ELECTRON TRANSPORT CHAIN

1. M.Prasad Naidu MSc Medical Biochemistry, Ph.D.Research Scholar Electron transport chain

2. INTRODUCTION • ETC is the 4th and final stage of aerobic respiration. • Through ETC, the E needed for the cellular activities is released in the form of ATP. • ETC is an O2 dependent process which occurs in the inner mitochondrial membrane.

3. What is ETC ? • The energy rich carbohydrates (Glu), FA and AAs undergo a series of metabolic reactions and finally get oxidized to CO2 and H20. • The reducing equivalents from various metabolic intermediates are transferred to NAD+ and FAD to produce NADH and FADH2. • The latter two reduced coenzymes pass through the ETC or respiratory chain and finally reduce O2 to H20. • The passage of electrons through the ETC is associated with the loss of free energy. • A part of this free E is utilized to generate ATP from ADP and Pi.…

4. DEFINITION • This is the final common pathway in aerobic cells by which electrons derived from various substrates are transferred to oxygen. • ETC is series of highly organized oxidation-reduction enzymes.

5. Location • ETC is localized in Mitochondria. • MC are the centres for metabolic oxidative reactions to generate reduced coenzymes (NADH and FADH2) which in turn, are utilized in ETC to liberate E in the form of ATP. • Hence, MC is regarded as Power House of the Cell.

6. Mitochondrial Organization • 5 distinct parts. • 1.the outer membrane • 2.the inner membrane • 3.the inter membrane space • 4.the cristae • 5.the matrix.

7. Inner mitochondrial membrane • ETC and ATP synthesizing system are located on IMM. • IMM is rich in proteins. • It is impermeable to ions(H+,K+,Na+) and small molecules (ADP, ATP). • IMM is highly folded to form Cristae. • The surface area of the IMM is greatly increased due to Cristae. • The IMM Possesses specialized particles ( that look like lollipops ), the phosphorylating subunits which are the centres for ATP production.

8. Mitochondrial matrix • The interior ground substance. • Rich in enzymes responsible for TCA Cycle, oxidation of FA and the oxidation of amino acids.

9. Structural Organization of ETC • The IMM can be disrupted into 5 distinct enzyme complexes, denoted as Complex I, II, III, IV and V • The complex I-IV are carriers of electrons while V is responsible for ATP synthesis. • Besides these enzyme complexes, there are certain mobile e- carriers in ETC. • These include NADH, Coenzyme Q, Cytochrome C and Oxygen. • The complexes (I-IV) and the mobile carriers are collectively involved in the transport of e- which ultimately combine with O2 to produce H2O. • Most of the O2 supplied to the Body is utilized by MC for ETC.

10. Components of the ETC • Complex I(NADH-CoQ reductase), Complex II(Succinate Co.Q reductase), Complex III(CoQ-Cyt C reductase) Complex IV(Cyt.oxidase) & Complex V(ATP synthetase) There are 5 distinct carriers that participate in the ETC.Viz • 1.Nicotinamide nucleotides • 2.Flavo proteins • 3.Iron-Sulfur proteins • 4.Coenzyme Q • 5.Cytochromes. • These carriers are sequentially arranged and are responsible for the transfer of e- from a given substrate to ultimately combine with proton and O2 to form H2O.

11. Nicotinamide Nucleotides • Of the 2 coenzymes NAD+ and NADP+ derived from the vit. Niacin, NAD+ is more actively involved in the ETC. • NAD+ is reduced to NADH + H+ by dehydrogenases with the removal of 2H atoms from the substrate. • The substrates include Gly-3-P, Pyruvate, isocitrate, α-KG, and malate. • NADPH + H+ produced by NADP+ -dependent dehydrogenase is not usually a substrate for ETC. • NADPH is more effectively utilized for anabolic reactions • Eg: FA synthesis, Cholesterol synthesis.

12. Flavoproteins • The enzyme NADH dehydrogenase (NADH-CoQ reductase) is a flavo protein. • FMN is the prosthetic group. • FMN accepts 2e- and a proton to form FMNH2. • NADH dehydrogenase is a complex enzyme closely associated with non-heme iron proteins (NHI) or iron-sulfur proteins. • NADH+ H++ FMN--NAD+ + FMNH2 • SDH(Succinate-Co.Q reductase) is an enzyme found in the IMM. • It is also a flavoprotein with FAD as the coenzyme. • SDH can accept 2 H atoms(2H+ + 2e- ) from succinate. • Succinate+FAD- Fumarate + FADH2

13. Iron Sulfur proteins • A group of quinones has been found to be present in MC namely FeS, Fe2S2, Fe4S4 and Fe3S4 etc., • FeS proteins exist in the oxidized(Fe3+) or reduced (Fe2+). • About 6 FeS proteins connected with ETC have been identified. • The machanism of FeS proteins in ETC is not clearly understood. • One FeS participates in the transfer of electrons from FMN to Co.Q • Other FeS proteins associated with cyt.b and cyt.c1 participate in the transport of electrons. • Vit E, D and plastoquinones also involved in ETC.

14. FeS Proteins • FeS: It has a single Fe coordinated to the side chain-SH groups of 4 Cys.residues. • Fe2S2: It contains 2 iron atoms, 2 inorganic sulfides and 4 –SH groups. Each iron is linked to 2-SH and 2-sulfur groups. • Fe4S4: It consists of 4 iron atoms and 4 cys-SH groups and 4 inorganic sulfides. Each iron remains linked to 1-SH, 3-inorganic sulfides while each sulfide is coordinated to 3 iron atoms. • Fe3S4: It consists of 3 Fe, 4- SH and 4inorganic sulfides. • Each FeS protein transfers only one e- at a time. • The enzymes may have one or more of the combinations

15. Coenzyme Q • Also called Ubiquinone since it is ubiquitous in living system. • It is a quinone derivative with a variable isoprenoid side chain. • The mammalian tissues possess a quinone with 10 isoprenoid units which is known as coenzyme Q10.(CoQ10) • CoQ is a lipophilic e- carrier. • CoQ is not found in Mitochondria • Vit k performs similar function as CoQ in these organisms.

16. Cytochromes • Cytochromes are conjugated proteins. • Contains Heme group. • The heme group of cyt differ from that Hb and Mb. • The Iron of Heme in cyt is alternately oxidized(Fe3+) and reduced(Fe2+), which is essential for the transport of e- in ETC. • This is in contrast to the Hb and Mb iron which remains in the Fe2+ state.

17. Cytochromes • Cyt are identified by their characteristic absorption spectra. • Ferricytochromes show diffuse and non-characteristic absorption spectra. • Ferrocytochromes exhibit characteristic absorption bands called α, β and γ–soret bands. • Cytochromes are characterized into different groups according to the light wavelength at which the alpha band shows its peak(α-abs.max.)

18. Types of cytochromes • cyt.c:- Since it is largely available , it is the best studied of the cyts. • It is a central member of ETC with an intermediate redox potential) • Water soluble-loosely bound to IMM-easy to extract. • Shows characteristic absorption spectra in the reduced form at 550,521 and 416mµ • Oxidized form @530mµ and 400mµ • The iron content of cyt.c. is 0.38% • Heme is attached with protein by means of 2 thioester linkages involving sulfur of 2 cys and apoprotein. • Cyt.c is incapable of combining with O2/CO. • a protein with 1-PPchain 104aa (mw12400-13000) • NADPH-Cyt.c.reductase can readily reduce Cyt.c

19. Types of cytochromes • Cyt.c1:like cyt.c – contains an ironprotoporphyrinIX complex-heme-c. • It has abs.maxima @554,524&418mµ • Incapable of combining w O2,CO,CN- • Cyt.b:also- protoporphyrinIXcomplex-(heme-b). But the apoprotein is diff. • Tightly bound to Flavo proteins and ubiquinones in the MC. • The Ferrocyt.b has an abs.max.@563mµ, 530mµ & 430mµ. • It is thermostable & not easily extractable. • It also does not react with O2,CO or CN- • Normally its oxidation requires the presence of Cyt.c,a,& a3.

20. Cytochrome a & a3 (cyt.oxidase) • Complex IV of the ETC. • Both contain an identical type of iron porphyrin complex • Inspite of identical hemes, cyt.a & a3 differ in e-affinity & bio.activity.This is bcos of their location of hemes • One heme is located along with one Cu ion. This heme is called heme-a • This Cyt.a functions as anaerobic oxidizing unit.

21. Cytochrome a & a3 (cyt.oxidase) • The other heme is located along with the 2nd Cu ion and is called heme-a3 (functions as aerobic reducing unit). • Cyt.a-abs.max-605,517&414;Cyt.a3600&445mµ. • Cyt.a does not react with O2,CO/CN- where as Cyt.a3 is autooxidizable and forms compounds with CO & CN-.

22. PEROXIDASES • Heme containing enzymes. • Found in bacteria, fungi and animals. • On the basis of sequence similarity peroxidases are grouped into two super families. • 1.fungal, plant & bac.peroxidases • 2.animals form the 2nd super family of peroxidases. • P.ases use H2O2 as the e- acceptor to catalyze a no. of oxidative reactions. • P.ases contain heme group and this heme group is responsible for carrying out the activity of peroxidases • Heme consists of a protoporphyrin ring and a central iron atom in +3 oxidation state. • A protoporphyrin is made up of 4 pyrrole rings linked by methine bridges.--- with diff. side chains.

23. CATALASES • Heme containing redox enzymes. • Produced by all aerobic organisms ranging from bacteria to man. • Converts H2O2 to H2O and mole.O. • Utilizes H2O2 both as an e-acceptor and an e- donor. • Catalase also catalyzes • RCOOH + HQOH  ROH + QO + H2O where R is an alkyl or acyl group and HQOH is a 2e- donor. • Most catalases exist as tetramers of 60-75KD. • Each subunit contains an active heme group buried deep within the structure. • The stable structure of catalases is resistant to PH, thermal denaturation and proteolysis. • About heme

24. Classes of catalases • 1. Monofunctional heme containing catalases. • 2. Bifunctional heme containing catalase-peroxidases that are closely related to plant peroxidases. • 3. Non-heme manganese containing catalases.

25. Haemocyanin (Blue blood) • Are copper containing dioxygen carriers. • Responsible for Di O2 transport in molluscs and Arthopods • High mol.wts and multiple subunits. • Each subunit has a mol.wt of 76000D • Each subunit is made up of 3 domains. • Di O2 is bound to the active site in 2nd domain. • There are 2 Cu atoms with an oxidation state of +1 • So a PP chain of Arthopod HeCN binds 1 O2 molecule. • The structure of Molluscan HeCN is diff. from that of Artho. in Wt, subunits structure and O2 binding capacity. • Mol.wt of HeCN is 290000D with 2 Cu atoms for every 50KD • So 1 PP chain can bind 6-8 O2 molecules.

26. Haemerythrin • Is a biological Di O2 carrier. • Responsible for Di O2 transport in marine invertebrates. • The 4 diff.phyla of invertebrates are Sipuniculids, priapulids, Brachiopods & annelid worm magelona • HeEy is found as an oligomer. • Blood contains an octameric form & tissues contain trimeric or tetrameric • Octameric HeEy consists of 8 subunits which are very similar to 40 structure to MyoHeEy. • Although diff oligomers are known, all of them share a DiIron active site. • The 2 iron atoms are 3.25-3.30Ao apart. • The 2 Fe atoms are bound to 5 His.residues.

27. Model synthetic complexes • Study of model compound involves the structure determination, physical measurements and reactions of simple co-ordination compounds. • A model can give only a partial view of real system and provide valuable evidence for the study of the real systems.

28. BIO INORGANIC CHEMISTRY-I • ESSENTIAL & TRACE ELEMENTS • Essential for life of animals, plants & microbes. • They include Na, K(alkali metals), Ca, Mg (alkali earth metals) & transition metals (V,Cr,Mn,Fe,Co,Zn, Mo and Cd.) • These elements are required for biological processes and are called essential elements. • Trace metals----occurs low conc in animal and plant cells. They are a part of good nutrition. • In high doses they may be toxic to the body or produce deficiencies in other trace metals. • For Eg. High levels of Zn can result in the def. of Cu.

29. Role of metal ions in bio.processes • Regulatory axn is exercised by Na+, K+,Mg2+ & Ca2+ ions. • As cellular regulators they are involved in nerve transmission, • Maintanence of cell membrane permeability and • Regulation of osmotic pressure • Ca regulates muscle contraction, cell division and growth, & enzyme activities. • Also – blood coagulation system • Mg,Ca, and Zn ions have structural role. • Ca is a component of bones, teeth and animal shells. • Zn – structural role in fingures • Mg helps to stabilize 3D-structure of RNA& DNA.

30. Role of metal ions in bio.processes • Metallo enzymes catalyze several biological reactions. • Metal ions are at the active site of these enzymes. • Imp.metal enzymes – CP(Zn), Urease (Ni) & vit.B12(Co) • Metal ions play imp.role in diO2 tpt and storage. • A diO2 carrier protein contains diO2 binding site. This active site is a complex of Fe/Cu. • The 3 imp.DiO2 carriers are Hb, HeEy & HeCN • Hb:- found in RBC----respiration---- the active site of Hb consists of iron- porphyrin( heme) group. • HeEy:- found in marine invertebrates. The O2 binding site contains a pair of Fe atoms. • HeCN:- Cu containing diO2 carriers found in molluscs and arthopods.

31. Role of metal ions in bio.processes • Mb stores O2 in muscles. It contains a heme group • Metal ions play an imp. Role in e- transfer agents include ferredoxin, rubredoxin and cytochromes. • Ferredoxin and rebredoxin contain Fe-S sites. • These sites are involved in e- transfer • Cytochromes serve as e- carriers in both plants and animals.

32. Role of zinc • 2nd most abundant transition element in the hu. body. • About 2 gms of Zn and requires a daily intake of (RDA) 8-13mg. • Stimulates the activity of 100 enzymes.Eg: CA, CP • Plays structural role in proteins called zinc fingures. • Also required in plants for leaf formation and synthesis of auxin. • Zn ion is good lewis acid in biochemical systems. • Zn2+ can be 4,5,6- coordinate. • Zn2+ complexes show easily 4 to 5 coordinate interconversion. If the interconversion is fast, catalysis is also fast. • Zn complexes are labile than Ni2+/ Mg2+ complexes

33. Role of Calcium • Ca has a structural role. • Chief component of bones, teeth and animal shells. • Imp. In cellular messenger system. • Muscle contraction, secretion, ion transport, cell division and growth and blood clotting. • Ca and P are imp for bone formation. • 99% of Ca is stored in bones • Ca is necessary for the growth of children.

34. Role of Iron • Humans contain about 4 gms of iron. • Functions as the principal e- carrier in biological oxidation & reduction reactions. • Fe-S proteins are present in all forms of life. • Fe-S sites occur in ferredoxins and rubredoxins. • They are involved in intra protein and inter protein e- transfer.