Introduction: electron carriers Any of various molecules that are capable ofaccepting one or two electrons from one moleculeand donating them to another in the process ofelectron transport. As the electrons are transferred from oneelectron carrier to another, their energy leveldecreases, and energy is released. Cytochromes andquinones (such as coenzyme Q) are some examplesof electron carriers.
Electron carriers of the transport chain Protein Electron Prosthetic Group CarriersFlavoprotein Flavin Mononucleotide (FMN)Iron-sulfur Protein Iron and SulfurCytochromes Heme GroupMost electron carriers are proteins. Ubiquinone (Q) is theonly electron carrier of the transport chain that is notbound to a protein.
The electron carriers that ferry electrons from NADH andFADH2 to O2 are associated with the inner mitochondrialmembrane. Some of these redox centers are mobile, andothers are components of integral membrane proteincomplexes. Electron Carriers: NADH, Flavoproteins, Cytochromes, Iron–Sulfur Proteins, Quinones1.) NADH.This is water-soluble pyridine derivative used bydehydrogenases, enzymes. Important to note that NADHtransfers 2 electrons at a time in the form of a hydride. NAD+ + 2e- + H+ NADH
2.) Flavoproteins.Some redox enzymes use redoxgroups derived from riboflavin;these are called flavoproteins.The redox groups are the FlavinMono-Nucleotide (FMN) and theFlavin Adenine Dinucleotide(FAD).Flavoproteins can accept or donateelectrons one at time or two at atime because their semiquinoneforms are stable. FAD + 2e- + 2H+ FADH2; FMN + 2e- + 2H+ FMNH2
3.) Coenzyme Q (CoQ) Coenzyme Q (CoQ, Q or ubiquinone) is lipid-soluble. It is a soluble electron carrier in the hydrophobic bilipid layer of the inner mitochondrial membrane. The only electron carrier not bound to a protein. It can accept/donate 1 or 2 e-. O OCH3O CH3 CH3O CH3 CH3 e CH3CH3O (CH2 CH C CH2)nH CH3O (CH2 CH C CH2)nH O O coenzyme Q coenzyme Q • e + 2 H+ OH CH3O CH3 CH3 CH3O (CH2 CH C CH2)nH OH coenzyme QH2
3.) Cytochromes.Cytochromes are proteinsthat contain heme prostheticgroups which function as oneelectron carriers.The heme iron is involved inone electron transfersinvolving the Fe2+ and Fe3+oxidation states.• Some cytochromes (b,c1,a,a3) are part of large integral membrane protein complexes. This is the structure of iron- protoporphyrin IX, one of• Cytochrome c is a small, the most common hemes. water-soluble protein.
4.) Iron-Sulfur Proteins In the ETC we will encountermany iron-sulfer proteins whichparticipate in one electron transfersinvolving the the Fe2+ and Fe3+oxidation states.These are non-heme iron-sulfurproteins.• The simplest iron-sulfer protein is FeS in which iron is tetrahedrally coordinated by four cysteines.• The second form is Fe2S2 which contains two irons complexed to 2 cysteine residues and two inorganic sulfides.
• The third form is Fe3S4 which contains 3 iron atoms coordinated to three cysteine residues and 4 inorganic sulfides.• The last form is the most complicated Fe4S4 which contains 4 iron atoms coordinated to 4 cysteine residues and 4 inorganic sulfides.5.) Copper Proteins Copper bound proteins participate in one electron transfersinvolving the Cu+ and Cu2+ oxidation states.
Overview of the Electron Transport Chain.Electrons move along the electron transport chain goingfrom donor to acceptor until they reach oxygen theultimate electron acceptor.The components of the electron transport chain areorganized into 4 complexes.
Each complex contains several different electron carriers.1. Complex I also known as the NADH-coenzyme Q reductase or NADH dehydrogenase.2. Complex II also known as succinate-coenzyme Q reductase or succinate dehydrogenase.3. Complex III also known as coenzyme Q reductase.4. Complex IV also known as cytochrome c reductase.Complex I accepts electrons from NADH and serves as the linkbetween glycolysis, the citric acid cycle, fatty acid oxidation and theelectron transport chain.Complex II includes succinate dehydrogenase and serves as adirect link between the citric acid cycle and the electron transportchain.
• Complexes I and II both produce reduced coenzyme Q,CoQH2 which is the substrate for Complex III.• Complex III transfers the electrons from CoQH2 to reduce cytochrome c which is the substrate for Complex IV.• Complex IV transfers the electrons from cytochrome c to reduce molecular oxygen into water. Electron carriers operate in sequence.• Each of these complexes are large multisubunit complexes embedded in the inner mitochondrial membrane.
C | Complex I Accepts Electrons from NADH oxidoreductase or NADH dehydrogenase, is a large enzyme composed of 42 different polypeptide chains, including an FMN- containing flavoprotein and at least six iron-sulfur centers. High- resolution electron microscopy shows Complex I to be L-shaped, with one arm of the L in the membrane and the other extending into the matrix. Matrix arm. Hydrophilic domainMembrane arm
Complex I FMNComplex I Contains Multiple Coenzymes. [2Fe–2S] [4Fe–4S]• one molecule of flavin mononucleotide• eight or nine iron–sulfur clusters as the prosthetic groups of iron–sulfur proteinsComplex I catalyzes two simultaneousand obligately coupled processes:• (1) the exergonic transfer to ubiquinone of a hydride ion from NADH and a proton from the matrix and• (2) the endergonic transfer of four protons from the matrix to the intermembrane space. The reaction of NADH dehydrogenase is: NADH + H+ + CoQ + 4H+in → NAD+ + CoQH2 + 4H+out
MechanismAll redox reactions take place Series ofin the extramembranous iron–sulfurportion. NADH initially binds clustersto NADH dehydrogenase, andtransfers two electrons to theflavin mononucleotide (FMN)prosthetic group of complex I,creating FMNH2. The electronacceptor - the isoalloxazine Electronsring - of FMN is identical to transfer paththat of FAD. FMN hydrophilic peripheral domain (Thermus thermophilus)
The electrons are then transferred CoQthrough the second prosthetic groupof NADH dehydrogenase via a seriesof iron-sulfur (Fe-S) clusters, andfinally to coenzyme Q (ubiquinone). Series ofThis electron flow changes the redox state iron–sulfurof the protein, inducing conformational clusterschanges of the protein which alters the pKvalues of ionizable side chain, and causesfour hydrogen ions to be pumped outof the mitochondrial matrix.Ubiquinone (CoQ) accepts two electronsto be reduced to ubiquinol (CoQH2). FMN
Complex II: Succinate to Ubiquinone (membrane-bound enzyme in the citric acid cycle)• Although smaller and simpler than Complex I, it contains five prosthetic groups of two types and four different protein subunits.• Subunits C and D are integral membrane proteins, each with three transmembrane helices. They contain a heme group, heme b, and a binding site for ubiquinone, the final electron acceptor in the reaction catalyzed by Complex II.• Subunits A and B extend into the matrix; they contain three 2Fe-2S centers, bound FAD, and a binding site for the substrate, succinate.
Intermembrane Phosphatidyl ethanolamine space (P side) D C QH2 Ubiquinone Matrix Fe-S centers(N side) Heme b FAD B Substrate binding site
Intermembrane Phosphatidyl ethanolamine space (P side) D C QH2 Ubiquinone Matrix Fe-S centers (N side) Heme b FAD B SubstrateElectron Transfer binding Pathway site
Structure of Complex II (succinate dehydrogenase).• (PDB ID 1ZOY) This complex (shown here is the porcine heart enzyme) has two transmembrane subunits, C and D; the cytoplasmic extensions contain subunits A and B. Just behind the FAD in subunit A is the binding site for succinate. Subunit B has three sets of Fe-S centers; ubiquinone is bound to subunit B; and heme b is sandwiched between subunits C and D. Two phosphatidylethanolamine molecules are so tightly bound to subunit D that they show up in the crystal structure. They serve to occupy the hydrophobic space below the heme b• Electrons move (blue LINE) from succinate to FAD, then through the three Fe-S centers to ubiquinone. The heme b is not on the main path of electron transfer but protects against the formation of reactive oxygen species (ROS) by electrons that go astray.
Complex III: Ubiquinone to Cytochrome c• The next respiratory complex, Complex III, also called cytochrome bc1 complex or ubiquinone:cytochrome c oxidoreductase, couples the transfer of electrons from ubiquinol (QH2) to cytochrome c with the transport of protons from the matrix to the intermembrane space.• The functional unit of Complex III is a dimer, with the two monomeric units of cytochrome b surrounding a ―cavern‖ in the middle of the membrane, in which ubiquinone is free to move from the matrix side of the membrane (site QN on one monomer) to the intermembrane space (site QP of the other monomer) as it shuttles electrons and protons across the inner mitochondrial membrane.
P" side (inter membrane space), RieskeCytochrome bc1 complex Heme c1 iron 2Fe-2S(Complex III). sulfur Cytochrome c1 proteinThe complex is a dimer ofidentical monomers, each with Heme bL Heme bL11 different subunits. Heme bH Heme bH(a) The functional core of eachmonomer is three subunits: Cytochrome bcytochrome b (green) with itstwo hemes (bH and bL); theRieske iron-sulfur protein(purple) with its 2Fe-2Scenters; and cytochrome c1(blue) with its heme (PDB ID1BGY). Cavern "N" side (matrix)
Reaction MechanismThe reaction mechanism for complex III is known as the ubiquinone ("Q")cycle. In this cycle four protons get released into the Positive "P" side (intermembrane space), but only two protons get taken up from the Negative"N" side (matrix). As a result a proton gradient is formed across themembrane. In the overall reaction, two ubiquinols are oxidized toubiquinones and one ubiquinone is reduced to ubiquinol. In the completemechanism, two electrons are transferred from ubiquinol to ubiquinone, viatwo cytochrome c intermediates.Overall:•2 x QH2 oxidised to Q•1 x Q reduced to QH2•2 x Cyt c1 reduced•4 x H+ released into intermembrane space•2 x H+ picked up from matrix
• Round 1:• Cytochrome b binds a ubiquinol and a ubiquinone. The 2Fe/2S center and BL heme each pull an electron off the bound ubiquinol, releasing two hydrogens into the intermembrane space.• One electron is transferred to cytochrome c1 from the 2Fe/2S centre, whilst another is transferred from the BL heme to the BH Heme.• Cytochrome c1 transfers its electron to cytochrome c (not to be confused with cytochrome c1), and the BH Heme transfers its electron to a nearby ubiquinone, resulting in the formation of a ubisemiquinone.• Cytochrome c diffuses. The first ubiquinol (now oxidised to ubiquinone) is released, whilst the semiquinone remains bound.
Round 2:• A second ubiquinol is bound by cytochrome b. The 2Fe/2S center and BL heme each pull an electron off the bound ubiquinol, releasing two hydrogens into the intermembrane space.• One electron is transferred to cytochrome c1 from the 2Fe/2S centre, whilst another is transferred from the BL heme to the BH Heme.• Cytocrome c1 then transfers its electron to cytochrome c, whilst the nearby semiquinone picks up a second electron from the BH Heme, along with two protons from the matrix.• The second ubiquinol (now oxidised to ubiquinone), along with the newly formed ubiquinol are released.
ReactionIt catalyzes the reduction of cytochrome c by oxidation of coenzyme Q(CoQ) and the concomitant pumping of 4 protons from the mitochondrialmatrix to the intermembrane space: QH2 + 2 cytochrome c (FeIII) + 2 H+in → Q + 2 cytochrome c (FeII) + 4 H+outIn the process called Q cycle, two protons are consumed from the matrix(M), four protons are released into the inter membrane space (IM) andtwo electrons are passed to cytochrome c.
Complex IV: Cytochrome cto O2In the final step of the Heme arespiratory chain, Complex IV,also called cytochrome oxidase,carries electrons fromcytochrome c to molecularoxygen, reducing it to H2O.Complex IV is a large enzyme(13 subunits; Mr 204,000) ofthe inner mitochondrialmembrane. Heme a3
Structure of cytochrome oxidase (Complex IV).This complex from bovine mitochondria has 13 subunits, but only fourcore proteins are shown here (PDB ID 1OCC). (a) Complex IV, withfour subunits in each of two identical units of a dimer. Subunit I(yellow) has two heme groups, a and a3, near a single copper ion, CuB(green sphere). Heme a3 and CuB form a binuclear Fe-Cu center.Subunit II (purple) contains two Cu ions complexed with the —SHgroups of two Cys residues in a binuclear center, CuA, that resemblesthe 2Fe-2S centers of iron-sulfur proteins. This binuclear center andthe cytochrome c–binding site are located in a domain of subunit IIthat protrudes from the P side of the inner membrane (into theintermembrane space). Subunit III (light blue) is essential for rapidproton movement through subunit II. The role of subunit IV (green) isnot yet known.
BiochemistrySummary reaction: 4 Fe2+-cytochrome c + 8 H+in + O2 → 4 Fe3+-cytochrome c + 2 H2O + 4 H+outTwo electrons are passed from two cytochrome cs, through the CuA andcytochrome a sites to the cytochrome a3- CuB binuclear center, reducing themetals to the Fe+2 form and Cu+1. The hydroxide ligand is protonated and lost aswater, creating a void between the metals that is filled by O2. The oxygen is rapidlyreduced, with two electrons coming from the Fe+2cytochrome a3, which isconverted to the ferryl oxo form (Fe+4=O). The oxygen atom close to CuB picks upone electron from Cu+1, and a second electron and a proton from the hydroxyl ofTyr(244), which becomes a tyrosyl radical: The second oxygen is converted to ahydroxide ion by picking up two electrons and a proton. A third electron arisingfrom another cytochrome c is passed through the first two electron carriers to thecytochrome a3- CuB binuclear center, and this electron and two protons convertthe tyrosyl radical back to Tyr, and the hydroxide bound to CuB+2 to a watermolecule. The fourth electron from another cytochrome c flows through CuA andcytochrome a to the cytochrome a3- CuB binuclear center, reducing the Fe+4=O toFe+3, with the oxygen atom picking up a proton simultaneously, regenerating thisoxygen as a hydroxide ion coordinated in the middle of the cytochrome a3- CuBcenter as it was at the start of this cycle. The net process is that four reducedcytochrome cs are used, along with 4 protons, to reduce O2 to two watermolecules.
StructureSubunit I and II of Complex IV excluding all other subunits, PDB2EIKThe complex is a large integral membrane protein composed ofseveral metal prosthetic sites and 13 protein subunits inmammals. In mammals, ten subunits are nuclear in origin, andthree are synthesized in the mitochondria. The complex containstwo hemes, a cytochrome a and cytochrome a3, and two coppercenters, the CuA and CuB centers. In fact, the cytochrome a3and CuB form a binuclear center that is the site of oxygenreduction. Cytochrome c reduced by the preceding component ofthe respiratory chain (cytochrome bc1 complex, complex III)docks near the CuA binuclear center, passing an electron to it andbeing oxidized back to cytochrome c containing Fe3+. The reducedCuA binuclear center now passes an electron on to cytochrome a,which in turn passes an electron on to the cytochrome a3- CuBbinuclear center. The two metal ions in this binuclear center are4.5 Å apart and coordinate a hydroxide ion in the fully oxidizedstate.
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