AOC3 is highly expressed in adipocytes and smooth muscle cells, but its function in these cells is currently unknown. The in vivo substrate(s) of AOC3 is/are also unknown, but could provide an invaluable clue to the enzyme's function. Expression of untagged, soluble human AOC3 in insect cells provides a relatively simple means of obtaining pure enzyme. Characterization of enzyme indicates a 6% titer for the active site 2,4,5-trihydroxyphenylalanine quinone (TPQ) cofactor and corrected kcat values as high as 7 s？1. Substrate kinetic profiling shows that the enzyme accepts a variety of primary amines with different chemical features, including nonphysiological branched-chain and aliphatic amines, with measured kcat/Km values between 102 and 104 M？1 s？1. Km(O2) approximates the partial pressure of oxygen found in the interstitial space. Comparison of the properties of purified murine to human enzyme indicates kcat/Km values that are within 3 to 4-fold, with the exception of methylamine and aminoacetone that are ca. 10-fold more active with human AOC3. With drug development efforts investigating AOC3 as an anti-inflammatory target, these studies suggest that caution is called for when screening the efficacy of inhibitors designed against human enzymes in non-transgenic mouse models. Differentiated murine 3T3-L1 adipocytes show a uniform distribution of AOC3 on the cell surface and whole cell Km values that are reasonably close to values measured using purified enzymes. The latter studies support a relevance of the kinetic parameters measured with isolated AOC3 variants to adipocyte function. From our studies, a number of possible substrates with relatively high kcat/Km have been discovered, including dopamine and cysteamine, which may implicate a role for adipocyte AOC3 in insulin-signaling and fatty acid metabolism, respectively. Finally, the demonstrated AOC3 turnover of primary amines that are non-native to human tissue suggests possible roles for the adipocyte enzyme in subcutaneous bacterial infiltration and obesity.
References
[1]
Janes SM, Mu D, Wemmer D, Smith AJ, Kaur S, et al. (1990) A new redox cofactor in eukaryotic enzymes: 6-hydroxydopa at the active site of bovine serum amine oxidase. Science 248: 981–987.
[2]
O'Sullivan J, Unzeta M, Healy J, O'Sullivan MI, Davey G, et al. (2004) Semicarbazide-sensitive amine oxidases: enzymes with quite a lot to do. Neurotoxicology 25: 303–315.
[3]
Mure M, Mills SA, Klinman JP (2002) Catalytic mechanism of the topa quinone containing copper amine oxidases. Biochemistry 41: 9269–9278.
[4]
Boomsma F, de Kam PJ, Tjeerdsma G, van den Meiracker AH, van Veldhuisen DJ (2000) Plasma semicarbazide-sensitive amine oxidase (SSAO) is an independent prognostic marker for mortality in chronic heart failure. Eur Heart J 21: 1859–1863.
[5]
Boomsma F, van den Meiracker AH, Winkel S, Aanstoot HJ, Batstra MR, et al. (1999) Circulating semicarbazide-sensitive amine oxidase is raised both in type I (insulin-dependent), in type II (non-insulin-dependent) diabetes mellitus and even in childhood type I diabetes at first clinical diagnosis. Diabetologia 42: 233–237.
[6]
Ferrer I, Lizcano JM, Hernandez M, Unzeta M (2002) Overexpression of semicarbazide sensitive amine oxidase in the cerebral blood vessels in patients with Alzheimer's disease and cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy. Neurosci Lett 321: 21–24.
[7]
Kurkijarvi R, Adams DH, Leino R, Mottonen T, Jalkanen S, et al. (1998) Circulating form of human vascular adhesion protein-1 (VAP-1): increased serum levels in inflammatory liver diseases. J Immunol 161: 1549–1557.
[8]
Marttila-Ichihara F, Auvinen K, Elima K, Jalkanen S, Salmi M (2009) Vascular adhesion protein-1 enhances tumor growth by supporting recruitment of Gr-1+CD11b+ myeloid cells into tumors. Cancer Res 69: 7875–7883.
[9]
Rea G, Laurenzi M, Tranquilli E, D'Ovidio R, Federico R, et al. (1998) Developmentally and wound-regulated expression of the gene encoding a cell wall copper amine oxidase in chickpea seedlings. FEBS Lett 437: 177–182.
[10]
Andres N, Lizcano JM, Rodriguez MJ, Romera M, Unzeta M, et al. (2001) Tissue activity and cellular localization of human semicarbazide-sensitive amine oxidase. J Histochem Cytochem 49: 209–217.
[11]
Morris NJ, Ducret A, Aebersold R, Ross SA, Keller SR, et al. (1997) Membrane amine oxidase cloning and identification as a major protein in the adipocyte plasma membrane. J Biol Chem 272: 9388–9392.
[12]
Salmi M, Jalkanen S (2005) Cell-surface enzymes in control of leukocyte trafficking. Nat Rev Immunol 5: 760–771.
[13]
Salmi M, Jalkanen S (1992) A 90-kilodalton endothelial cell molecule mediating lymphocyte binding in humans. Science 257: 1407–1409.
[14]
Barreiro O, Sanchez-Madrid F (2009) Molecular basis of leukocyte-endothelium interactions during the inflammatory response. Rev Esp Cardiol 62: 552–562.
[15]
Koskinen K, Vainio PJ, Smith DJ, Pihlavisto M, Yla-Herttuala S, et al. (2004) Granulocyte transmigration through the endothelium is regulated by the oxidase activity of vascular adhesion protein-1 (VAP-1). Blood 103: 3388–3395.
[16]
Kivi E, Elima K, Aalto K, Nymalm Y, Auvinen K, et al. (2009) Human Siglec-10 can bind to vascular adhesion protein-1 and serves as its substrate. Blood 114: 5385–5392.
[17]
Trayhurn P, Wood IS (2004) Adipokines: inflammation and the pleiotropic role of white adipose tissue. Br J Nutr 92: 347–355.
[18]
Fruhbeck G, Gomez-Ambrosi J, Muruzabal FJ, Burrell MA (2001) The adipocyte: a model for integration of endocrine and metabolic signaling in energy metabolism regulation. Am J Physiol Endocrinol Metab 280: E827–847.
Enrique-Tarancon G, Castan I, Morin N, Marti L, Abella A, et al. (2000) Substrates of semicarbazide-sensitive amine oxidase co-operate with vanadate to stimulate tyrosine phosphorylation of insulin-receptor-substrate proteins, phosphoinositide 3-kinase activity and GLUT4 translocation in adipose cells. Biochem J 350 Pt 1: 171–180.
[21]
Zorzano A, Abella A, Marti L, Carpene C, Palacin M, et al. (2003) Semicarbazide-sensitive amine oxidase activity exerts insulin-like effects on glucose metabolism and insulin-signaling pathways in adipose cells. Biochim Biophys Acta 1647: 3–9.
[22]
Forman HJ, Maiorino M, Ursini F (2010) Signaling functions of reactive oxygen species. Biochemistry 49: 835–842.
[23]
Rhee SG (2006) Cell signaling. H2O2, a necessary evil for cell signaling. Science 312: 1882–1883.
[24]
Mahadev K, Wu X, Zilbering A, Zhu L, Lawrence JT, et al. (2001) Hydrogen peroxide generated during cellular insulin stimulation is integral to activation of the distal insulin signaling cascade in 3T3-L1 adipocytes. J Biol Chem 276: 48662–48669.
[25]
Carpene C, Iffiu-Soltesz Z, Bour S, Prevot D, Valet P (2007) Reduction of fat deposition by combined inhibition of monoamine oxidases and semicarbazide-sensitive amine oxidases in obese Zucker rats. Pharmacol Res 56: 522–530.
[26]
Wishart DS, Knox C, Guo AC, Eisner R, Young N, et al. (2009) HMDB: a knowledgebase for the human metabolome. Nucleic Acids Res 37: D603–610.
[27]
Neumann R, Hevey R, Abeles RH (1975) The action of plasma amine oxidase on beta-haloamines. Evidence for proton abstraction in the oxidative reaction. J Biol Chem 250: 6362–6367.
[28]
Stephens JM, Lee J, Pilch PF (1997) Tumor necrosis factor-alpha-induced insulin resistance in 3T3-L1 adipocytes is accompanied by a loss of insulin receptor substrate-1 and GLUT4 expression without a loss of insulin receptor-mediated signal transduction. J Biol Chem 272: 971–976.
[29]
Ucar G, Topaloglu E, Burak Kandilci H, Gumusel B (2005) Elevated semicarbazide-sensitive amine oxidase (SSAO) activity in lung with ischemia-reperfusion injury: protective effect of ischemic preconditioning plus SSAO inhibition. Life Sci 78: 421–427.
[30]
Welford RW, Lam A, Mirica LM, Klinman JP (2007) Partial conversion of Hansenula polymorpha amine oxidase into a “plant” amine oxidase: implications for copper chemistry and mechanism. Biochemistry 46: 10817–10827.
[31]
Kao AW, Noda Y, Johnson JH, Pessin JE, Saltiel AR (1999) Aldolase mediates the association of F-actin with the insulin-responsive glucose transporter GLUT4. J Biol Chem 274: 17742–17747.
[32]
Merinen M, Irjala H, Salmi M, Jaakkola I, Hanninen A, et al. (2005) Vascular adhesion protein-1 is involved in both acute and chronic inflammation in the mouse. Am J Pathol 166: 793–800.
[33]
Singer TP, Ramsay RR (1995) Flauoprotein structure and mechanism 2. Monoamine oxidases: old friends hold many surprises. FASEB J 9: 605–610.
[34]
Jakobsson E, Nilsson J, Kallstrom U, Ogg D, Kleywegt GJ (2005) Crystallization of a truncated soluble human semicarbazide-sensitive amine oxidase. Acta Crystallogr Sect F Struct Biol Cryst Commun 61: 274–278.
[35]
Ernberg K, McGrath AP, Peat TS, Adams TE, Xiao X, et al. (2010) A new crystal form of human vascular adhesion protein 1. Acta Crystallogr Sect F Struct Biol Cryst Commun 66: 1572–1578.
[36]
Airenne TT, Nymalm Y, Kidron H, Smith DJ, Pihlavisto M, et al. (2005) Crystal structure of the human vascular adhesion protein-1: unique structural features with functional implications. Protein Sci 14: 1964–1974.
Johansen H, van der Straten A, Sweet R, Otto E, Maroni G, et al. (1989) Regulated expression at high copy number allows production of a growth-inhibitory oncogene product in Drosophila Schneider cells. Genes Dev 3: 882–889.
[39]
Booth RE, Misquitta SA, Bateman RC Jr (2003) Human pituitary glutaminyl cyclase: expression in insect cells and dye affinity purification. Protein Expr Purif 32: 141–146.
[40]
Brazeau BJ, Johnson BJ, Wilmot CM (2004) Copper-containing amine oxidases. Biogenesis and catalysis; a structural perspective. Arch Biochem Biophys 428: 22–31.
[41]
Suzuki S, Sakurai T, Nakahara A, Manabe T, Okuyama T (1983) Effect of metal substitution on the chromophore of bovine serum amine oxidase. Biochemistry 22: 1630–1635.
[42]
Cai D, Williams NK, Klinman JP (1997) Effect of metal on 2,4,5-trihydroxyphenylalanine (topa) quinone biogenesis in the Hansenula polymorpha copper amine oxidase. J Biol Chem 272: 19277–19281.
[43]
DuBois JLKJ (2005) Mechanism of post-translational quinone formation in copper amine oxidases and its relationship to the catalytic turnover. Arch Biochem Biophys 433: 255–265.
[44]
Bonaiuto E, Lunelli M, Scarpa M, Vettor R, Milan G, et al. (2010) A structure-activity study to identify novel and efficient substrates of the human semicarbazide-sensitive amine oxidase/VAP-1 enzyme. Biochimie 92: 858–868.
[45]
Davis EJ, De Ropp RS (1961) Metabolic origin of urinary methylamine in the rat. Nature 190: 636–637.
[46]
Mitchell SC, Zhang AQ, Smith RL (2000) Ethylamine in human urine. Clin Chim Acta 302: 69–78.
[47]
Li H, Luo W, Lin J, Lin Z, Zhang Y (2004) Assay of plasma semicarbazide-sensitive amine oxidase and determination of its endogenous substrate methylamine by liquid chromatography. J Chromatogr B Analyt Technol Biomed Life Sci 810: 277–282.
[48]
Miura Y (2000) Plasma beta-phenylethylamine in Parkinson's disease. Kurume Med J 47: 267–272.
[49]
Lee TL, Hsu CT, Yen ST, Lai CW, Cheng JT (1998) Activation of beta3-adrenoceptors by exogenous dopamine to lower glucose uptake into rat adipocytes. J Auton Nerv Syst 74: 86–90.
[50]
Bird MI, Nunn PB, Lord LA (1984) Formation of glycine and aminoacetone from L-threonine by rat liver mitochondria. Biochim Biophys Acta 802: 229–236.
[51]
Dupre S, Graziani MT, Rosei MA, Fabi A, Del Grosso E (1970) The enzymatic breakdown of pantethine to pantothenic acid and cystamine. Eur J Biochem 16: 571–578.
Elovaara H, Kidron H, Parkash V, Nymalm Y, Bligt E, et al. (2011) Identification of two imidazole binding sites and key residues for substrate specificity in human primary amine oxidase AOC3. Biochemistry 50: 5507–5520.
[54]
Heuts DP, Gummadova JO, Pang J, Rigby SE, Scrutton NS (2011) Reaction of Vascular Adhesion Protein-1 (VAP-1) with Primary Amines: Mechanistic Insights from Isotope Effects and Quantitative Structure-Activity Relationships. J Biol Chem 286: 29584–29593.
[55]
Salmi M, Yegutkin GG, Lehvonen R, Koskinen K, Salminen T, et al. (2001) A cell surface amine oxidase directly controls lymphocyte migration. Immunity 14: 265–276.
[56]
Olivieri A, Tipton K, O'Sullivan J (2007) L-lysine as a recognition molecule for the VAP-1 function of SSAO. J Neural Transm 114: 747–749.
[57]
Yegutkin GG, Salminen T, Koskinen K, Kurtis C, McPherson MJ, et al. (2004) A peptide inhibitor of vascular adhesion protein-1 (VAP-1) blocks leukocyte-endothelium interactions under shear stress. Eur J Immunol 34: 2276–2285.
[58]
Kaitaniemi S, Elovaara H, Gron K, Kidron H, Liukkonen J, et al. (2009) The unique substrate specificity of human AOC2, a semicarbazide-sensitive amine oxidase. Cell Mol Life Sci 66: 2743–2757.
[59]
Marti L, Abella A, De La Cruz X, Garcia-Vicente S, Unzeta M, et al. (2004) Exploring the binding mode of semicarbazide-sensitive amine oxidase/VAP-1: identification of novel substrates with insulin-like activity. J Med Chem 47: 4865–4874.
[60]
Nurminen EM, Pihlavisto M, Lazar L, Szakonyi Z, Pentikainen U, et al. (2010) Synthesis, in vitro activity, and three-dimensional quantitative structure-activity relationship of novel hydrazine inhibitors of human vascular adhesion protein-1. J Med Chem 53: 6301–6315.
[61]
Iffiu-Soltesz Z, Mercader J, Daviaud D, Boucher J, Carpene C (2011) Increased primary amine oxidase expression and activity in white adipose tissue of obese and diabetic db？/？mice. J Neural Transm 118: 1071–1077.
[62]
Scherer PE, Bickel PE, Kotler M, Lodish HF (1998) Cloning of cell-specific secreted and surface proteins by subtractive antibody screening. Nat Biotechnol 16: 581–586.
[63]
Bickel PE (2002) Lipid rafts and insulin signaling. Am J Physiol Endocrinol Metab 282: E1–E10.
[64]
Guyton KZ, Liu Y, Gorospe M, Xu Q, Holbrook NJ (1996) Activation of mitogen-activated protein kinase by H2O2. Role in cell survival following oxidant injury. J Biol Chem 271: 4138–4142.
[65]
Jalkanen S, Salmi M (2001) Cell surface monoamine oxidases: enzymes in search of a function. EMBO J 20: 3893–3901.
[66]
Gustafsson C, Govindarajan S, Minshull J (2004) Codon bias and heterologous protein expression. Trends Biotechnol 22: 346–353.
[67]
Maula SM, Salminen T, Kaitaniemi S, Nymalm Y, Smith DJ, et al. (2005) Carbohydrates located on the top of the “cap” contribute to the adhesive and enzymatic functions of vascular adhesion protein-1. Eur J Immunol 35: 2718–2727.
[68]
Chang CM, Klema VJ, Johnson BJ, Mure M, Klinman JP, et al. (2010) Kinetic and structural analysis of substrate specificity in two copper amine oxidases from Hansenula polymorpha. Biochemistry 49: 2540–2550.
[69]
Xiao S, Yu PH (2009) A fluorometric high-performance liquid chromatography procedure for simultaneous determination of methylamine and aminoacetone in blood and tissues. Anal Biochem 384: 20–26.
[70]
Schayer RW, Smiley RL, Kaplan EH (1952) The metabolism of epinephrine containing isotopic carbon. II. J Biol Chem 198: 545–551.
[71]
Tressel T, Thompson R, Zieske LR, Menendez MI, Davis L (1986) Interaction between L-threonine dehydrogenase and aminoacetone synthetase and mechanism of aminoacetone production. J Biol Chem 261: 16428–16437.
[72]
Edgar AJ (2002) The human L-threonine 3-dehydrogenase gene is an expressed pseudogene. BMC Genet 3: 18.
[73]
Bird MI, Nunn PB (1983) Metabolic homoeostasis of L-threonine in the normally-fed rat. Importance of liver threonine dehydrogenase activity. Biochem J 214: 687–694.
[74]
Ballevre O, Cadenhead A, Calder AG, Rees WD, Lobley GE, et al. (1990) Quantitative partition of threonine oxidation in pigs: effect of dietary threonine. Am J Physiol 259: E483–491.
[75]
Boylan SA, Dekker EE (1981) L-threonine dehydrogenase. Purification and properties of the homogeneous enzyme from Escherichia coli K-12. J Biol Chem 256: 1809–1815.
[76]
Bamshad M, Aoki VT, Adkison MG, Warren WS, Bartness TJ (1998) Central nervous system origins of the sympathetic nervous system outflow to white adipose tissue. Am J Physiol 275: R291–299.
[77]
Bukowski JF, Morita CT, Brenner MB (1999) Human gamma delta T cells recognize alkylamines derived from microbes, edible plants, and tea: implications for innate immunity. Immunity 11: 57–65.
[78]
Ghenghesh KS, Drucker DB (1989) Gas liquid chromatography of amines produced by the Enterobacteriaceae. Braz J Med Biol Res 22: 653–665.
[79]
Allison C, Macfarlane GT (1989) Influence of pH, nutrient availability, and growth rate on amine production by Bacteroides fragilis and Clostridium perfringens. Appl Environ Microbiol 55: 2894–2898.
[80]
Yuan B, Wang X, Wang Z, Wei J, Qing C, et al. (2010) Comparison of fibrogenesis caused by dermal and adipose tissue injury in an experimental model. Wound Repair Regen 18: 202–210.
[81]
Bouwman JJ, Visseren FL, Bouter KP, Diepersloot RJ (2008) Infection-induced inflammatory response of adipocytes in vitro. Int J Obes (Lond) 32: 892–901.
[82]
Caesar R, Fak F, Backhed F (2010) Effects of gut microbiota on obesity and atherosclerosis via modulation of inflammation and lipid metabolism. J Intern Med 268: 320–328.
[83]
Yu PH, Wright S, Fan EH, Lun ZR, Gubisne-Harberle D (2003) Physiological and pathological implications of semicarbazide-sensitive amine oxidase. Biochim Biophys Acta 1647: 193–199.
[84]
Komatsubara S, Murata K, Kisumi M, Chibata I (1978) Threonine degradation by Serratia marcescens. J Bacteriol 135: 318–323.
[85]
Aoyama Y, Motokawa Y (1981) L-Threonine dehydrogenase of chicken liver. Purification, characterization, and physiological significance. J Biol Chem 256: 12367–12373.
