecules: CO2 produced=6 NADH produced= 8x3=24 FADH2 produced=2x2=4 Total= 28 ATP ATP is formed by the controlled movement of H+ through the ATP synthesizing enzyme- chemiosmosis 92