Biochemistry and evolution of anaerobic energy metabolism in eukaryotes

Abstract

Major insights into the phylogenetic distribution, biochemistry, and evolutionary significance of organelles involved in ATP synthesis (energy metabolism) in eukaryotes that thrive in anaerobic environments for all or part of their life cycles have accrued in recent years. All known eukaryotic groups possess an organelle of mitochondrial origin, mapping the origin of mitochondria to the eukaryotic common ancestor, and genome sequence data are rapidly accumulating for eukaryotes that possess anaerobic mitochondria, hydrogenosomes, or mitosomes. Here we review the available biochemical data on the enzymes and pathways that eukaryotes use in anaerobic energy metabolism and summarize the metabolic end products that they generate in their anaerobic habitats, focusing on the biochemical roles that their mitochondria play in anaerobic ATP synthesis. We present metabolic maps of compartmentalized energy metabolism for 16 well-studied species. There are currently no enzymes of core anaerobic energy metabolism that are specific to any of the six eukaryotic supergroup lineages; genes present in one supergroup are also found in at least one other supergroup. The gene distribution across lineages thus reflects the presence of anaerobic energy metabolism in the eukaryote common ancestor and differential loss during the specialization of some lineages to oxic niches, just as oxphos capabilities have been differentially lost in specialization to anoxic niches and the parasitic life-style. Some facultative anaerobes have retained both aerobic and anaerobic pathways. Diversified eukaryotic lineages have retained the same enzymes of anaerobic ATP synthesis, in line with geochemical data indicating low environmental oxygen levels while eukaryotes arose and diversified.

Fig 1

Two organelles in comparison. (A) Generalized metabolic scheme of pyruvate oxidation and oxidative phosphorylation in a typical oxygen-respiring mitochondrion, for example, from rat liver. (B) Generalized metabolic scheme of fermentative pyruvate oxidation in trichomonad hydrogenosomes, as proposed in the early 1970s. The presence and absence of organellar genomes are indicated. End products are boxed. Abbreviations: CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; C, cytochrome c; A, ATPase; Fd, ferredoxin; [1], pyruvate:ferredoxin oxidoreductase; [2], acetate:succinate CoA-transferase; [3], succinyl-CoA synthetase; [4], hydrogenase; [5], malic enzyme; [6], pyruvate dehydrogenase complex.

Fig 2

Organelles of mitochondrial origin. (A) The mitochondrial family of organelles, divided along functional lines (classes 1 to 5). Class 1, the canonical, rat liver-type mitochondrion (as described in most textbooks), which uses oxygen as the terminal electron acceptor; class 2, an anaerobically functioning mitochondrion, which uses an endogenously produced electron acceptor, such as fumarate, instead of oxygen; class 3, a hydrogen-producing mitochondrion, which possesses (besides a proton-pumping electron transport chain) a hydrogenase and hence can use protons as a terminal electron acceptor and is therefore qualitate qua also a hydrogenosome; class 4, hydrogenosomes, anaerobically functioning ATP-producing organelles of mitochondrial origin that can use protons as an electron acceptor, which results in the formation of hydrogen; class 5, mitosomes, organelles of mitochondrial origin that are not involved in ATP production. Red indicates that oxygen is consumed in the production of ATP, blue indicates the production of ATP without the use of oxygen, and yellow indicates that the organelle is not involved in the production of ATP. (B) Criteria for the functional classification of organelles of mitochondrial origin.

Fig 3

Phylogenetic distribution of organelles of mitochondrial origin. The distribution is plotted across a schematic phylogeny for the six currently recognized major clades or supergroups of eukaryotes (4, 180, 237). Species for which metabolic maps are presented in this review are shown in boldface type. The mitochondrion class for each example species is indicated, and numbers in the column “organelle classes” correspond to the scheme described in the legend of Fig. 2. The presence of a genome is indicated by encircled DNA. Sizes of individual mitochondrial types are not drawn to scale. LECA, last eukaryotic common ancestor. Note that functional information is absent for the loriciferan organelle. Note the absence of lineages lacking organelles of mitochondrial origin among eukaryotes.

Fig 4

Major pathways of energy metabolism in anaerobic mitochondria of the adult parasitic platyhelminth Fasciola hepatica (common liver fluke). The map is redrawn based on data described previously (500). The main end products of the adult parasites are acetate and propionate, with minor amounts of lactate and succinate. In the presence of oxygen, this metabolism of the adult helminth remains unchanged. Aerobic mitochondrial metabolism, occurring in free-living and juvenile parasitic stages of F. hepatica in the mammalian host and leading to the reduction of oxygen and water production, is shaded in gray (and in all following metabolic figures as well) to distinguish it from anaerobic respiration and fermentation pathways, drawn in black. Abbreviations: CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; RQ, rhodoquinone; C, cytochrome c; A, ATPase; FRD, fumarate reductase; [2B], acetate:succinate CoA-transferase (subfamily 1B); [3], succinyl-CoA synthetase; [5], malic enzyme; [6], pyruvate dehydrogenase complex; [8], fumarase; [10], phosphoenolpyruvate carboxykinase (ATP dependent); [11], malate dehydrogenase; [14], methylmalonyl-CoA mutase; [15], methylmalonyl-CoA epimerase; [16], propionyl-CoA carboxylase. The photograph shows Fasciola hepatica at the adult stage, with a length of ca. 2.5 cm. (Photograph by Louk Herber, Utrecht University, Netherlands.)

Fig 5

Major metabolic pathways and electron transport chain in mitochondria of the adult nematode Ascaris lumbricoides (giant roundworm). The map is redrawn based on data reported previously (249). Mature forms of this animal, a large and very common parasitic worm, inhabit the small intestine of their hosts. Like most parasitic worms, Ascaris performs malate dismutation to obtain redox balance and uses rhodoquinone to donate electrons for fumarate reduction (434). However, Ascaris is unusual, as acetyl-CoA and propionyl-CoA are used for the production of two branched-chain fatty acids, 2-methylbutanoate and 2-methylpentanoate, which are formed by the condensation of an acetyl-CoA and a propionyl-CoA or two propionyl-CoAs, respectively, with the subsequent reduction of the condensation products (436, 437). These short branched-chain fatty acids are typical end products for Ascaris (249, 251, 298). Abbreviations: MOP-CoA, 2-methyl-3-oxo-pentanoyl-CoA; MOB-CoA, 2-methyl-3-oxo-butanoyl-CoA; CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; RQ, rhodoquinone; C, cytochrome c; A, ATPase; FRD, fumarate reductase; [2A, B], acetate:succinate CoA-transferase (subfamilies A and B); [3], succinyl-CoA synthetase; [5], malic enzyme; [6], pyruvate dehydrogenase complex; [8], fumarase; [11], malate dehydrogenase; [14], methylmalonyl-CoA mutase; [15], methylmalonyl-CoA epimerase; [16], propionyl-CoA carboxylase; [17], phosphoenolpyruvate carboxykinase (ITP/GTP dependent); [44], condensing enzyme; [45], 2-methyl acetoacyl-CoA reductase; [46], hydratase; [47], 2-methyl branched-chain enoyl-CoA reductase; [48], acyl-CoA transferase. The photograph shows Ascaris lumbricoides worms at the adult stage, with lengths of ca. 15 cm (top, male) and 30 cm (bottom, female). (Photograph by Rob Koelewijn, Harbor Hospital, Rotterdam, Netherlands.)

Fig 6

Major pathways of facultative anaerobic energy metabolism in mitochondria of the free-living mollusk Mytilus edulis (blue mussel). The map is redrawn based on data reported previously (107). Living attached to hard substrates, for example, rocks in intertidal habitats, the bivalve has to face anaerobiosis periodically. Oxygen-independent cytosolic energy metabolism produces ATP via substrate-level phosphorylation accompanied by the formation of various end products, including alanopine, strombine, and alanine (11). Under conditions of prolonged anaerobiosis, propionate is preferentially formed instead of succinate in mitochondria. Fumarate reduction is electron transfer chain coupled, and rhodoquinone serves as an electron donor to fumarate reductase, as in other anaerobic mitochondria (107, 531). The light gray rectangle highlights metabolic pathways preferentially employed during the early phase of anaerobiosis. The gray shading of lines is described in the legend of Fig. 4. Abbreviations: CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; RQ, rhodoquinone; C, cytochrome c; A, ATPase; FRD, fumarate reductase; [2B], acetate:succinate CoA-transferase (subfamily 1B); [3], succinyl-CoA synthetase; [5], malic enzyme; [6], pyruvate dehydrogenase complex; [8], fumarase; [9], pyruvate kinase; [10], phosphoenolpyruvate carboxykinase (ATP dependent); [11], malate dehydrogenase; [14], methylmalonyl-CoA mutase; [15], methylmalonyl-CoA epimerase; [16], propionyl-CoA carboxylase; [18], alanine aminotransferase; [19], aspartate aminotransferase; [26], strombine dehydrogenase; [27], octopine dehydrogenase. The photograph shows Mytilus edulis at the adult stage, with a length of ca. 6 cm. (Photograph by Louk Herber, Utrecht University, Netherlands.)

Fig 7

Major pathways of energy metabolism in mitochondria of the free-living annelid polychaete Arenicola marina (lugworm or sandworm). The worm inhabits intertidal sediments and produces ATP predominantly by an aerobic respiratory chain during high tide. At low tide, it is exposed for several hours to hypoxia (445) and also has to deal with high concentrations of H₂S (up to 2 mM), which is a potent inhibitor of respiratory complex IV (495, 496). A. marina protects itself intrinsically by means of membrane-bound flavoprotein sulfide:quinone oxidoreductase, which oxidizes hydrogen sulfide and transfers the electrons to the quinone pool. The generation of the final oxidized sulfur species, thiosulfate, can occur in an oxygen-dependent manner in cooperation with a sulfur dioxygenase and a sulfur transferase (189). The animal can also reduce fumarate in anaerobic respiration by utilizing rhodoquinone (531) and can produce acetate and ATP via substrate-level phosphorylation. The gray shading of lines is described in the legend of Fig. 4; the light gray rectangle highlights metabolic pathways preferentially employed during the early phase, as described in the legend of Fig. 6. Abbreviations: CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; RQ, rhodoquinone; C, cytochrome c; A, ATPase; FRD, fumarate reductase; SQR, sulfide:quinone oxidoreductase; ST, sulfur transferase; SD, sulfur dioxygenase; [2B], acetate:succinate CoA-transferase (subfamily 1B); [3], succinyl-CoA synthetase; [5], malic enzyme; [6], pyruvate dehydrogenase complex; [8], fumarase; [9], pyruvate kinase; [10], phosphoenolpyruvate carboxykinase (ATP dependent); [11], malate dehydrogenase; [14], methylmalonyl-CoA mutase; [15], methylmalonyl-CoA epimerase; [16], propionyl-CoA carboxylase; [18], alanine aminotransferase; [19], aspartate aminotransferase; [20], alanopine dehydrogenase; [26], strombine dehydrogenase. The photograph shows Arenicola marina at the adult stage, with a length of ca. 20 cm. (Photograph by Auguste Le Roux [http://commons.wikimedia.org/wiki/File:Arenicola_marina.JPG] [Wikimedia Commons].)

Fig 8

Major pathways of energy metabolism in mitochondria of the peanut worm, Sipunculus nudus. The map is redrawn based on data reported previously (169). This free-living worm inhabits sandy marine sediments ranging from intertidal to subtidal zones down to a 900-m depth, where it often encounters hypoxia. S. nudus deals with low oxygen by using the same pathways as those found in other marine invertebrates facing hypoxic or anoxic conditions. Fumarate serves as the terminal electron acceptor in the anaerobic branch of the respiratory chain and enables the synthesis of ATP not only via oxidative phosphorylation but also through propionate production from succinate. Additional ATP can be generated from succinyl-CoA in the pyruvate degradation pathway, leading to acetate as an end product (169). The gray shading of lines is described in the legend of Fig. 4. Abbreviations: CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; RQ, rhodoquinone; C, cytochrome c; A, ATPase; FRD, fumarate reductase; [2B], acetate:succinate CoA-transferase (subfamily 1B); [3], succinyl-CoA thiokinase; [5], malic enzyme; [6], pyruvate dehydrogenase complex; [8], fumarase; [9], pyruvate kinase; [11], malate dehydrogenase; [14], methylmalonyl-CoA mutase; [15], methylmalonyl-CoA epimerase; [16], propionyl-CoA carboxylase; [17], phosphoenolpyruvate carboxykinase (ITP/GTP dependent); [18], glutamate:pyruvate transaminase; [19], glutamate:oxaloacetate transaminase; [26], strombine dehydrogenase; [27], octopine dehydrogenase. The photograph shows Sipunculus nudus at the adult stage, with a length of ca. 15 cm. (Photograph courtesy of Matt du Fort.)

Fig 9

Mixed-acid fermentative metabolism of the hydrogenosome-bearing anaerobic chytridiomycete fungus Piromyces sp. E2, a rumen inhabitant. The map is redrawn based on data reported previously (47). Contrary to anaerobic parabasalian parasites, which use pyruvate:ferredoxin oxidoreductase, these common rumen inhabitants of many herbivorous mammals use pyruvate:formate lyase for pyruvate catabolism in their hydrogenosomes (47). Carbohydrate can also be metabolized to the end products succinate, lactate, formate, and ethanol in the cytosol. Bifunctional alcohol dehydrogenase E (ADHE), having both alcohol dehydrogenase and acetaldehyde dehydrogenase activities, mediates the cytosolic formation of ethanol. Note that metabolic maps reported previously by Yarlett et al. (570) and Boxma et al. (47) differ with respect to the involvement of ferredoxin in the H₂-producing reactions. With regard to the problematic circumstance that H₂ production from NAD(P)H is sketched, see the text. Abbreviations: Fd, ferredoxin; [2], acetate:succinate CoA-transferase; [3], succinyl-CoA synthetase; [4], hydrogenase; [5], malic enzyme; [8], fumarase; [9], pyruvate kinase; [10], phosphoenolpyruvate carboxykinase (ATP dependent); [11], malate dehydrogenase; [12], lactate dehydrogenase; [22], pyruvate:formate lyase; [23], NAD(P)H:ferredoxin oxidoreductase; [24], fumarate reductase (soluble); [25], alcohol dehydrogenase E. The photograph shows Piromyces sp. E2; the fluorescing region (sporangia and hyphae) is ca. 250 μm across. (Reprinted from reference with permission from Elsevier.)

Fig 10

Major metabolic pathways in the anaerobic intestinal parasite Entamoeba histolytica, a parasitic amoeboid protist and the causative agent of amoebiasis. The map is redrawn based on data reported previously (354). The amoebozoan infects humans and other primates; its energy metabolic pathways are localized in the cytosol, which also harbors mitosomes (5, 507). Pyruvate:ferredoxin oxidoreductase is used for pyruvate decarboxylation/oxidation, as in parabasalian parasites; ATP is synthesized through substrate-level phosphorylation via acetyl-CoA synthetase (ADP forming) (431). Although not involved in energy metabolism, mitosomes in E. histolytica harbor enzymes of sulfate activation as a major function (332). Abbreviations: Fd, ferredoxin; [1], pyruvate:ferredoxin oxidoreductase; [5], malic enzyme; [9], pyruvate kinase; [11], malate dehydrogenase; [18], alanine aminotransferase; [25], alcohol dehydrogenase E; [28], phosphoenolpyruvate carboxytransferase (PP_i dependent); [29], pyruvate:orthophosphate dikinase; [30], acetyl-CoA synthetase (ADP forming). The photograph shows an Entamoeba histolytica trophozoite with an ingested erythrocyte, with a length of ca. 20 μm. (Photograph from the Centers for Disease Control and Prevention [CDC], Atlanta, GA.)

Fig 11

Major pathways of energy metabolism in a parasite of the small intestine, Giardia intestinalis (also called Giardia lamblia), a flagellated protist parasite and the causative agent of giardiasis, a diarrheal infection of humans. The map is redrawn based on data reported previously (354), taking hydrogenase activity into account (287). Giardia mitosomes are not directly involved in energy metabolism but are involved in FeS cluster biogenesis (508). In Giardia, as in Entamoeba, the typically hydrogenosomal (and sometimes mitochondrial) enzymes PFO and [Fe]-Hyd have been recompartmentalized to the cytosol during evolution. G. lamblia produces molecular hydrogen under strictly anoxic conditions (287), as indicated by the dashed line. Abbreviations: Fd, ferredoxin; [1], pyruvate:ferredoxin oxidoreductase; [4], hydrogenase; [5], malic enzyme; [9], pyruvate kinase; [11], malate dehydrogenase; [17], phosphoenolpyruvate carboxykinase (GTP dependent); [18], alanine aminotransferase; [25], alcohol dehydrogenase E; [29], pyruvate:orthophosphate dikinase; [30], acetyl-CoA synthetase (ADP forming). The photograph shows Giardia intestinalis at the trophozoite stage, with a length of 15 to 20 μm. (Photograph from the CDC, Atlanta, GA.)

Fig 12

Major pathways of anaerobic, molecular hydrogen-producing, fermentative metabolism in hydrogenosomes of the flagellated protist parasite Trichomonas vaginalis, a sexually transmitted pathogen of the human urogenital tract. The map is redrawn based on data reported previously (209, 349). Hydrogenosomal pyruvate breakdown involves pyruvate:ferredoxin oxidoreductase and functional 51-kDa and 24-kDa subunits of the NADH dehydrogenase module in complex I, which reoxidize NADH stemming from malate oxidation (71, 209, 349). The 51-kDa and 24-kDa subunits of mitochondrial complex I function in association with [Fe]-Hyd in Trichomonas (209), possibly in a manner similar to that of the trimeric [Fe]-Hyd of Thermotoga (448) (see the text). Additional major end products of cytosolic fermentations in T. vaginalis include alanine, lactate, ethanol, and glycerol (469). Abbreviations: Fd, ferredoxin; [1], pyruvate:ferredoxin oxidoreductase; [2C], acetate:succinate CoA-transferase subfamily 1C (503); [3], succinyl-CoA synthetase; [4], hydrogenase; [5], malic enzyme; [9], pyruvate kinase; [11], malate dehydrogenase; [12], lactate dehydrogenase; [17], phosphoenolpyruvate carboxykinase (GTP dependent); [18], alanine aminotransferase; [31], 51-kDa and 24-kDa subunits of the NADH dehydrogenase module of complex I; [32], pyruvate decarboxylase; [34], alcohol dehydrogenase (NADPH dependent); [49], glycerol-3-phosphate dehydrogenase; [50], glycerol-3-phosphatase. The light micrograph shows Trichomonas vaginalis at the trophozoite stage, with a length of 7 to 30 μm (Photograph from the CDC, Atlanta, GA.) The transmission electron micrograph shows hydrogenosomes (H) and the nucleus (N). (Photograph courtesy of Kathrin Bolte, University of Marburg, Germany.)

Fig 13

Major pathways of energy metabolism leading to acetate end product formation in hydrogenosomes of the flagellated protist parasite Tritrichomonas foetus, a pathogen of the bovine urogenital tract. The map was redrawn based on data reported previously (354, 469). The hydrogenosomal fermentation of T. foetus is very similar to that of Trichomonas vaginalis. Pyruvate:ferredoxin oxidoreductase mediates the generation of acetyl-CoA, and protons serve as the terminal electron acceptor; NADPH can be reoxidized through alcohol dehydrogenase (469). The 51- and 24-kDa subunits of complex I (209) have so far not been identified in T. foetus. Cytosolic succinate production through soluble fumarate reductase is lacking in metronidazole-resistant Tritrichomonas foetus strains (256). Abbreviations: Fd, ferredoxin; [1], pyruvate:ferredoxin oxidoreductase; [2C], acetate:succinate CoA-transferase subfamily 1C (503); [3], succinyl-CoA synthetase; [4], hydrogenase; [5], malic enzyme; [8], fumarase; [9], pyruvate kinase; [11], malate dehydrogenase; [17], phosphoenolpyruvate carboxykinase (GTP dependent); [18], alanine aminotransferase; [24], fumarate reductase (soluble); [32], pyruvate decarboxylase; [34], alcohol dehydrogenase (NADPH dependent); [49], glycerol-3-phosphate dehydrogenase; [50], glycerol-3-phosphatase. The photograph shows Tritrichomonas foetus at the trophozoite stage, with a length of ca. 15 μm. (Reprinted from reference with permission from Elsevier.)

Fig 14

Major pathways of compartmentalized energy metabolism in bloodstream (a) and procyclic (b) life cycle stages of the flagellated and fully oxygen-dependent protist parasite Trypanosoma brucei. The maps are redrawn based on data reported previously (54, 500a, 538). (a) In bloodstream forms, Trypanosoma brucei mitochondria help to maintain cell redox homeostasis via glycerol-3-phosphate dehydrogenase and alternative oxidase, which, together with ubiquinone, constitute the whole O₂-consuming respiratory chain, which does not, however, produce ATP. The inner mitochondrial membrane ATPase works in reverse; that is, it pumps protons to the intermembrane space while hydrolyzing ATP to ADP. The major metabolic end product pyruvate is generated in the cytosol. The photograph shows a long slender bloodstream form of Trypanosoma brucei with a length of ca. 25 μm (circles in the background are erythrocytes). (Photograph by Rob Koelewijn, Harbor Hospital, Rotterdam, Netherlands.) (b) Energy metabolism is more complex in procyclic T. brucei stages. Succinate is generated both in glycosomes by a soluble fumarate reductase and in mitochondria via a recently functionally characterized soluble mitochondrial fumarate reductase (90). Acetate is produced in mitochondria mainly by means of the acetate:succinate CoA-transferase/succinyl-CoA synthetase cycle, but an alternative pathway is also present (413). In T. brucei, no lactate is formed from pyruvate (538); instead, another end product commonly found in facultative anaerobic animals, alanine, is produced by alanine aminotransferase. The respiratory chain contains not only an alternative oxidase and glycerol-3-phosphate dehydrogenase but also an alternative rotenone-insensitive NADH dehydrogenase (90). Abbreviations: CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; C, cytochrome c; A, ATPase; AOX, alternative oxidase; AN, alternative, rotenone-insensitive NADH dehydrogenase; GPDH, glycerol-3-phosphate dehydrogenase; G3P, glycerol-3-phosphate; DHAP, dihydroxyacetone phosphate; GAP, glyceraldehyde-3-phosphate; PGA, 3-phosphoglycerate; [2A], acetate:succinate CoA-transferase (subfamily 1A); [3], succinyl-CoA synthetase; [5], malic enzyme; [6], pyruvate dehydrogenase complex; [8], fumarase; [9], pyruvate kinase; [10], phosphoenolpyruvate carboxykinase (ATP dependent); [11], malate dehydrogenase; [18], alanine aminotransferase; [24], fumarate reductase (soluble, NADH dependent); [42], α-ketoglutarate dehydrogenase; [43], citrate synthase.

Fig 15

Wax ester fermentation in mitochondria of the facultatively anaerobic photosynthetic flagellate Euglena gracilis. The map is redrawn based on data reported previously (514). Under anaerobic conditions, this photosynthetic euglenid, which acquired its plastids through secondary endosymbiosis, uses acetyl-CoA produced by pyruvate:NADP⁺ oxidoreductase (419) as the terminal electron acceptor, leading to the formation of an unusual end product among eukaryotes: wax esters (213, 443, 514). Mitochondrial wax ester fermentation includes anaerobic fumarate respiration and the same propionyl-CoA formation pathway as the one found in mitochondria of facultative anaerobic animals excreting and/or accumulating propionate. Abbreviations: CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; RQ, rhodoquinone; C, cytochrome c; A, ATPase; FRD, fumarate reductase; [3], succinyl-CoA synthetase; [5], malic enzyme; [6], pyruvate dehydrogenase complex; [7], pyruvate:NADP⁺ oxidoreductase; [8], fumarase; [9], pyruvate kinase; [10], phosphoenolpyruvate carboxykinase (ATP dependent); [11], malate dehydrogenase; [14], methylmalonyl-CoA mutase; [15], methylmalonyl-CoA epimerase/racemase; [16], propionyl-CoA carboxylase; [35], α-ketoacyl synthase; [36], β-ketoacyl reductase; [37], β-hydroxyacyl dehydrogenase; [38], trans-2-enoyl-CoA reductase. (Photograph of Euglena gracilis [American Type Culture Collection] by D. J. Patterson, L. Amaral-Zettler, M. Peglar, and T. Nerad, reprinted from the Encyclopedia of Life [http://eol.org/pages/918864/overview], which was published under a Creative Commons license.)

Fig 16

Tentative map of major pathways of energy metabolism in hydrogen-producing mitochondria of the anaerobic ciliate Nyctotherus ovalis. The map is redrawn based on data reported previously (49). The ciliate lives in the hindgut of cockroaches and harbors organelles that provided a link between mitochondria and hydrogenosomes (49). The Krebs cycle is incomplete and is likely used in the reductive direction (177). A proton gradient is generated, probably by a functional respiratory complex I, which passes the electrons from the NADH pool through rhodoquinone to complex II, acting as fumarate reductase synthesizing succinate (101). Redox balance is also achieved with the help of hydrogenase, releasing molecular hydrogen, hence the term hydrogen-producing anaerobic mitochondria (49). ATP can be synthesized by substrate-level phosphorylation, producing acetate. Abbreviations: CI, respiratory complex I; RQ, rhodoquinone; CII, fumarate reductase/succinate dehydrogenase; [2A], acetate:succinate CoA-transferase subfamily 1A (503); [3], succinyl-CoA synthetase; [4], hydrogenase; [5], malic enzyme; [6], pyruvate dehydrogenase complex; [8], fumarase (predicted); [9], pyruvate kinase; [12], lactate dehydrogenase; [18], alanine aminotransferase; [32], pyruvate decarboxylase; [33], alcohol dehydrogenase (NADH dependent); [42], α-ketoglutarate dehydrogenase (predicted); [41], glutamate dehydrogenase. The photograph on the left shows Nyctotherus ovalis, with a length of ca. 80 μm. The hydrogenosomes (H) are surrounded by endosymbiotic methane-producing archaebacteria (dark spots). N, macronucleus; n, micronucleus; V, vacuole. (Reprinted from reference with permission of Macmillan Publishers Ltd.) The right photograph shows a closeup view of a Nyctotherus hydrogenosome (H) and an associated methanogen (M). (Reprinted from reference with permission of Macmillan Publishers Ltd.)

Fig 17

Putative major pathways of anaerobic energy metabolism in the organelle of mitochondrial origin in Blastocystis hominis, a protozoan parasite and a common inhabitant of the human gastrointestinal tract. The map is redrawn based on data reported previously (270). The conversion of pyruvate to acetyl-CoA is carried out by pyruvate:NADP⁺ oxidoreductase (PNO) (270), the same fusion protein found in Euglena gracilis. The transformation of acetyl-CoA to acetate leads to the synthesis of ATP via substrate-level phosphorylation. The alternative oxidase might help Blastocystis cope with oxygen stress conditions in the host intestine and prevent the formation of reactive oxygen species. Hydrogen production has not yet been shown for Blastocystis (as indicated by a question mark), although [Fe]-Hyd localizes to the organelle (468). In this sense, the mitochondria of Blastocystis resemble the hydrogen-producing mitochondria found in the ciliate Nyctotherus ovalis and represent another example demonstrating that mitochondria and hydrogenosomes utilize a common enzyme toolkit. The question mark indicates another possible pathway (103, 270, 468). Abbreviations: CI, respiratory complex I; Q?, quinone (of an uncertain nature); AOX, alternative oxidase; [2B, C], acetate:succinate CoA-transferase (members of subfamilies 1B and 1C [503] are present in the genome); [3], succinyl-CoA synthetase; [4], hydrogenase; [5], malic enzyme; [7], pyruvate:NADP⁺ oxidoreductase; [9], pyruvate kinase; [11], malate dehydrogenase; [12], lactate dehydrogenase; [17], phosphoenolpyruvate carboxykinase (GTP dependent). The photograph shows Blastocystis sp. at the trophozoite stage, with a length of ca. 35 μm. (Photograph courtesy of C. G. Clark, London School of Hygiene and Tropical Medicine, London, United Kingdom.)

Fig 18

Major pathways of anaerobic energy metabolism in the green alga Chlamydomonas reinhardtii, whereby the suggestions for their compartmentation are still putative. The map was redrawn based on data reported previously (331). This typical soil inhabitant can produce oxygen but is often faced with anoxic conditions in nature (331). When grown anaerobically, C. reinhardtii generates virtually the same end products as those generated by Trichomonas but utilizing pyruvate:formate lyase, pyruvate:ferredoxin oxidoreductase, lactate dehydrogenase, and pyruvate decarboxylase (357). The localization of fermentative pathways leading to the formation of acetate, ethanol, and formate end products has yet to be fully clarified. Abbreviations: CI to CIV, respiratory complexes I to IV; UQ, ubiquinone; C, cytochrome c; A, ATPase; Fd, ferredoxin; [1], pyruvate:ferredoxin oxidoreductase; [4], hydrogenase; [6], pyruvate dehydrogenase complex; [9], pyruvate kinase; [12], lactate dehydrogenase; [22], pyruvate:formate lyase; [25], alcohol dehydrogenase E; [32], pyruvate decarboxylase; [33], alcohol dehydrogenase (NADH dependent); [39], phosphotransacetylase; [40], acetate kinase. The photograph shows Chlamydomonas reinhardtii, with a length of ca. 10 μm. (Photograph courtesy of Cytographics Inc., Victoria, Australia.)

Fig 19

Presence or absence of genes for anaerobic energy metabolism across major eukaryotic groups. For each protein, sequences of suitable functionally characterized enzymes were used to search the databases via BLAST (12); the presence or absence of the gene and the average amino acid identity in pairwise local alignments between the eukaryotic and prokaryotic proteins are color coded as shown. Note the pattern of the presence or absence of genes for anaerobic energy metabolism and glycolysis and those most frequent in archaebacteria across major groups of eukaryota, proteobacteria, and archaea (see Table S1 in the supplemental material). For each protein, suitable functionally characterized sequences (Table 1) were used to search the available sequence data for the organisms shown via BLAST (12). The presence of a certain gene was scored as the best match at an E value threshold of 10⁻¹⁰. For the eukaryotes, the colors show if the genes were found by EST data (light blue) or protein data (green) or comprised the searched sequence itself (dark blue). In the cases of the prokaryotes, the colors indicate the average global pairwise identity of the homolog in the corresponding prokaryote to all eukaryotic homologs (coded as shown). The archaeal genes were included as a control. Eukaryotic proteomes and ESTs were downloaded from RefSeq, GenBank, and, in the case of Fusarium oxysporum, the Broad Institute website (www.broadinstitute.org/). Prokaryotic proteomes were downloaded from RefSeq, version March 2011. Data for the archaebacterial genes were reported previously (139) and serve to demonstrate that archaebacterially related genes in eukaryotes are readily detected.

Fig 20

Summary of major events in Earth history relating to the appearance of eukaryotic groups and the appearance of O₂ in the atmosphere and in marine environments (see the text, and see also references and 386). Ma, million years. (Reprinted from reference with permission.)