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Hideto Miyoshi - One of the best experts on this subject based on the ideXlab platform.
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Pinpoint Chemical Modification of Asp160 in the 49 kDa Subunit of Bovine Mitochondrial Complex I via a Combination of Ligand-Directed Tosyl Chemistry and Click Chemistry
2014Co-Authors: Takahiro Masuya, Masatoshi Murai, Hironobu Morisaka, Hideto MiyoshiAbstract:Through a ligand-directed tosyl (LDT) chemistry strategy using the synthetic Acetogenin ligand AL1, we succeeded in the pinpoint alkynylation (−CCH) of Asp160 in the 49 kDa subunit of bovine complex I, which may be located in the inner part of the putative quinone binding cavity of the enzyme [Masuya, T., et al. (2014) Biochemistry, 53, 2307–2317]. This study provided a promising technique for diverse chemical modifications of complex I. To further improve this technique for its adaptation to intact complex I, we here synthesized the new Acetogenin ligand AL2, possessing an azido (−N3) group in place of the terminal alkyne in AL1, and attempted the pinpoint azidation of complex I in bovine heart submitochondrial particles. Careful proteomic analyses revealed that, just as in the case of AL1, azidation occurred at 49 kDa Asp160 with a reaction yield of ∼50%, verifying the high site specificity of our LDT chemistry using Acetogenin ligands. This finding prompted us to speculate that a reactivity of the azido group incorporated into Asp160 (Asp160-N3) against externally added chemicals can be employed to characterize the structural features of the quinone/inhibitor binding cavity. Consequently, we used a ring-strained cycloalkyne possessing a rhodamine fluorophore (TAMRA-DIBO), which can covalently attach to an azido group via so-called click chemistry without Cu1+ catalysis, as the reaction partner of Asp160-N3. We found that bulky TAMRA-DIBO is capable of reacting directly with Asp160-N3 in intact complex I. Unexpectedly, the presence of an excess amount of short-chain ubiquinones as well as some strong inhibitors (e.g., quinazoline and fenpyroximate) did not interfere with the reaction between TAMRA-DIBO and Asp160-N3; nevertheless, bullatacin, a member of the natural Acetogenins, markedly interfered with this reaction. Taking the marked bulkiness of TAMRA-DIBO into consideration, it appears to be difficult to reconcile these results with the proposal that only a narrow entry point accessing to the quinone/inhibitor binding cavity exists in complex I [Baradaran, R., et al. (2013) Nature, 494, 443–448]; rather, they suggest that there may be another access path for TAMRA-DIBO to the cavity
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critical role of a methyl group on the γ lactone ring of annonaceous Acetogenins in the potent inhibition of mitochondrial complex i
Bioorganic & Medicinal Chemistry Letters, 2013Co-Authors: Naoto Kojima, Masato Abe, Yuki Suga, Kazufumi Ohtsuki, Tetsuaki Tanaka, Hiroki Iwasaki, Masayuki Yamashita, Hideto MiyoshiAbstract:Abstract C34-epi and C34-epi-C35-trifluoro analogues of solamin, a mono-THF annonaceous Acetogenin, were synthesized. Their inhibitory activity, along with previously synthesized analogues (C35-fluoro, C35-difluoro, and C35-trifluorosolamins), against bovine mitochondrial NADH–ubiquinone oxidoreductase (complex I) was determined. The present study revealed that the methyl group on the γ-lactone moiety is critical to the potent inhibition of complex I by natural Acetogenins.
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bis thf motif of Acetogenin binds to the third matrix side loop of nd1 subunit in mitochondrial nadh ubiquinone oxidoreductase
Biochimica et Biophysica Acta, 2011Co-Authors: Sayo Nakanishi, Masato Abe, Shuhei Yamamoto, Masatoshi Murai, Hideto MiyoshiAbstract:Abstract Natural Acetogenins are among the most potent inhibitors of bovine heart mitochondrial NADH-ubiquinone oxidoreductase (complex I). Our photoaffinity labeling study suggested that the hydroxylated bis-THF ring moiety of Acetogenins binds at “site A” in the third matrix-side loop connecting the fifth and sixth transmembrane helices in the ND1 subunit [Kakutani et al. (2010) Biochemistry 49 , 4794–4803]. Nevertheless, since this proposition was led using a photoreactive Δlac-Acetogenin derivative, it needs to be directly verified using a natural Acetogenin-type probe. We therefore conducted photoaffinity labeling using a photoreactive natural Acetogenin mimic ([ 125 I] d i a zinylated n atural a cetogenin, [ 125 I]DANA), which has a small photolabile diazirine group, in place of a hydroxy group, attached to the bis-THF ring moiety. Analysis of the photocross-linked protein in bovine heart submitochondrial particles unambiguously revealed that [ 125 I]DANA binds to the membrane subunit ND1 with high specificity. The photocross-linking was completely blocked in the presence of just a 5-fold excess of bullatacin, indicating that [ 125 I]DANA is an excellent mimic of natural Acetogenins and hence binds to the site that accommodates natural products. Careful examination of the fragmentation patterns of the cross-linked ND1 generated by different proteases and their combinations indicated that the cross-linked residue is predominantly located at the supposed site A in the third matrix-side loop.
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synthesis and characterization of photoaffinity probe of Acetogenin a strong inhibitor of mitochondrial complex i
Tetrahedron Letters, 2011Co-Authors: Shuhei Yamamoto, Masato Abe, Sayo Nakanishi, Masatoshi Murai, Hideto MiyoshiAbstract:Abstract Acetogenins are valuable inhibitor probes to get an insight into the structural and functional properties of mitochondrial NADH-ubiquinone oxidoreductase (complex I). We synthesized a photoreactive Acetogenin mimic ([125I]DANA) which retained a strong inhibitory activity. The preliminary photoaffinity labeling with bovine heart submitochondrial particles revealed that [125I]DANA binds to the ND1 subunit among 45 different subunits with high specificity.
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exploring the binding site of δlac Acetogenin in bovine heart mitochondrial nadh ubiquinone oxidoreductase
Biochemistry, 2010Co-Authors: Nobuyuki Kakutani, Masatoshi Murai, Naoto Sakiyama, Hideto MiyoshiAbstract:Biochemical characterization of the inhibition mechanism of Deltalac-Acetogenins synthesized in our laboratory indicated that they are a new type of inhibitor of bovine heart mitochondrial NADH-ubiquinone oxidoreductase (complex I) [Murai, M., et al. (2006) Biochemistry 45, 9778-9787]. To identify the binding site of Deltalac-Acetogenins with a photoaffinity labeling technique, we synthesized a photoreactive Deltalac-Acetogenin ([(125)I]diazinylated Deltalac-Acetogenin, [(125)I]DAA) which has a small photoreactive diazirine group attached to a pharmacophore, the bis-THF ring moiety. Characterization of the inhibitory effects of DAA on bovine complex I revealed unique features specific to, though not completely the same as those of, the original Deltalac-Acetogenin. Using [(125)I]DAA, we carried out photoaffinity labeling with bovine heart submitochondrial particles. Analysis of the photo-cross-linked protein by Western blotting and immunoprecipitation revealed that [(125)I]DAA binds to the membrane subunit ND1 with high specificity. The photo-cross-linking to the ND1 subunit was suppressed by an exogenous short-chain ubiquinone (Q(2)) in a concentration-dependent manner. Careful examination of the fragmentation patterns of the cross-linked ND1 generated by limited proteolysis using lysylendopeptidase, endoprotease Asp-N, or trypsin and their changes in the presence of the original Deltalac-Acetogenin strongly suggested that the cross-linked residues are located at two different sites in the third matrix-side loop connecting the fifth and sixth transmembrane helices.
Jerry L Mclaughlin - One of the best experts on this subject based on the ideXlab platform.
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Chemical defense in the zebra swallowtail butterfly, Eurytides marcellus, involving annonaceous Acetogenins
Journal of Natural Products, 1999Co-Authors: John M. Martin, Stephen R. Madigosky, Jinn Wu, Zhe-ming Gu, Dawei Zhou, Jerry L MclaughlinAbstract:Few herbivores feed on the foliage of the North American paw paw tree, Asimina triloba; notable exceptions are the larvae of the zebra swallowtail butterfly, Eurytides marcellus. Toxic annonaceous Acetogenins, produced by A. triloba, are responsible for the relative unpalatability of the leaves. Acetogenins found in A. triloba extracts are potent pesticidal and antineoplastic agents and have emetic activity in vertebrates. In this study, partitioned aqueous MeOH fractions of the bioactive CH2Cl2 extracts, of freeze-dried and pulverized larvae, and of mature butterflies revealed Acetogenin content through the use of HPLC coupled to tandem MS (LC-MS/MS). This sensitive technique provides an uncomplicated method for the detection of trace compounds and, in this instance, has confirmed tissue presence of Acetogenins that serve a probable role as chemical defense agents against bird predation in zebra swallowtail larvae and adults.
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Membrane conformations and their relation to cytotoxicity of asimicin and its analogues.
Biochemistry, 1998Co-Authors: Hiroko Shimada, John B. Grutzner, John F. Kozlowski, Jerry L MclaughlinAbstract:Certain plant species belonging to the family Annonaceae produce Annonaceous Acetogenins, which are a unique class of long-chain fatty acid derivatives with potent cytotoxicity. Putative protein targets of the Acetogenins are membrane-associated proteins, including complex I. Asimicin and its analogues constitute a class of Annonaceous Acetogenins containing two tetrahydrofuran (THF) rings with hydrocarbon chains tethered to each ring; an alpha,beta-unsaturated gamma-lactone ring is terminal to one of the alkyl chains. The compounds examined in this study differ in the length of the alkyl chain between the THF rings and the lactone ring. The positions of both the THF and the lactone rings within liposomal membranes were determined by proton (1H) nuclear magnetic resonance spectroscopy. The depth of membrane penetration of Acetogenins, coupled to membrane diffusion, controls the conformation of Acetogenins as they diffuse to an active site. Based on 1H intermolecular nuclear Overhauser effects (NOEs), the THF rings of all Acetogenins studied reside near the polar interfacial head group region of the DMPC. This was corroborated by 1H two-dimensional NOE spectroscopy and differential scanning calorimetry studies. The 1H difference NOE spectra indicated that the lactone rings of asimicin and parviflorin, the latter of which has two fewer carbons in its alkyl chain, are located below the glycerol backbone in the membrane. In contrast with asimicin and parviflorin, the lactone ring of longimicin B, an asimicin analogue with an alkyl chain four carbons shorter, resides close to the midplane in the membrane. This was corroborated by manganese-induced broadening studies. Since the THF rings are located near the center of the Acetogenin molecules and the lactone ring is terminal to a long alkyl chain, these observations indicate that an asimicin-type Acetogenin can be in either sickle-shaped or U-shaped conformations, depending on the length of the alkyl chain between the THF rings and the lactone ring. Interestingly, longimicin B does not exhibit significant cytotoxicity, but parviflorin is as cytotoxic as asimicin. The cytotoxicity of the asimicin-type of Acetogenins would seem to be strongly related to the membrane conformation. This is the first report elucidating the conformation of Annonaceous Acetogenins in membranes.
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the annonaceous Acetogenin bullatacin is cytotoxic against multidrug resistant human mammary adenocarcinoma cells
Cancer Letters, 1997Co-Authors: Nicholas H Oberlies, Vicki L Croy, Marietta L Harrison, Jerry L MclaughlinAbstract:Cytotoxic effects of the Annonaceous Acetogenin, bullatacin, were studied in multidrug-resistant (MDR) human mammary adenocarcinoma (MCF-7/Adr) cells vs. the parental non-resistant wild type (MCF-7/wt) cells. Bullatacin was effectively cytotoxic to the MCF-7/Adr cells while it was more cytostatic to the MCF-7/wt cells. ATP depletion is the mode of action of the Annonaceous Acetogenins, and these agents offer a special advantage in the chemotherapeutic treatment of MDR tumors that have ATP-dependent mechanisms.
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the absolute configuration of adjacent bis thf Acetogenins and asiminocin a novel highly potent asimicin isomer from asimina triloba
Bioorganic & Medicinal Chemistry, 1996Co-Authors: Gengxian Zhao, Jinfeng Chao, Matthew J Rieser, Lu Zeng, Jerry L MclaughlinAbstract:Abstract A novel Acetogenin, asiminocin (1), was isolated by activity-directed fractionation from the stem bark of the paw paw tree, Asimina triloba . By spectral and chemical methods, 1 was identified as (30 S )-hydroxy-4-deoxyasimicin. The absolute configuration of 1, along with those of previously reported Acetogenins asimin, asiminacin, bullatin, (30 S )-bullanin, and (30 R ) bullanin, was determined by Mosher ester methodology. Compound 1 was highly inhibitory to three human solid tumor cell lines with over a billion times the potency of adriamycin.
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SARs of annonaceous Acetogenins in rat liver mitochondria.
Natural toxins, 1996Co-Authors: Dorothée Alfonso, Holly A. Johnson, Trina Colman-saizarbitoria, Christopher P. Presley, George P. Mccabe, Jerry L MclaughlinAbstract:The Annonaceous Acetogenins represent a class of compounds with diverse bioactivities, including promising cytotoxicites. These are due, at least in part, to inhibition of complex I in the oxidative phosphorylation pathway in mitochondria. Fourteen Annonaceous Acetogenins were tested in a rat liver mitochondrial oxygen uptake assay to probe additional structure-activity relationships. In this subcellular assay, the activity of non-adjacent bis-THF ring Acetogenins depends on the distance between the two THF rings; the activity decreases to that of a mono-THF ring Acetogenin if the distance is too long. When one THF ring is replaced with a tetrahydropyran ring, the activity remains comparable. The configuration of the THF ring, in mono ring compounds, seems to be more important than stereochemical differences in the rings of adjacent bis-THF ring compounds. Bullatacin, an adjacent bis-THF ring Acetogenin, was used as a standard compound in every run to normalize the data.
William J Keller - One of the best experts on this subject based on the ideXlab platform.
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Identification of annonaceous Acetogenins in the ripe fruit of the North American pawpaw ( Asimina triloba ).
Journal of Agricultural and Food Chemistry, 2009Co-Authors: Kirk W. Pomper, Jeremiah D. Lowe, Sheri B. Crabtree, William J KellerAbstract:The North American pawpaw [ Asimina triloba (L.) Dunal] is a tree fruit in the early stages of commercial production in the United States. This plant contains annonaceous Acetogenins in the twigs, unripe fruit, seeds, roots, and bark tissues, which display antitumor, pesticidal, antimalarial, anthelmintic, piscicidal, antiviral, and antimicrobial effects, suggesting many potentially useful applications. However, commercial development of these compounds, based on twig extracts, has been problematic due to limited availability of biomass for extraction. Additionally, Acetogenin compounds contained in fruit of pawpaw relatives (soursop or Annona muricata ) and tea made from the leaves of these plants may lead to an increased risk of atypical Parkinsonism later in life with overconsumption of these compounds. Therefore, the objectives of this study were (1) to determine if extracts of ripe pawpaw fruit pulp displayed Acetogenin activity, (2) to identify potential Acetogenin compounds in the fruit tissue, and (3) to determine if the Acetogenin activity varied in diverse pawpaw genotypes and closely related Annona species. Extracts of ripe fruit had total extract weights and bioactivity using the brine shrimp bioassay similar to those from 'NC-1' pawpaw twig tissue. Pulp from soursop, cherimoya, and several additional pawpaw cultivars ('Mitchell', 'Overleese', 'NC-1','Zimmerman', 'Wells', and 'Sunflower') also displayed bioactivity, but peach or banana pulp did not. Ripe pawpaw pulp extract subjected to HPLC-MS analysis identified three prominent Acetogenins: asimicin, bullatacin, and bullatalicin. This study points to pawpaw fruit pulp serving as a new biomass source for the extraction of Acetogenin compounds for product development. An assessment of the potential human health risk of overconsumption of fruit and Acetogenin bioavailability and degradation studies should be pursued.
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identification of annonaceous Acetogenins in the ripe fruit of the north american pawpaw asimina triloba
Journal of Agricultural and Food Chemistry, 2009Co-Authors: Kirk W. Pomper, Jeremiah D. Lowe, Sheri B. Crabtree, William J KellerAbstract:The North American pawpaw [Asimina triloba (L.) Dunal] is a tree fruit in the early stages of commercial production in the United States. This plant contains annonaceous Acetogenins in the twigs, unripe fruit, seeds, roots, and bark tissues, which display antitumor, pesticidal, antimalarial, anthelmintic, piscicidal, antiviral, and antimicrobial effects, suggesting many potentially useful applications. However, commercial development of these compounds, based on twig extracts, has been problematic due to limited availability of biomass for extraction. Additionally, Acetogenin compounds contained in fruit of pawpaw relatives (soursop or Annona muricata) and tea made from the leaves of these plants may lead to an increased risk of atypical Parkinsonism later in life with overconsumption of these compounds. Therefore, the objectives of this study were (1) to determine if extracts of ripe pawpaw fruit pulp displayed Acetogenin activity, (2) to identify potential Acetogenin compounds in the fruit tissue, and (...
Alicia Bardon - One of the best experts on this subject based on the ideXlab platform.
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tucupentol a novel mono tetrahydrofuranic Acetogenin from annona montana as a potent inhibitor of mitochondrial complex i
Chemistry & Biodiversity, 2009Co-Authors: Olga Alvarez Colom, Adriana Neske, Nadia Chahboune, Carmen M Zafrapolo, Alicia BardonAbstract:Ten Acetogenins, one of them new, were isolated from leaves and twigs of a Bolivian collection of Annona montana. The new compound that we named tucupentol (1) is a mono-tetrahydrofuran-pentahydroxy-Acetogenin. The inhibitory potency of tucupentol (1) on the mitochondrial complex I was evaluated, and this activity was compared with that of the known Acetogenins, annonacin-A, cis-annonacin-10-one, aromin, and gigantetronenin, also isolated from this plant material. The mentioned Acetogenins acted as selective inhibitors of mitochondrial complex I in the 0.8-5.4-nM range.
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toxic effects of annonaceous Acetogenins from annona cherimolia magnoliales annonaceae on spodoptera frugiperda lepidoptera noctuidae
Journal of Pest Science, 2007Co-Authors: Alvarez O Colom, Adriana Neske, Susana Popich, Alicia BardonAbstract:Plants belonging to the family Annonaceae have been commonly described in traditional medicine as remedies against head lice, and for their insecticidal properties. Characteristic constituents from a few genera of these plants are the annonaceous Acetogenins. Fourteen annonaceous Acetogenins have been isolated from our Argentine collection of the seeds of A. cherimolia. We report herein the antifeedant and insecticidal effects of nine of those Acetogenins on Spodoptera frugiperda (J. E. Smith). The Acetogenin squamocin, one of the major constituents of the extract, displayed toxic effects on early larval instars when incorporated to the larval diet at a dose of 50 μg per g of diet. The remaining annonaceous Acetogenins tested, itrabin, asimicin, neoanonin, cherimolin-1, cherimolin-2, almunequin, motrilin, and tucumanin produced pupal mortality and adult malformations leading to death, when incorporated to the larval diet at the same dose. The evaluation of indices of food consumption, growth, and food utilization indicated that squamocin was the only tested Acetogenin to produce significant decrease in the growth rate and to reduce the efficiency with which larvae converted ingested food into biomass. All the Acetogenins produced more than 80% pupal mortality with no dependence on the position of the THF rings or the number and location of the OH groups.
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toxic effects of annonaceous Acetogenins from annona cherimolia magnoliales annonaceae on spodoptera frugiperda lepidoptera noctuidae
Journal of Pest Science, 2007Co-Authors: Alvarez O Colom, Adriana Neske, Susana Popich, Alicia BardonAbstract:Plants belonging to the family Annonaceae have been commonly described in traditional medicine as remedies against head lice, and for their insecticidal properties. Characteristic constituents from a few genera of these plants are the annonaceous Acetogenins. Fourteen annonaceous Acetogenins have been isolated from our Argentine collection of the seeds of A. cherimolia. We report herein the antifeedant and insecticidal effects of nine of those Acetogenins on Spodoptera frugiperda (J. E. Smith). The Acetogenin squamocin, one of the major constituents of the extract, displayed toxic effects on early larval instars when incorporated to the larval diet at a dose of 50 μg per g of diet. The remaining annonaceous Acetogenins tested, itrabin, asimicin, neoanonin, cherimolin-1, cherimolin-2, almunequin, motrilin, and tucumanin produced pupal mortality and adult malformations leading to death, when incorporated to the larval diet at the same dose. The evaluation of indices of food consumption, growth, and food utilization indicated that squamocin was the only tested Acetogenin to produce significant decrease in the growth rate and to reduce the efficiency with which larvae converted ingested food into biomass. All the Acetogenins produced more than 80% pupal mortality with no dependence on the position of the THF rings or the number and location of the OH groups.
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tucumanin a β hydroxy γ lactone bistetrahydrofuranic Acetogenin from annona cherimolia is a potent inhibitor of mitochondrial complex i
Planta Medica, 2004Co-Authors: Isabel Barrachina, Adriana Neske, Alicia Bardon, Nadia Chahboune, Susana Granell, Almudena Bermejo, Noureddine El Aouad, Olga Alvarez, Carmen M ZafrapoloAbstract:A new beta-hydroxy-gamma-methyl-gamma-lactone bistetrahydrofuranic Acetogenin, tucumanin, with the infrequent symmetrical threo/trans/threo/trans/threo relative configuration at the tetrahydrofuran rings was isolated from Annona cherimolia (Annonaceae) seeds. The inhibitory potency on the mitochondrial complex I of Acetogenins with this relative configuration (tucumanin and asimicin)was compared with that shown by the corresponding pairs with an asymmetrical threo/trans/threo/trans/erythro relative configuration (laherradurin/rolliniastatin-2, and itrabin/molvizarin). All these compounds act as selective inhibitors of mitochondrial complex I in the 0.18 - 1.55 nM range.
Masato Abe - One of the best experts on this subject based on the ideXlab platform.
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critical role of a methyl group on the γ lactone ring of annonaceous Acetogenins in the potent inhibition of mitochondrial complex i
Bioorganic & Medicinal Chemistry Letters, 2013Co-Authors: Naoto Kojima, Masato Abe, Yuki Suga, Kazufumi Ohtsuki, Tetsuaki Tanaka, Hiroki Iwasaki, Masayuki Yamashita, Hideto MiyoshiAbstract:Abstract C34-epi and C34-epi-C35-trifluoro analogues of solamin, a mono-THF annonaceous Acetogenin, were synthesized. Their inhibitory activity, along with previously synthesized analogues (C35-fluoro, C35-difluoro, and C35-trifluorosolamins), against bovine mitochondrial NADH–ubiquinone oxidoreductase (complex I) was determined. The present study revealed that the methyl group on the γ-lactone moiety is critical to the potent inhibition of complex I by natural Acetogenins.
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bis thf motif of Acetogenin binds to the third matrix side loop of nd1 subunit in mitochondrial nadh ubiquinone oxidoreductase
Biochimica et Biophysica Acta, 2011Co-Authors: Sayo Nakanishi, Masato Abe, Shuhei Yamamoto, Masatoshi Murai, Hideto MiyoshiAbstract:Abstract Natural Acetogenins are among the most potent inhibitors of bovine heart mitochondrial NADH-ubiquinone oxidoreductase (complex I). Our photoaffinity labeling study suggested that the hydroxylated bis-THF ring moiety of Acetogenins binds at “site A” in the third matrix-side loop connecting the fifth and sixth transmembrane helices in the ND1 subunit [Kakutani et al. (2010) Biochemistry 49 , 4794–4803]. Nevertheless, since this proposition was led using a photoreactive Δlac-Acetogenin derivative, it needs to be directly verified using a natural Acetogenin-type probe. We therefore conducted photoaffinity labeling using a photoreactive natural Acetogenin mimic ([ 125 I] d i a zinylated n atural a cetogenin, [ 125 I]DANA), which has a small photolabile diazirine group, in place of a hydroxy group, attached to the bis-THF ring moiety. Analysis of the photocross-linked protein in bovine heart submitochondrial particles unambiguously revealed that [ 125 I]DANA binds to the membrane subunit ND1 with high specificity. The photocross-linking was completely blocked in the presence of just a 5-fold excess of bullatacin, indicating that [ 125 I]DANA is an excellent mimic of natural Acetogenins and hence binds to the site that accommodates natural products. Careful examination of the fragmentation patterns of the cross-linked ND1 generated by different proteases and their combinations indicated that the cross-linked residue is predominantly located at the supposed site A in the third matrix-side loop.
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synthesis and characterization of photoaffinity probe of Acetogenin a strong inhibitor of mitochondrial complex i
Tetrahedron Letters, 2011Co-Authors: Shuhei Yamamoto, Masato Abe, Sayo Nakanishi, Masatoshi Murai, Hideto MiyoshiAbstract:Abstract Acetogenins are valuable inhibitor probes to get an insight into the structural and functional properties of mitochondrial NADH-ubiquinone oxidoreductase (complex I). We synthesized a photoreactive Acetogenin mimic ([125I]DANA) which retained a strong inhibitory activity. The preliminary photoaffinity labeling with bovine heart submitochondrial particles revealed that [125I]DANA binds to the ND1 subunit among 45 different subunits with high specificity.
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Synthesis of photolabile Δlac-Acetogenin for photoaffinity labeling of mitochondrial complex I
Journal of Pesticide Science, 2006Co-Authors: Masatoshi Murai, Masato Abe, Takaaki Nishioka, Naoya Ichimaru, Hideto MiyoshiAbstract:Δlac-Acetogenins are a novel type of inhibitor acting at the terminal electron transfer step of mitochondrial NADH-ubiquinone oxidoreductase (complex I). To identify the binding site of Δlac-Acetogenins by photoaffinity labeling, we synthesized a photolabile Δlac-Acetogenin that possesses a biotin probe to enable the detection and the isolation of the labeled peptide without the use of a radioisotope. The photolabile Δlac-Acetogenin synthesized in this study elicited potent inhibition of bovine heart mitochondrial complex I at the nanomolar level. © Pesticide Science Society of Japan
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Synthesis and Inhibition Mechanism of Δlac-Acetogenins, a Novel Type of Inhibitor of Bovine Heart Mitochondrial Complex I†
Biochemistry, 2005Co-Authors: Naoya Ichimaru, Masato Abe, Takeshi Hamada, Takaaki Nishioka, Masatoshi Murai, Hidefumi Makabe, Yohsuke Yamada, Sae Makino, Asami Makino, Toshihide KobayashiAbstract:We have synthesized Deltalac-Acetogenins that are new Acetogenin mimics possessing two n-alkyl tails without an alpha,beta-unsaturated gamma-lactone ring and suggested that their inhibition mechanism may be different from that of common Acetogenins [Hamada et al. (2004) Biochemistry 43, 3651-3658]. To elucidate the inhibition mechanism of Deltalac-Acetogenins in more detail, we carried out wide structural modifications of original Deltalac-Acetogenins and characterized the inhibitory action with bovine heart mitochondrial complex I. In contrast to common Acetogenins, both the presence of adjacent bis-THF rings and the stereochemistry around the hydroxylated bis-THF rings are important structural factors required for potent inhibition. The inhibitory potency of a derivative possessing an n-butylphenyl ether structure (compound 7) appeared to be superior to that of the original Deltalac-Acetogenins and equivalent to that of bullatacin, one of the most potent natural Acetogenins. Double-inhibitor titration of steady-state complex I activity showed that the extent of inhibition of compound 7 and bullatacin is not additive, suggesting that the binding sites of the two inhibitors are not identical. Competition tests using a fluorescent ligand indicated that the binding site of compound 7 does not overlap with that of other complex I inhibitors. The effects of compound 7 on superoxide production from complex I are also different from those of other complex I inhibitors. Our results clearly demonstrate that Deltalac-Acetogenins are a novel type of inhibitor acting at the terminal electron-transfer step of bovine complex I.