D J Morré

Purdue University, West Lafayette, Indiana, United States

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Publications (522)1856.73 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: ENOX (ECTO-NOX) proteins are proteins of the external surface of the plasma membrane that catalyze oxidation of both NADH and hydroquinones as well as carry out protein disulfide-thiol interchange. They exhibit both prion-like and time-keeping (clock) properties. The oxidative and interchange activities alternate to generate a regular period of 24 min in length. Here we report the cloning, expression, and characterization of a plant candidate constitutive ENOX (CNOX or ENOX1) protein from Arabidopsis lyrata. The gene encoding the 335 (165) amino acid protein is found in accession XP-002882467. Functional motifs characteristics of ENOX proteins previously identified by site-directed mutagenesis and present in the candidate ENOX1 protein from plants include adenine nucleotide and copper binding motifs along with essential cysteines. However, the drug binding motif (EEMTE) sequence of human ENOX2 is absent. The activities of the recombinant protein expressed in E. coli were unaffected by capsaicin, EGCg, and other ENOX2-inhibiting substances. Periodic oxidative activity was exhibited both with NAD(P)H and reduced coenzyme Q as substrate. Bound copper was necessary for activity and activity was inhibited by the ENOX1- specific inhibitor simalikalactone D. Addition of melatonin phased the 24-min period such that the next complete period began 24 min after the melatonin addition as appeared to be characteristic of ENOX1 activities in general. Periodic protein disulfide-thiol interchange activity also was demonstrated along with the 2 oxidative plus 3 interchange activity pattern characteristics of the 24- min ENOX1 protein period. Concentrated solutions of the purified plant ENOX1 protein formed insoluble aggregates, devoid of enzymatic activity, resembling amyloid. Activity was restored to aggregate preparations by isoelectric focusing.
    Advances in Biological Chemistry 02/2015; 5:1-15.
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    ABSTRACT: ENOX (ECTO-NOX) proteins of the external surface of the plasma membrane catalyze oxidation of both NADH and hydroquinones and protein disulfide-thiol interchange. They exhibit both prionlike and time-keeping (clock) properties. The oxidative and interchange activities alternate to generate a regular period of 24 min in length. Here we report the cloning, expression and characterization of a constitutive plant ENOX protein activated by both natural (Indole-3-acetic acid, IAA) and synthetic (2,4-dichlorophenoxyacetic acid, 2,4-D) auxin plant growth regulators with an optimum of about 1 μM, higher concentrations being less effective. The gene encoding the 213 amino acid protein (ABP20) is found in EMBL accession number U81162. Functional motifs characteristic of ENOX1 proteins, previously identified by site-directed mutagenesis, are present in the candidate auxin-activated ENOX (dNOX, ENOX5), including adenine nucleotide and copper binding motifs along with essential cysteines and a motif having homology with a previously identified auxinbinding motif. Periodicity was exhibited by both the oxidative and protein disulfide-thiol interchange activities as is characteristic for other ENOX proteins. Activity was blocked by the ENOX2- specific quassinoid inhibitor glaucarubolone and other ENOX2 inhibitors but not by the ENOX1- specific quassinoid inhibitor simalikalactone D. Activity required both auxin and bound copper. The inactive auxin 2,3-D was without effects.
    Advances in Biological Chemistry 12/2014; 4(07):415-427. DOI:10.4236/abc.2014.47047
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    ABSTRACT: ME-143 (NV-143), a synthetic isoflavone under clinical evaluation for efficacy in the management of ovarian and other forms of human cancer, blocked the activity of a cancer-specific and growth-related cell surface ECTO-NOX protein with both oxidative (hydroquinone) and protein disulfide-thiol interchange activity designated ENOX2 (tNOX) and inhibited the growth of cultured cancer cells with EC50s in the range of 20‐50 nM. Purified recombinant ENOX2 also bound ME-143 with a K d of 43 (40‐50) nM. Both the oxidative and protein disulfide-thiol interchange activities of ENOX proteins that alternate to generate a complex set of oscillations with a period length of 22 min compared to 24 min for the constitutive counterpart ENOX1 (CNOX) that characterizes ENOX proteins responded to ME-143. Oxidation of NADH or reduced coenzyme Q10 was rapidly blocked. In contrast, the protein disulfide-thiol interchange activity measured from the cleavage of dithiodipyridine (EC50 of ca. 50 nM) was inhibited progressively over an interval of 60 min that spanned three cycles of activity. Inhibition of the latter paralleled the inhibition of cell enlargement and the consequent inability of inhibited cells to initiate traverse of the cell cycle. Activities of constitutive ENOX1 (CNOX) forms of either cancer or noncancer cells were unaffected by ME-143 over the range of concentrations inhibiting ENOX2. Taken together, the findings show that ME-143 binds to ENOX2 with an affinity 4 to 10 times greater than that reported previously for the related anticancer isoflavone, phenoxodiol.
    Oncology Research Featuring Preclinical and Clinical Cancer Therapeutics 10/2014; 22(1). DOI:10.3727/096504014X14077751730270 · 0.92 Impact Factor
  • X Tang, DM Morré, DJ Morré
    08/2014; DOI:10.4267/2042/54027
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    ABSTRACT: Age-related NADH oxidase (arNOX), a cell surface-located hydroquinone oxidase capable of superoxide generation, appears at age 30 and increases with age thereafter. The ectodomain of arNOX is shed from the cell surface into body fluids including sera and saliva where its activity was measured spectrophotometrically using a reduction of ferricytochrome c as a measure of superoxide generation. The autofluorescence of advanced glycation end products correlates with epidermal arNOX activity as well. To demonstrate protein cross-linking, a fluorescence-labeled analog of tyrosine, tyramine, that would react with proteins carrying arNOX-generated tyrosyl radicals was used. The assay demonstrated the potential for arNOX-induced oxidative damage (dityrosine formation) to human collagen and elastin and to other surface proteins of intact human embryo fibroblasts and frozen sections from epidermal punch biopsies. The findings support a role for arNOX as a major source of oxidative damage leading to cross-linking of skin proteins.
    Archives for Dermatological Research 06/2014; DOI:10.1007/s00403-014-1472-8 · 2.27 Impact Factor
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    Claudia Hanau, D James Morré, Dorothy M Morré
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    ABSTRACT: Experts agree that one of the more promising strategies in cancer management is early detection coupled with early intervention. In this study, we evaluated an early cancer detection strategy of cancer presence based on serum levels of the cancer-specific transcript variants of ENOX2 in serum coupled with an ENOX2-targeted nutraceutical preparation of green tea concentrate plus Capsicum (Capsol-T(R)) as a strategy of Curative Prevention(R) involving early detection coupled with early intervention in early stage cancer when in its most susceptible and manageable stages.Experimental design: One hundred ten (110) subjects were tested for cancer presence using the ONCOblot(R) Tissue of Origin 2-D gel/western blot protocol for detection of serum presence of transcription variants of the ENOX2 protein. Subjects testing positive for ENOX2 received 350 mg of Capsol-T(R) in capsule form every 4 h including during the night for periods of at least 3 to 6 months or longer after which they were again tested for ENOX2 presence using the ONCOblot(R) Tissue of Origin Cancer Test protocol. Of the 110 subjects, both male and female, ages 40 to 84, with no evidence of clinical symptoms of cancer, 40% were positive for ENOX2 presence in the ONCOblot(R) Tissue of Origin Cancer Test. After completion of 3 to 17 months of Capsol-T(R) use, 94% of subjects subsequently tested negative for ENOX2 presence. Oral Capsol-T(R) is well tolerated and, for ENOX2 presence in serum in the absence of clinical cancer symptoms, is consistently effective in reducing the serum ENOX2 levels to below detectable limits.
    Clinical Proteomics 01/2014; 11(1):2. DOI:10.1186/1559-0275-11-2
  • Advances in Biological Chemistry 01/2014; 04(05):339-350. DOI:10.4236/abc.2014.45039
  • World Journal of Cardiovascular Diseases 01/2014; 04(03):119-129. DOI:10.4236/wjcd.2014.43018 · 0.22 Impact Factor
  • Greg B Forney, D James Morré, Dorothy M Morré
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    ABSTRACT: ABSTRACT Reactive oxygen species that are produced by aerobic metabolism and signaling cascades have the potential to play important roles in maintaining homeostatic redox and cell proliferation. When the balance between the production and elimination of reactive oxygen species is perturbed toward production, the result is oxidative stress. High levels of oxidative stress are a general characteristic of cancer. The altered redox state within a tumor microenvironment confers a growth advantage through increased proliferation rates, evasion of apoptosis, and increased resistance to therapeutic compounds. We have tested a synergistic combination of green tea-Camellia sinensis-concentrate and powdered Capsicum powder (TeaFense™/Capsol-T™) as a dietary supplement to reduce oxidative stress as an approach to elimination of malignant cells. Here, we demonstrate that the green tea-powdered Capsicum mixture effectively reduces levels of oxidative stress in both cancer (HeLa) and noncancer (MCF-10A) cells as determined from measurements of levels of the oxidative stress indicator Nrf-2 by western blot analysis. Nrf-2 is a transcription factor that controls an antioxidant response element. Increased expression of Nrf-2 is linked to high levels of oxidative stress and vice versa. Based on levels of Nrf-2, the mixture of green tea concentrate plus powdered Capsicum reduced oxidative stress by more than 50% compared with 15% by the green tea concentrate alone.
    Journal of Dietary Supplements 12/2013; 10(4):318-24. DOI:10.3109/19390211.2013.830673
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    ABSTRACT: arNOX, an age-related NADH oxidase, has been shown to play a role in the formation of the autophagosome. The role of autophagy in tumorigenesis and tumor proliferation led to the hypothesis that abnormal arNOX levels might play a role in the pathology of cancer. Serum samples from cancer patients and age- and gender-matched healthy controls were assayed to measure arNOX activity by a standard assay involving measurements of superoxide production based on the reduction of ferricyanide c by superoxide monitored from the increase in absorbance at 550 nm with reference at 540 nm. Superoxide dismutase was added to the end of the assay to ensure return of the rate of ferricytochrome c reduction to the base line. Cancer and cardiovascular diseases are inversely correlated. Similarly, arNOX levels were decreased in cancer patients when compared to non-cancer control subjects while high arNOX levels have been implicated as an important cause of LDL oxidation and increased risk of cardiovascular disease. These findings may help further the understanding of the pathologies of cancer and cardiovascular disease and have implications for risk assessment for both diseases as well as for future therapeutic intervention strategies.
    Journal of medicine and life 07/2013; 1(2):38. DOI:10.14511/jlm.2013.010204
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    ABSTRACT: Age-related NADH oxidase (arNOX) is a cell surface protein shed into the circulation and other body fluids, which generates superoxide. The activity increases with age in human tissues and body fluids (serum, saliva, and perspiration) and is a potential source of age-related oxidative damage. We measured arNOX activity in the coelomic fluid of sea urchin species with different life spans. Coelomic fluid of long-lived sea urchin species Strongylocentrotus purpuratus and Strongylocentrotus franciscanus exhibited low levels of arNOX compared to the short-lived urchin species Lytechinus variegatus. arNOX activity was positively correlated with animal size in L. variegatus, whereas with S. purpuratus and S. franciscanus, arNOX activity and animal size were inversely correlated. The inverse correlation of arNOX activity with life span and decreased levels of arNOX with age in the long-lived species is consistent with a contribution of reduced arNOX activity to slower aging.
    01/2013; 5(1). DOI:10.1186/2008-6970-5-2
  • Advances in Biological Chemistry 01/2013; 03(03):320-328. DOI:10.4236/abc.2013.33036
  • Advances in Biological Chemistry 01/2013; 03(05):505-511. DOI:10.4236/abc.2013.35055
  • Advances in Biological Chemistry 01/2013; 03(02):187-197. DOI:10.4236/abc.2013.32024
  • Source
    Hans Löw, Frederick L Crane, D James Morré
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    ABSTRACT: The observation that high cellular concentrations of NADH were associated with low adenylate cyclase activity led to a search for the mechanism of the effect. Since cyclase is in the plasma membrane, we considered the membrane might have a site for NADH action, and that NADH might be oxidized at that site. A test for NADH oxidase showed very low activity, which could be increased by adding growth factors. The plasma membrane oxidase was not inhibited by inhibitors of mitochondrial NADH oxidase such as cyanide, rotenone or antimycin. Stimulation of the plasma membrane oxidase by iso-proterenol or triiodothyronine was different from lack of stimulation in endoplasmic reticulum. After 25 years of research, three components of a trans membrane NADH oxidase have been discovered. Flavoprotein NADH coenzyme Q reductases (NADH cytochrome b reductase) on the inside, coenzyme Q in the middle, and a coenzyme Q oxidase on the outside as a terminal oxidase. The external oxidase segment is a copper protein with unique properties in timekeeping, protein disulfide isomerase and endogenous NADH oxidase activity, which affords a mechanism for control of cell growth by the overall NADH oxidase and the remarkable inhibition of oxidase activity and growth of cancer cells by a wide range of anti-tumor drugs. A second trans plasma membrane electron transport system has been found in voltage dependent anion channel (VDAC), which has NADH ferricyanide reductase activity. This activity must be considered in relation to ferricyanide stimulation of growth and increased VDAC antibodies in patients with autism.
    The international journal of biochemistry & cell biology 06/2012; 44(11):1834-8. DOI:10.1016/j.biocel.2012.06.032 · 4.24 Impact Factor
  • D. J. Morré, D. M. Morré
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    ABSTRACT: A homodimeric, growth-related and time-keeping hydroquinone oxidase (ENOX1) of the eukaryotic cell surface capable of oxidizing extracellular NADH exhibits properties of the ultradian driver of the biological 24 h circadian clock by exhibiting a complex 2 + 3 set of oscillations with a period length of 24 min. The oscillations require bound copper, are recapitulated by aqueous solutions of copper salts and appear to derive from 30 to 40 sec periodic variations in the ratios of ortho and para nuclear spins of the paired hydrogen atoms of the elongated octahedral structure of the ENOX2 protein bound copperII hexahydrates. Based on a heartbeat model of limit cycle oscillations, these oscillations correlate with generation of the 24 min periodicity as a form of carrier wave. A corollary of these observations is that the ortho-para oscillations must occur in a highly synchronized manner. Attendant oscillatory changes in redox potential offer an opportunity to monitor oscillations in synchronous populations of water molecules. The periodicity of the ENOX1/copper/water clock can be phased by brief 10 to 20 sec exposure to very low frequency electromagnetic fields. The synchronized populations of oscillating water molecules give rise to oscillatory electromagnetic fields that apparently are perceived by adjacent water molecules to create a collectively coherent synchronous system. Two asynchronous water samples placed adjacent to one another but separated by a thin non-metal barrier become fully synchronized in a matter of several min. A barrier of metal foil prevents the synchronization. The corollary of these observations is that contiguous water molecules function synchronously perhaps even over relatively long distances. Two samples of water from contiguous still or flowing bodies of water collected from different locations and analyzed simultaneously were synchronous in their oscillations in redox potential measured as changes in rates of NADH oxidation. Our findings suggest that water molecules communicate with each other via very low frequency electromagnetic fields.
    WATER AND SOCIETY 2011; 12/2011
  • D James Morré, Dorothy M Morré
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    ABSTRACT: The key role of coenzyme Q (ubiquinone or Q) is in mitochondrial and prokaryotic energetics. Less well investigated is the basis for its presence in eukaryotic membrane locations other than mitochondria and in plasma where both antioxidant and potentially more targeted roles are indicated. Included in the latter is that of a lipid-soluble electron transfer intermediate that serves as the transmembrane component of plasma membrane and Golgi apparatus electron transport, which regulates cytosolic NAD(+) /NADH ratios and is involved in vectorial membrane displacements and in the regulation of cell growth. Important protective effects on circulating lipoproteins and in the prevention of coronary artery disease ensue not only from the antioxidant role of CoQ(10) but also from its ability to directly block protein oxidation and superoxide generation of the TM-9 family of membrane proteins known as age-related NADH oxidase or arNOX (ENOX3) and their shed forms that appear after age 30 and some of which associate specifically with low-density lipoprotein particles to catalyze protein oxidation and crosslinking.
    BioFactors 09/2011; 37(5):355-60. DOI:10.1002/biof.156 · 3.00 Impact Factor
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    ABSTRACT: HUVEC or mouse 3T3 cells infected with SV-40 generate within 3 to 5 days post-infection an ENOX2 species corresponding to the exon-4 minus splice variant of a tumor-associated NADH oxidase (ENOX2 or tNOX) expressed at the cancer cell surface. This study was to seek evidence for splicing factors that might direct formation of the exon 4 minus ENOX2 splice variant. To determine if silencing of ENOX2 exon 4 occurs because of motifs located in exon 4, transfections were performed on MCF-10A (mammary non-cancer), BT-20 (mammary cancer), and HeLa (cervical cancer) cells using a GFP minigene construct containing either a constitutively spliced exon (albumin exon 2) or the alternatively spliced ENOX2 exon 4 between the two GFP halves. Removal of exon 4 from the processed RNA of the GFP minigene construct occurred with HeLa and to a lesser extent with BT-20 but not in non-cancer MCF-10A cells. The Splicing Rainbow Program was used to identify all of the possible hnRNPs binding sites of exon 4 of ENOX2. There are 8 Exonic Splicing Silencers (ESSs) for hnRNP binding in the exon 4 sequences. Each of these sites were mutated by site-directed mutagenesis to test if any were responsible for the splicing skip. Results showed MutG75 ESS mutation changed the GFP expression which is a sign of splicing silence, while other mutations did not. As MutG75 changed the ESS binding site for hnRNP F, this result suggests that hnRNP F directs formation of the exon 4 minus variant of ENOX2.
    Molecular and Cellular Biochemistry 05/2011; 357(1-2):55-63. DOI:10.1007/s11010-011-0875-5 · 2.39 Impact Factor
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    ABSTRACT: Constituents and inhibitors of intermediary metabolism resulting in alterations in levels of cytosolic NADH, stimulation of sphingomyelinase and inhibition of sphingosine kinase were evaluated for effects on growth inhibition and induction of apoptosis by the ENOX2 inhibitors EGCG, the principal catechin of green tea, and phenoxodiol, a naturally occurring isoflavone. Responses were evaluated from dose-response curves of the metabolites and metabolic inhibitors in which growth of HeLa cells, apoptosis based on DAPI fluorescence and cytosolic NADH levels were correlated with sphingomyelinase and spingosine kinase activities and levels of ceramide and sphingosine1-phosphate. Growth inhibition correlated with the modulation of localized cytosolic NADH levels by metabolites and metabolic inhibitors, the response of sphingomyelinase and sphingosine kinase located near the inner surface of the plasma membrane, and apoptosis. Based on findings with metabolites, we conclude that apoptosis in cancer cell lines caused by ENOX2 inhibitors such as EGCG and phenoxodiol is a direct response to elevated levels of cytosolic NADH that result from ENOX2 inhibition. The findings help to explain why increased NADH levels resulting from ENOX2 inhibition result in decreased prosurvival sphingosine-1-phosphate and increased proapoptotic ceramide, both of which may be important to initiation of the ENOX2 inhibitor-induced apoptotic cascade.
    Biochimica et Biophysica Acta 05/2011; 1810(8):784-9. DOI:10.1016/j.bbagen.2011.04.011 · 4.66 Impact Factor
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    ABSTRACT: ECTO-NOX proteins are growth-related cell surface proteins that catalyze both hydroquinone or NADH oxidation and protein disulfide interchange and exhibit time-keeping and prion-like properties. A bacterially expressed truncated recombinant 46 kDa ENOX2 with full ENOX2 activity bound ca 2 moles copper and 2 moles of zinc per mole of protein. Unfolding of the protein in trifluoroacetic acid in the presence of the copper chelator bathocuproine resulted in reversible loss of both enzymatic activities and of a characteristic pattern in the Amide I to Amide II ratios determined by FTIR with restoration by added copper. The H546-V-H together with His 562 form one copper binding site and H582 represents a second copper site as determined from site-directed mutagenesis. Bound copper emerges as having an essential role in ENOX2 both for enzymatic activity and for the structural changes that underly the periodic alternations in activity that define the time-keeping cycle of the protein.
    Journal of Bioenergetics 10/2010; 42(5):355-60. DOI:10.1007/s10863-010-9305-8 · 2.71 Impact Factor

Publication Stats

10k Citations
1,856.73 Total Impact Points


  • 1965–2013
    • Purdue University
      • • Department of Medicinal Chemistry and Molecular Pharmacology (MCMP)
      • • Department of Biological Sciences
      • • Department of Animal Sciences
      • • Department of Botany and Plant Pathology
      West Lafayette, Indiana, United States
  • 2006
    • University of Alabama at Birmingham
      • Department of Biology
      Birmingham, AL, United States
  • 1980–1998
    • University of Geneva
      • Department of Botany and Plant Biology
      Genève, Geneva, Switzerland
  • 1987–1995
    • University of Cordoba (Spain)
      • Departamento de Biología Celular, Fisiología e Inmunología
      Cordoue, Andalusia, Spain
    • Virginia Polytechnic Institute and State University
      Blacksburg, Virginia, United States
  • 1993
    • Hungarian Academy of Sciences
      • Institute of Biophysics
      Budapeŝto, Budapest, Hungary
  • 1992
    • University of Gothenburg
      Goeteborg, Västra Götaland, Sweden
  • 1988
    • Johannes Gutenberg-Universität Mainz
      • Institute of General Botany
      Mayence, Rheinland-Pfalz, Germany
  • 1986
    • Universität Heidelberg
      Heidelburg, Baden-Württemberg, Germany
    • University of Hamburg
      Hamburg, Hamburg, Germany
  • 1973–1974
    • Agricultural Research Service
      ERV, Texas, United States
  • 1970
    • Pennsylvania State University
      University Park, Maryland, United States