Biochem 2 PDF

Lysosomes -house importantenzymes in...

Lysosomes -house importantenzymes in breaking down molecules `Lecture 2 the pathway when they are not required. Five Principles of Metabolic Pathways 5. Metabolic Pathway in eukaryotic organisms 1. Metabolic Pathways are irreversible occur in specific cellular locations metabolic pathway A highly exergonic reaction (having a -form of compartmentalizing has direction to a ensure that the large negative free energy change) is -ex. Since lysosomes house break down importantenzyme to be its butsome of environment Free energy irreversible; that is it goes to can lysosomal enzyme only functions atacidic condition used properly competition. ___________________________________ It confers directionality on the pathway, thats is, it makes the entire pathway irreversible ‘The endergonic processes that maintain the living state are driven by the exergonic 2. Catabolic and Anabolic Pathways must reactions of nutrient oxidation.’ differ catabolic - breakdown of If two metabolites are metabolically molewles into interconvertible, the pathway from the ___________________________________ simpler molecules first to the second must differ from the AP of cell anabolic pathway from the second back to the energy currency the - synthesis macromolecules of into simpler molecules first adenine It allows independent control of the two processes ↑residue substrate EX. intermediate nitrogeneous base -based from this ex. - ribose the conversion of the two substrate need a different enzyme Phosphate be they tend to be group irreversible that -itis important so pathways won't the clash or resultto futile cycle. 3. Every Metabolic Pathway has a committed step committed steps are investmentprocess Most of the component reactions -consuming energyin the function close to equilibrium form of ATP Irreversible (exergonic) reaction that APP - is an example of nucleotide - committed step rate contains which is ribose sugar residue - “commits” the intermediate it produces a limiting step to continue down the pathway -so basically, AP has 3 phosphate attached to Rate Limiting Step Carbon 5 of Ribose via linkage. phosphresther 4. All Metabolic Pathways are regulated -catalyzed by the the ribose is attached to adenine residue via nitrogenous base enzymes that are In order to exert control on the flux of -each phosphate are connected by a special type of bond called tigthly regulated metabolites through a metabolic phospho anhydride bonds dictates now fastor pathway, it is necessary to regulate its - slow particular Phosphoester rate-limiting step. a metabolic pathway -connects the methyl group, C5 of Ribose to to will proceed, depending The first committed step is often one residue on the first the phosphate called a phosphate physiological biochemical status or of its rate-limiting steps. the cell. of An efficient way to exert control Phosphoanhydride bonds because it prevents the unnecessary -link the phosphate groups the bet 8 prous, the ex. cell is under starvation synthesis of metabolites further along alpha, beta a gamma -these are reservior the free of is utilized by the energy -itneeds to extract endergonic reaction thatthey favorably so proceed energy from available frels, ex, carbs Pyrophosphate residue!! bond thatrelease therefore, regulatory controls combination the of phosphoanhydride for reaction must be in place to activate huge amount endergonic the limiting step rate for metabolic pathways proceed -higher amountreleased.  Phosphoryl-Transfer Reactions Phosphoryl -transfer reactions, are of enormous metabolic significance. Some of the most important reactions of this type DNAto synthesis of involve the synthesis and hydrolysis of ATP: * RNA does not require ATP but Nip still depends to biochemical -provide slightly condition of the higher energy than cell the firstone Remember: Highly exergonic reactions are ATP be in the middle bes its the easiest to make!! OK, so when we coupled to numerous endergonic use As itis importantthat biochemical processes so as to drive outside we a regenerate them through phosphate-containing compound higher that ATP!! them to completion ATP is regenerated by coupling its Free energy: In vitro vs In vivoGinside -most which of present are formation to a more highly exergonic The △G’s of hydrolysis of in a brain tissue muscle metabolic process phosphorylated compounds are therefore highly dependent on pH, Some overall coupled reaction involving divalent metal ion concentration ATP (Mg2+), and ionic strength For example, ATP hydrolysis △G = Endergonic - thermodynamically -50 kJ/mol (in vivo) vs = -30.5 - unfavorable -no action can occur kJ/mol in vitro on its own Exergonic - vice versa!! The concentrations of most substances vary both with location and time. The concentrations of many ions, coenzymes and metabolites commonly vary by several orders of magnitude across membranous organelle boundaries ___________________________________ Rationalizing the Energy in High Energy Compounds ‘The bioenergetic utility of phosphoryl-transfer High energy bonds ,-25 kJ/mol reactions stems from their kinetic stability to hydrolysis combined with their capacity to transmit relatively large amounts of free energy.’ ____________________________________  What is responsible for the high-energy character of ATP? ○ The resonance stabilization of a phosphoanhydride bond is less than that of its hydrolysis Resonance Stabilization -makes the phospho anhydride When this is bond more ammenable to hydrolised hydrolysis which will yield high free energy the productwould - be less resonance stabilized ○ Destabilizing effect of the electrostatic repulsions between the charged groups of a phosphoanhydride in comparison to that if its hydrolysis products II. Enol phosphates -> negative charges repel each other *the greater the repulsion more ammenable hydrolysis to solvation energy ○ Smaller solvation energy of a III. Phosphoguanidines -capacityof water molecules phosphoanhydride in to hydrate charge molecules comparison to that if its - if hydrolysis products I found in brain -has higher density of negative charges therefore, this is solvation ammenable to Other High-Energy Compounds I. Acyl Phosphates 1,3-Biphosphoglycerate ATP occupies the middle rank ○ Most commonly occur in the early stages of carbohydrates metabolism ATP serves a conduit between low energy and high energy Endergonic and exergonic reactions must be coupled Creatine Kinase and Adenylate Kinase (AK) maintain ATP levels in tissues with high, fluctuating energy demands such as brain and muscle ____________________________________ How ATP is formed? ___________________________________ How ATP is consumed? Substrate Level Phosphorylation ○ Direct transfer of a phosphoryl group from a “high-energy” Early stages of nutrient breakdown compound to ADP  Interconversion of nucleoside High levels of products tend to inhibit triphosphates the conversion of substrates to ○ Nucleoside triphosphates products (NTPs) are synthesized from ATP and the corresponding nucleoside diphosphate (NDP) in reactions catalyzed by the nonspecific enzyme nucleoside diphosphate kinase The △G = 0 ATP + NDP ADP + NTP Feedback Control Physiological processes ○ Protein folding (chaperone A ——> B ——> C ——> D ——> E assisted) ○ Muscle contraction ○ Active transport High Endergonic reactions have extra source of energy Allosteric Enzymes Lecture 3 What will happen if enzymes are operating at its maximum rate? Intermediates would pile up Products would be produced in vast excess Depletion of raw materials Other pathways would shut down Two ways by which enzyme can be controlled via allosteric either Substrate-Level Control activation or inhibition The higher the substrate concentration Allosteric means the substrate could is, the more rapidly the reaction bind inside other than active site occurs  causing conformational change This is also demonstrated by same (changes the shapes of the enzyme) protein hemoglobin as influence by other factors not necessarily the 2 Types of Allostery: Homo and Hetero subunits themselves (alpha and beta), it could be the protons, H+, Homoallostery bicarbonate, carbon dioxide (slightly acidic) The enzyme subunit does not directly cause it. For example, hemoglobin can assume two states: Tense (T) which has lower affinity for oxygen and Relax ( R) which has higher affinity for oxygen. The influence of heteroallosteric inhibitors drives the The subunits themselves affect each graph towards the T state. So from other, they enhance cooperative sigmoidal, it drops down a little binding of the substrate. So, alpha 1 because of the inhibitors either influences alpha 2 and beta 1 competitive or uncompetitive. influences beta 2. Activators does the opposite, It shifts This happens to proteins with multi the hemoglobin towards the R state subunits such as hemoglobin with 2 alpha and 2 beta. These subunits are Regulation of Enzyme Activity by Covalent identical, most likely they demonstrate Modification homoallosteric wherein binding of oxygen to 1 subunit would influence Phosphorylation -> a type of covalent the affinity of the other subunits for modification wherein another enzyme oxygen phosphorylates another protein downstream Initially a protein demonstrating leading to cascade of events homoallosteric or cooperative binding, Addition of PO43- to the -OH group of at the first lower concentrations, they Ser, Tyr, Thr (occurs in specific amino seem to be no adequate response in acids) - results in the repulsive terms of maximum velocity, But if you electrostatic interaction that lead to increase the substrate concentration, conformational change it increases dramatically Cooperative: Sigmoidal (curve) Catalyzed by ATP dependent protein Non-cooperative : Hyperbole (curve) kinases Made reversible by phosphatases Heteroallostery (conformational change shape recovered, inactive) Regulates many metabolic pathway Other Covalent/Chemical Modifications: gene that codes for this particular enzymes Ribosylation that affects any of ○ addition of ribose moieties to a these in variant amino protein specifically at the ADP acid, it cannot be residue effectively Acetylation phosphorylated ○ Can remarkably or therefore it cannot be dramatically change the gene activated downstream expression or the switching of Requires a donor of genes in a chromosome phosphate and ATP ATO is also - ○ Can influence the expression a donor of usually served the role of our genes at the level of our phosphate in as the -- donor for the chromosomes phosphorylation phosphorylation of of enzyme enzymes ○ Phosphatases - removal of Phosphorylation the phosphate reversible through the action of phosphatases ○ Kinase & Phosphatase - one of the prominent covalent allows tight regulation of modification that occurs in certain enzyme activity especially in enzymes in metabolic pathways response to a stimulus reversible through the action of phosphatases Addition of PO43- (has high density of Frequently under hormonal control negative charge that can trigger either attractive or repulsive forces on its proximity to a particular amino acid) to the -OH group of Ser, Tyr, Thr phosphorylation ○ results in the repulsive electrostatic interactions that lead to conformational change Ex: Phosphate is placed within the vicinity of the enzyme that contains lysine (+), therefore, it would stimulate attractive forces between There are 2 enzymes that control these residues which can influence the global phosphorylation and these are: conformational changes. Conversely, if the ○ Kinase - facilitates the phosphate is placed in proximity of aspartic phosphorylation of enzymes acid (-), it will cause repulsive forces between and in variant (specific amino these 2 residues. In effect, it will also result in acid residues in enzymes, they the global conformational change in the are not phosphorylated at enzyme shape. This conformational changes random residues, it has to be what triggers the activity of the enzymes. at exact Ser, Tyr, Thr residues) When the phosphate is removed by the such us Serine, Tyrosine, phosphatases, the original conformation of the Threonine enzyme is recovered and it is rendered In the event that there inactive. is a mutation and the  responsible for the GLUT 4 transporters. These are synthesized in the rough endoplasmic reticulum where the protein synthesis machinery is and it is translocated unto the surface of the cell and begins to transport glucose into the cell where its subsequently oxidized to produce ATP as a source of free energy Insulin’s (protein hormone) main Regulation of Enzyme Activity by Protein function is to stimulate the cell’s Backbone Cleavage absorbed glucose when the levels of glucose becomes high in the bloodstream Insulin binds to the insulin receptor on the surface of the cell. Binding of insulin to its receptor triggers conformational change leading to the phosphorylation of a domain of this insulin receptor, specifically tyrosine residues or underneath the cell surface (leading to activation of IRS which is also phosphorylated). In variant tyrosine residues become phosphorylated. So this Pancreas phosphorylation triggers the activities One of the major organs responsible of the receptor which in turn for the synthesis of a multitude of phosphorylates downstream proteins enzymes. Many of these enzymes at specific Serine and threonine perform a role in the digestive process residues in the small intestine You can see that this is a cascade of Proteolytic (backbone) cleavage phosphorylation events affected by Proteolytic: Hydrolyze protein into kinases. That’s why in the insulin specific fragments; Cleavage: cutting signaling, you will encounter enzymes Happen in small intestine and that will perform a role as kinase or pancreas which produces the inactive kinase kinase or even kinase kinase form of these enzymes kinase A number of enzymes undergo Cascade of events culminate a proteolytic cleavage including Trypsin, change in gene expression in chymotrypsin, carboxypeptidase and particular in an increased expression elastase. All are synthesized in the of certain proteins that play a role in pancreas and they are secreted the absorption of glucose from the through the pancreatic duct all the way extracellular environment of cells, to the small intestine in response to a specifically skeletal muscle cells. hormone generated when high protein GLUT 4 gene is expressed or carbohydrate meal is eaten  Zymogens -> inactive form terminus and residue 122 which are (PROelastase, trypsinOGEN) both Cysteine residues ○ Green boxes not active; The product resulting from the image Orange boxes active is π Chymotrypsin. It's the product of irreversible the cleavage of arginine and ○ Because there is a battery of isoleucine at residues 15 and 16 potent proteases in the respectively. pancreas, that would digest The production of alpha-Chymotrypsin itself via auto-catalytic events. (π Nature designed us this way so that Chymotrypsin activates its own to these enzymes will not be destructive produce more to produce more of the organ that produces it, so they alpha-Chymotrypsin) are synthesized by the pancreas in ○ In this way, we can see how their inactive form (zymogens) zymogens undergo Proteolytic of these zymogens to their auto-catalysis to form their cognate active enzymes is irreversible cognate active enzyme X-ray diffraction - a technique used by scientists to resolve the details in proteolytic cleavage resulting from the crystal forms of these proteins which is called X-ray Crystallography First step: Activation of trypsin from the trypsinogen. Without this step the other enzymes will not be activated. Once active trypsin is present it will activate other trypsinogen to make more trypsin. This activation is called auto-catalysis. The activation of chymotrypsin from its These battery of enzymes trypsin, zymogen chymotrypsinogen is one of chymotrypsin, elastase and the most studied proteolytic activation carboxypeptidase and together with Initial step: Trypsin once activated another enzyme that is secreted by cleaves the bond between Arginine our stomach cells the Pepsin, they (15) and Isoleucine (16) residues. This are secreted into the intestinal wall is a highly specific cut, it would NOT cells. They are capable of digesting cut anywhere else other than those most of the proteins in our food into two residues smaller peptides, even free amino N-terminal fragment of acids. These smaller molecules are chymotrypsinogen is not completely absorbed by intestinal epithelium. released, it remains attached to the For example: Pepsin in the stomach is molecule because of the disulfide only active at strongly acidic bond linkage between the amino  conditions which is under pH 1 and 3 inhibitor produced by the pancreas which performs the initial proteolysis itself will not be sufficient to inhibit the of the food that we eat which gives activated enzymes. This happens in a rise to smaller peptides emerging from clinical case called Acute the stomach all the way down to the Pancreatitis (bangungot in PH) small intestine where the zymogens are released through the pancreatic Notes: ducts.These zymogens are acted Trypsinogen when acted upon by upon by the Endopeptidases in the enteropeptidase produces trypsin. small intestine or duodenum which Trypsin itself activates facilitates the auto-catalytic events enteropeptidase to produce more The enzymes themselves are trypsin from trypsinogen. continually subjected to mutual Chymotrypsin digest itself to produce digestion and auto digestion so that alpha chymotrypsin (self high levels of these enzymes never digestion/auto catalytic produces its accumulate in the intestine cause they own) might digest or damage the small Kinase use ATP as a cofactor; as a intestine itself source of phosphate In fact, even the inactive zymogens Phosphorylase can use other source that are synthesized in the pancreas, of phosphate other than ATP such as they are a potential source of danger glycogen phosphorylase (difference to the pancreas itself. It is because from kinase is that it's not meant to trypsin activation can be activate an enzyme it simply auto-catalytic. This could set an phosphorylates to produce glycogen) activation cascade in motion prematurely How do pancreas protect itself from these premature activation of the enzymes? The pancreas synthesizes a protein called Secretory pancreatic trypsin inhibitor (PTI) which protects the pancreas against the prematurely activated zymogen so that it will not digest the pancreas itself. This is a competitive inhibitor of the substrate for specific active enzymes that binds so tightly to the active site trypsin itself that it effectively inactivates it even at a very low concentration. There are some cases that the Secretory PTI is insufficient to mask the activity of trypsin which can be catastrophic for the patient. For example, when the pancreatic duct is blocked, the zymogens are prematurely activated and the trypsin Lecture 4 Oxidative phosphorylation constitutes the third stage of the metabolic Oxidative Phosphorylation: An Overview oxidation of substrates NADH and FADH2 are reoxidized by electron transport proteins bound to the inner mitochondrial membrane ○ You need important substrates for you to be able to synthesize ATP in the mitochondria.These substrates are called Reduced electron carriers including: Nicotinamide Adenine-dinucleotide (NADH) Flavin Adenine-dinucleotide (FADH2) Electron Transport Chain both are reduced form refers to how electrons are shuttled ○ NADH is generated in from one carrier molecule to the next glycolysis while the FADH2 is Oxidative Phosphorylation from the TCA and oxidation of Culmination of this events leading the lipids such as fatty acids. synthesis of ATP in the mitochondria ○ Glycolysis takes places in the -> Kreb's cytosol while TCA takes cycle The average adult human synthesizes place inside the mitochondria ATP at a rate of nearly 1021 molecules specifically in the per second mitochondrial matrix ○ ATP is not directly synthesized ○ NADH and FADH2 carry high from ADP + Pi energy electrons when ○ Synthesis of ATP is highly shuttled to electron transport endergonic reaction, the free chain energy works around positive A series of linked oxidation and 32 kJ/mol. For you to recover reduction occurs, with e-s being ATP you need a few passed along a series of e- carriers mechanisms to do so such as known as the electron transport chain Substrate Level Phosphorylation, Oxidative Mitochondrion Phosphorylation and High Bean-shaped organelle in the cell energy Phosphates specifically in eukaryotic cells. High (phosphocreatine) metabolically active cells have Glycolysis and the citric acid cycle by thousands of mitochondria like cardiac themselves generate relatively little cells, cardiomyocytes and skeletal ATP directly muscle cells. So density of mitochondria in a given cell would depend on their metabolic activity.  Mitochondria is made up of 2 Protein Complexes membranes: (double membranous Complex I-IV are involved in the organelle) shuttling or mobilization of electrons ○ (1) Outer membrane from one complex to another -- ○ Intermembrane space Complex V is responsible for the space between the ○ (2)Inner membrane synthesis of ATP - two membrane Made up of interfolded Complex I functions as a membranous proton pump, it where the complexes I -> III via embedded. invaginations called pumps electron from CoQ cristae the mitochondrial hydrophobic matrix to the Mitochondrial matrix protein carrier The culmination of cellular aerobic intermembrane shuttling of electron generating + respiration phosphorylation is oxidative space Pumps 4 protons ATP receives electron The NADH formed by glycolysis and from NADH the NADH and FADH2 formed by TCA (cofactor) and are utilized to reduce O2 and generate transfer it to another ATP carrier Coenzyme Q The electron transport chain consists (hydrophobic lipid carrier) of series of proteins that receive high NADH is oxidized to energy e-s from NADH and FADH2 NAD+. It releases its The e-s are more along the protein electrons to complex complexes and on the the final e- I. These electrons acceptor are carried by CoQ Proton gradient is established to to Complex III generate ATP Complex II does NOT function II -> III via as a proton pump Coq 0 protons are pumped since it is not a proton pump receives electron from FADH2 (requires fat as a cofactor) which is generated from the 7th step of the citric acid cycle the oxidizes succinate to fumarate Serves as the enzyme for the oxidation of succinate to fumarate Complex III functions as a III -> IV via proton pump Cyt C Pumps 4 protons  receives electron is not good for the from CoQ, it does cell not require a Thus, the Complex cofactor V relieves the It receives electron electron gradient by form CoQ, and in allowing protons to the process it be shuttled back to stimulates the the inner matrix extrusion of protons Once protons go from the matrix to to back to complex V, the intermembrane this where the free space energy is generated This creates an The free energy that even greater was generated from electrical potential the influx of protons difference between back to the inner the matrix and matrix is harnessed intermembrane to produce ATP from space ADP and inorganic Complex III requires Phosphate Cytochrome C which is a mobile electron carrier that What happens when most of the protons enables the (pumped out) move to the inter membrane transport of space? electrons to There will be an electron chemical Complex IV gradient or electrical potential difference between the matrix and the Complex IV functions as a proton pump intermembrane space. O2 -> H20 Pumps 2 protons What happens when we have an electron From Cytochrome C chemical gradient, would it produce energy its transferred to the right away? ultimate electron No. But this is important to build free acceptor which is - energy later on to enable Complex V the oxygen then to synthesize ATP reduced to water completing the When is the free energy released? oxidation of It will be released once the protons precursor such as shuttle back to the inner matrix which glucose happens in Complex V (not in the build up of the electron gradient) Complex V Since the matrix and intermembrane space have a Notes: greater build up of Electron do not just move around the electrochemical protein complexes, there must be a gradient or potential series of electron carrying events difference, there within these protein complexes must be a way for ATP is not directly synthesized from this gradient to ADP + Pi normalize because it ↓ for equilibrium  Oxidative phosphorylation links the amenable to movement across the free energy generated from the lipid bilayer because of its lipidic electrical potential difference in the chemical nature mitochondria and the synthesis of ATP Complex I also functions as a proton by coupling ADP and inorganic pump. As it receives electron, it is also phosphate stimulated to pump electrons from the matrix into the intermembrane space. Recap This is the preliminary stage of the These 4 complexes are crucial in the production of the electrochemical shuttling of the electrons which gradient generate the electrochemical gradient Complex II essential for the synthesis of ATP Complex II receives electron from which is facilitated by the 5th complex. FADH2 which is a high energy These complexes are found in the electron carrier from the citric acid inner membrane of mitochondria cycle/TCA. which separate the mitochondrial Citric acid cycle is a highly conserved matrix and intermembrane space wher metabolic pathway for carbohydrate Complex I and II receives electrons metabolism from reducing equivalents or high The oxidation of succinate and energy electron carriers that are fumarate is catalyzed by an enzyme generated from glycolysis (NADH) and called succinate dehydrogenase. A citric acid cycle/TCA (NADH and class of enzymes that facilitate FADH2). These electrons are crucial oxidation reactions. in the generation for the electron In the process, FADH2 which is also a chemical gradient. cofactor of the same enzyme receives These complexes are found in the electrons from this oxidation process. inner membrane of mitochondria Why is the citric acid cycle directly which separate the mitochondrial linked to the electron transport chain? matrix and intermembrane space Because the enzyme succinate where the protons are extruded when dehydrogenase catalyzes the electrons pass along this complexes oxidation of succinate to fumarate is Intermembrane space is the matrix the very enzyme that makes up the between the outer and inner complex II. Complex II and succinate membrane. dehydrogenase are the enzyme Complex I enzymes. It is a membrane-bound Complex I receives electrons from enzyme embedded in the inner NADH and these electrons are passed membrane of mitochondria. along the series of electrons carriers Electrons are received from the right along complex I. These electrons FADH2 and passed along to a series do not just wander around, these of electron built-in carrier right within protein complexes have built-in complex II electron carriers within them. Once complex II receives its electrons The electrons received by complex I is from its own cofactor, it shuttles the then passed along to a mobile carrier electrons to coenzyme Q which is the which coenzyme Q. It is a highly same mobile carrier that receives hydrophobic molecule which makes it electrons from complex I.  The difference between complex I and This very action dissipates the complex II is that complex II is not a electrochemical gradient and it is proton pump. It does not pump converted to a massive amount of free electrons into the intermembrane energy which is used in the direct space. It is not generally involved in coupling of ADP and inorganic the generation of the electrochemical phosphate to produce ATP gradient Complex III Respiratory Complexes: I, II, III, IV The electrons in the CoQ are now carried over to Complex III which is a Some inhibitors would inhibit Complex large protein complex and has built-in I, so basically it blocks the movement electron carriers which receive 1 of electrons from Complex I to CoQ. electron at a time from CoQ. Once this happens, there will be a In the process, it also serves as proton dissipation of electrochemical pump that extrudes proton from the gradient. Therefore, it also results in matrix to the intermembrane space the reduction of ATP. Complex III passes the electron to There are also inhibitors that target another mobile carrier called specific complexes like complex III, Cytochrome C which is a mobile this blocks the movement of electrons protein carrier. from complex I and II which effectively Complex IV dissipates the electrochemical The electrons from the Cytochrome C gradient and ATP production shuts in Complex III is passed to the last down. complex which is Complex IV. Using this inhibitors, scientists are From complex IV, it passed the able to determine the specific electrons to the ultimate electron functions of each of protein complexes acceptor which is the oxygen which is and their role in the generation of eventually reduced to water electrochemical gradient Complex IV is a proton pump. Respiratory Complex I The series of electron shuttling yields a massive electrochemical gradient which conserves an osmotic energy. This osmotic energy generated by the unequal distribution of protons between the matrix and the intermembrane space build up NADH-Coenzyme Q reductase however it remains unutilizable until (Complex I) such time the Complex V allows the The main donor of electrons into the exodus of these protons back into the respiratory chain because it received matrix high energy electrons from the Complex V reducing equivalent called reduced It is also a proton pump but does the nicotinamide nucleotide (NADH) opposite. It pumps electrons back to Large, membrane embedded the matrix from the intermembrane multisubunit complex ( 1000 kDA) with space about 45 separate polypeptide chains  Complex I is L-shaped, a domain oxidizes succinate to fumarate using a sticks out (receives electrons) in the coenzyme called FAD+. mitochondrial matrix which must be polar/hydrophilic. It is directly exposed to the NADH. While the other domain associated by the lipid bilayer of the inner membrane (hydrophobic) consists of built-in electron carriers It has also a domain sticking out the right within the complex I itself: matrix and the other domain ○ Flavin Mononucleotide embedded in the inner membrane (FMN) It has built-in electron carriers that ○ Iron Sulfur clusters (Fe-S) receives electrons from FADH2: *There are 8 of theme inside ○ Iron-Sulfur Clusters (Fe-S) complex I ○ Cytochrome B (Heme) NADH is oxidized to NAD+, electrons will not just float around, it will be Respiratory Complex III received by a series of cofactors. (1) FMN receives electrons directly from NADH and transfers them into Fe-S clusters. *1 electron at a time. When it does so, the electrons are passed along CoQ. Respiratory Complex II Coenzyme Q: Cytochrome c Oxidoreductase Catalyzes the transfer of electrons from CoQH2 to cytochrome c Two-electron donor, CoQH2, is transferring electrons to one-electron acceptors the cytochromes (can only Succinate-Coenzyme Q reductase process 1 electron at a time) Uses and FAD coenzyme to extract Like Complex I and II, it has also a electrons from succinate domain sticking out the matrix and the FADH2 is the same coenzyme or other domain embedded in the inner cofactor that receives electrons from membrane which basically anchors the oxidation of succinate to fumarate. this complexes into the mitochondrial This particular reaction occurs in the inner membrane citric acid cycle. So there is direct It has built-in electron carriers: connection between TCA and electron ○ Cytochrome B transport chain through complex II ○ Iron-Sulfur Cluster because it is also the enzyme that  ○ Cytochrome c1 Chemiosmotic Coupling It has a domain that is exposed to the Enzymes catalyzing theses intermembrane space (the space dehydrogenations are asymmetrically between the inner and outer oriented in the inner membrane membrane) Protons are always taken up from ○ Rieske Iron-Sulfur Protein inside the matrix and release in the It receives electrons intermembrane space from electron carriers This proton pumping by respiratory built-in Complex III proteins results in conversion of the The electron transport through energy of electron transport to osmotic Complex III is coupled to the extrusion energy, in the form of an on protons from matrix to the electrochemical gradient intermembrane space Respiratory Complex IV Cytochrome c oxidase The final stage of electron transport is carried out by cytochrome c oxidase This electrochemical gradient is the It does not serve as bridge between 2 unequal distribution of the protons electron carriers since they are continuously being What happens to the electrons from pumped out by complexes. This Cytochrome C transferred to Complex massive electrochemical gradient IV, would there be a recipient? It would generates osmotic energy. be the Oxygen. Osmotic energy remains latent until Catalyzes the transfer of electrons another complex come to play by from reduced cytochrome c to oxygen facilitating the re-entry of the protons ○ Initial oxidation of Cytochrome to the matrix through a channel called C is facilitated by Iron-Copper F0/F1 or Complex V (ATP Synthase) Centers. The two Fe-Cu This complex serves as an enzyme centers are the path of that directly couples the free energy electrons from the Cyt C to the that results from the dissipation of this molecular oxygen electrochemical gradient. How much ATP is formed? That will depend on the reducing equivalent. ○ For every mole of NADH, it yields 2.5 moles of ATP  are embedded these complexes in inner mitochondrial matrix electron NADH gives up - NADHonly I b ecame so it NADH is the inter membrace works in complex s O mitochondrial matrix in the proton gradient. This NADHdonates e here where C1 and space to there were It happens from complexes, makes itsuper marged! 113,94 pumps proton from from intermembrane space -complex I then a Now the protons mito mat to interspace 5or ATP synthase through complex -dat Complex Itransfer make proton will flow to make it etO - COG mitochondrial matrix to equilibrium. Is the 200 will then only works in convertADP-> ATP approach we complex 2 - the goal here is to itwill but be impossible with of energy input pass electrons - itcannot be than AP be ADP has lesser energy super charged so electrons will itcannotpump protone be back to WOQ C 7 the final electron receptor electrone here will go to - -now world be oxygen. Oxygen splits to 2 makes it ⑲not1 Complex 3 a when the oxygen proton spile,protons inthe super charged, makes gradient O L electrons a water mouse from complex's the 7 here will be passed to cytic - will HeU+H20 electrons where 4 be pass to complex super charged as making it well, make proston gradient