Ukr.Biochem.J. 2022; Volume 94, Issue 4, Jul-Aug, pp. 67-82


Distribution and polymorphism of enzymes involved in antioxidant protection and xenobiotics biotransformation in the mediterranean mussel Mytilus galloprovincialis

V. А. Toptikov, I. Yu. Chubyk*, S. V. Chebotar

Odesa National Mechnуkov University, Ukraine;

Received: 18 July 2022; Revised: 17 October 2022;
Accepted: 04 November 2022; Available on-line: 14 November 2022

The aim of the work was to determine the tissue distribution, activity and polymorphism of 13 enzymes involved in antioxidative protection and xenobiotics biotransformation in a five bivalve mussel organs (hepatopancreas, ctenidia, mantle, leg, adductor muscles). Iso-allozyme analysis was performed by electrophoresis, correlation relationships between the studied enzymes in the whole mussel body were carried out using Spearman’s correlation analysis. It was found that all studied enzymes are polymorphic, each organ differed in the level of enzyme activity and a set of multiple forms. The coordinated functioning of protective enzymes in various mussel organs was demonstrated.

Keywords: , , , ,


  1. Harrang E, Lapègue S, Morga B, Bierne N. A high load of non-neutral amino-acid polymorphisms explains high protein diversity despite moderate effective population size in a marine bivalve with sweepstakes reproduction. G3 (Bethesda). 2013;3(2):333-341. PubMed, PubMedCentral, CrossRef
  2. Zouros E, Foltz DW. Possible explanations of heterozygote deficiency in bivalve molluscs. Malacologia. 1984; 25: 583-591.
  3. Mallet A, Zouros E, Gartner-Kepkay KE, Freeman K, Dickie LM. Larval viability and heterozygote deficiency in populations of marine bivalves: evidence from pair mating of mussels. Mar Biol. 1985; 87(2): 165-172.
  4. Raymond, M, Vaanto, RL, Thomas, F, Rousset, F, De Meuss, T, Renaud, F. Heterozygote deficiency in the mussel Mytilus edulis species complex revisited. Mar Ecol Progr Ser. 1997; 156: 225-237.
  5. Myrand B, Tremblay R, Sévigny JM. Selection against blue mussels (Mytilus edulis L.) homozygotes under various stressful conditions. J Hered. 200g;93(4):238-248. PubMed, CrossRef
  6. McDonald JH, Seed R, Koehn RK. Allozymes and morphometric characters of three species of Mytilus in the Northern and Southern Hemispheres. Mar Biol. 1991; 111: 323-333. CrossRef
  7. Gardner J, Thompson R. High levels of shared allozyme polymorphism among strongly differentiated congeneric clams of the genus Astarte (Bivalvia: Mollusca). Heredity. 1999;82:89-99. CrossRef
  8. Cárcamo C, Comesańa AS, Winkler FM, Sanjuan A. Allozyme identification of mussels (Bivalvia: Mytilus) on the Pacific coast of South America. J Shellfish Res. 2005;24(4):1101-1115. CrossRef2.0.CO;2″]
  9. Sáenz LA, Seibert E, Zanette J, Fiedler HD, Curtius AJ, Ferreira JF, Alves de Almeida E, Marques MRF, Bainy ACD. Biochemical biomarkers and metals in Perna perna mussels from mariculture zones of Santa Catarina, Brazil. Ecotoxicol Environ Saf. 2010;73(5):796-804. PubMed, CrossRef
  10. Pes K, Friese A, Cox CJ, Laizé V, Fernández I. Biochemical and molecular responses of the Mediterranean mussel (Mytilus galloprovincialis) to short-term exposure to three commonly prescribed drugs. Mar Environ Res. 2021;168:105309. PubMed, CrossRef
  11. Krasota LL. Assessment of the quality of the environment of the North-West parts of the Black Sea according to the results of biotesting of waters in 2008-2014 years. Sci Notes Ternopil Nat Pedagog Univ Volodymyr Hnatyuk. Series: Biology. 2015; 64(3-4): 358-361. (In Ukrainian).
  12. Nikolić, M, Kuznetsova, T, Kholodkevich, S, Gvozdenović, S, Mandić, M, Joksimović, D, Teodorović, I. Cardiac activity in the Mediterranean mussel (Mytilus galloprovincialis Lamarck, 1819) as a biomarker for assessing sea water quality in Boka Kotorska Bay, South Adriatic Sea. Mediterr Mar Sci. 2019;20(4):680-687. CrossRef
  13. Bakhmet IN, Sazhin A, Maximovich N, Ekimov D. In situ long-term monitoring of cardiac activity of two bivalve species from the White Sea, the blue mussel Mytilus edulis and horse mussel Modiolus modiolus. J Mar Biolog Assoc UK. 2019;99(4):833-840. CrossRef
  14. Newton TJ, Cope WG. Biomarker responses of unionid mussels to environmental contaminants. In: Freshwater Bivalve Ecotoxicology. (Farris JL, Van Hassel JH, eds). Boca Raton: CRC Press, 2007. P. 257-284.
  15. Faucet J, Maurice M, Gagnaire B, Renault T, Burgeot T. Isolation and primary culture of gill and digestive gland cells from the common mussel Mytilus edulis. Methods Cell Sci. 2003;25(3-4):177-184. PubMed, CrossRef
  16. Trisciani A, Perra G, Caruso T, Focardi S, Corsi I. Phase I and II biotransformation enzymes and polycyclic aromatic hydrocarbons in the Mediterranean mussel (Mytilus galloprovincialis, Lamarck, 1819) collected in front of an oil refinery. Mar Environ Res. 2012;79:29-36. PubMed, CrossRef
  17. Manduzio H, Rocher B, Durand F, Galap C, Leboulenger F. The point about oxidative stress in molluscs. Invertebr Surviv J. 2005; 20: 814-823.
  18. Bartosz G. Reactive oxygen species: destroyers or messengers? Biochem Pharmacol. 2009;77(8):1303-1315. PubMed, CrossRef
  19. Valavanidis A, Vlahogianni T, Dassenakis M, Scoullos M. Molecular biomarkers of oxidative stress in aquatic organisms in relation to toxic environmental pollutants. Ecotoxicol Environ Saf. 2006;64(2):178-189. PubMed, CrossRef
  20. Alves de Almeida E, Celso Dias Bainy A, Paula de Melo Loureiro A, Regina Martinez G, Miyamoto S, Onuki J, Fujita Barbosa L, Carrião Machado Garcia C, Manso Prado F, Eliza Ronsein G, Alexandre Sigolo C, Barbosa Brochini C, Maria Gracioso Martins A, Helena Gennari de Medeiros M, Di Mascio P. Oxidative stress in Perna perna and other bivalves as indicators of environmental stress in the Brazilian marine environment: antioxidants, lipid peroxidation and DNA damage. Comp Biochem Physiol A Mol Integr Physiol. 2007;146(4):588-600. PubMed, CrossRef
  21. Lushchak VI. Environmentally induced oxidative stress in aquatic animals. Aquat Toxicol. 2011;101(1):13-30. PubMed, CrossRef
  22. Finkel T. Signal transduction by reactive oxygen species. J Cell Biol. 2011;194(1):7-15. PubMed, PubMedCentral, CrossRef
  23. Sies H, Jones DP. Reactive oxygen species (ROS) as pleiotropic physiological signalling agents. Nat Rev Mol Cell Biol. 2020;21(7):363-383. PubMed, CrossRef
  24. Toptikov VA. Genetic and biochemical studies of the adaptability of animals and their groups. Educational and methodical manual. (Toptikov VA, Ershova OM, Kovtun OO, Lavrenyuk TI, Totsky VM, eds). Odesa: ONU named after II Mechnikov, 2017. 140 p. (In Ukrainian).
  25. Toptikov V, Aleksyeyeva T, Kovtun O. Hydrolytic enzymes of Rapana venosa digestive system. Saarbrüken (Germany): LAP LAMBERT Academic Publishing, 2017. 65 p.
  26. Davis BI. Disc elektrophoresis. II. Method and application to human serum proteins. Ann N Y Acad Sci. 1964;121(2):404-427. PubMed, CrossRef
  27. Manchenko GP. Handbook of detection of enzymes on electrophoretic gels. CRC Press, 2003. 568 p.
  28. Meijer AE, Bloem JH. Improved histochemical demonstration of carbonate dehydratase. Acta Histochem. 1966;25(5):239-241. PubMed
  29. Lojda Z. A new method for demonstrating myeloperoxidase in paraffin sections (in Czech). Cs Patol. 1967;3:31-33.
  30. Podzharsky MA, Rybalka DG. AnaIS – Spectrum Image Analyzer. 2004. [Electronic resource] Site access mode:
  31. Atramentova LA. Easier nowhere. Research planning. Data analysis. Presentation of results. Kh: NTMT, 2018. 260 p. (In Russian).
  32. Ighodaro OM, Akinloye OA. First line defence antioxidants-superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX): Their fundamental role in the entire antioxidant defence grid. Alex J Med. 2018;54(4):287-293. CrossRef
  33. Geret F, Manduzio H, Company R, Leboulenger F, Bebianno MJ, Danger JM. Molecular cloning of superoxide dismutase (Cu/Zn-SOD) from aquatic molluscs. Mar Environ Res. 2004;58(2-5):619-623. PubMed, CrossRef
  34. Manduzio H, Monsinjon T, Galap C, Leboulenger F, Rocher B. Seasonal variations in antioxidant defences in blue mussels Mytilus edulis collected from a polluted area: major contributions in gills of an inducible isoform of Cu/Zn-superoxide dismutase and of glutathione S-transferase. Aquat Toxicol. 2004;70(1):83-93. PubMed, CrossRef
  35. Fernández C, San Miguel E, Fernández-Briera A. Superoxide dismutase and catalase: tissue activities and relation with age in the long-lived species Margaritifera margaritifera. Biol Res. 2009;42(1):57-68. PubMed
  36. Wu J, Bao M, Ge D, Huo L, Lv Z, Chi C, Liao Z, Liu H. The expression of superoxide dismutase in Mytilus coruscus under various stressors. Fish Shellfish Immunol. 2017;70:361-371. PubMed, CrossRef
  37. Gostyukhina OL, Andreenko TI. Activity of superoxide dismutase and catalase in tissues of three species of Black Sea bivalve mollusks: Cerastoderma glaucum (Bruguiere, 1789), Anadara kagoshimensis (Tokunaga, 1906) and Mytilus galloprovincialis Lam. in connection with adaptation to the conditions of their habitat. Zh Evol Biokhim Fiziol. 2020;56(2):108-118. (In Russian).
  38. Petrov, SA, Andriyevsky, OM, Budnyak, OK, Chernadchuk, SS, Sorokin, AV, Fedorko, NL, Karavansky, YuV, Zamorov, VV, Myronov, DA, Podgorny VV. Antioxidant protection system in the tissues of the Antarctic krill Euphausia superba and of the Black Sea shrimp Palaemon elegans. Hydrob J. 2022;58(5):78-84. CrossRef
  39. Nemoto M, Ren D, Herrera S, Pan S, TamuraT, Inagaki K, Kisailus D. Integrated transcriptomic and proteomic analyses of a molecular mechanism of radular teeth biomineralization in Cryptochiton stelleri. Sci Rep. 2019;9(1):856. PubMed, PubMedCentral, CrossRef
  40. Bannister JV, Bannister WH, Hill HA, Mahood JF, Willson RL, Wolfenden BS. Does caeruloplasmin dismute superoxide? No. FEBS Lett. 1980;118(1):127-129. PubMed, CrossRef
  41. Goldstein IM, Kaplan HB, Edelson HS. Ceruloplasmin: an acute phase reactant that scavenges oxygen-derived free radicals. Ann N Y Acad Sci. 1982;389:368-379. PubMed, CrossRef
  42. Sergeev AG, Pavlov AR, Revina AA, Yaropolov AI. The mechanism of interaction of ceruloplasmin with superoxide radicals. Int J Biochem. 1993;25(11):1549-1554. PubMed, CrossRef
  43. Zhang Y, Zhang R, Zou J, Hu X, Wang S, Zhang L, Bao Z. Identification and characterization of four ferritin subunits involved in immune defense of the Yesso scallop (Patinopecten yessoensis). Fish Shellfish Immunol. 2013;34(5):1178-1187. PubMed, CrossRef
  44. Sumithra TG, Neethu BR, Reshma KJ, Anusree VN, Reynold P, Sanil NK. A novel ferritin subunit gene from Asian green mussel, Perna viridis (Linnaeus, 1758). Fish Shellfish Immunol. 2021;115:1-6. PubMed, CrossRef
  45. Shcherbak GY, Tsarichkova DB, Verves YuG. Zoology of invertebrates: textbook: in 3 books. K: Lybid, 1996. 320 p. (In Ukrainian).
  46. Gerdol M, Gomez-Chiarri M, Castillo MG, Figueras A, Fiorito G, Moreira R, Novoa B, Pallavicini A, Ponte G, Roumbedakis K, Venier P, Vasta GR. Immunity in molluscs: recognition and effector mechanisms, with a focus on Bivalvia. In: Advances in Comparative Immunology. (Cooper E, eds). Cham: Springer, 2018. P. 225-341.
  47. Yurimoto T. Seasonal changes in glycogen contents in various tissues of the edible bivalves, pen shell Atrina lischkeana, ark shell Scapharca kagoshimensis, and manila clam Ruditapes philippinarum in West Japan. J Mar Biol. 2015. ID: 593032. CrossRef
  48. Cong R, Sun W, Liu G, Fan T, Meng X, Yang L, Zhu L. Purification and characterization of phenoloxidase from clam Ruditapes philippinarum. Fish Shellfish Immunol. 2005;18(1):61-70. PubMed, CrossRef
  49. Yang MS, Chan HW, Yu LC. Glutathione peroxidase and glutathione reductase activities are partially responsible for determining the susceptibility of cells to oxidative stress. Toxicology. 2006;226(2-3):126-130. PubMed, CrossRef
  50. Alfonso-Prieto M, Biarnés X, Vidossich P, Rovira C. The molecular mechanism of the catalase reaction. J Am Chem Soc. 2009;131(33):11751-11761. PubMed, CrossRef
  51. Gostyukhina OL, Golovina IV. Peculiarities of antioxidant defense system organization of the black sea mollusks Mytilus galloprovincialis Lam. and Anadara inaequivalvis Br. Ukr Biokhim Zhurn. 2012;84(3):31-36. (In Russian). PubMed
  52. Kulinsky VI, Kolesnichenko LS. Glutathione system. I. Synthesis, transport, glutathione transferases, glutathione peroxidases. Biomed Khim. 2009;55(3):255-277. (In Russian). PubMed
  53. Regoli F, Giuliani ME. Oxidative pathways of chemical toxicity and oxidative stress biomarkers in marine organisms. Mar Environ Res. 2014;93:106-117. PubMed, CrossRef
  54. Marques A, Pilo D, Araujo O, Pereira F, Guilherme S, Carvalho S, Santos AM, Pacheco M, Pereira P. Propensity to metal accumulation and oxidative stress responses of two benthic species (Cerastoderma edule and Nephtys hombergii): are tolerance processes limiting their responsiveness? Ecotoxicology. 2016;25(4):664-676. PubMed, CrossRef
  55. Gozhenko AI, Andreytsova NI, Kvasnytska OB. Biotransformation of exogenous oxidants in humans and animals. Actual Probl Transport Med. 2009;(4(18)):8-18. (In Ukrainian).
  56. Yanovych DO, Yanovych NE. Biotransformation of xenobiotics and mechanisms of its regulation. Scientific Messenger LNUVMBT named after SZ Gzhytskyj. 2011; 13(2(2)):305-311. (In Ukrainian).
  57. Sipes IG, Gandolfi AJ. Biotransformation of toxicants. In: Casarett and Doull’s toxicology: the basic science of poisons. (Klaasen CD, Amdur MO, Doull J, eds). 3rd edn. NY: Macmillan, 1986. P. 64-98.
  58. Li X, Schuler MA, Berenbaum MR. Molecular mechanisms of metabolic resistance to synthetic and natural xenobiotics. Annu Rev Entomol. 2007;52:231-253. PubMed, CrossRef
  59. Liu J, Zhao M, Song W, Ma L, Li X, Zhang F, Diao L, Pi Y, Jiang K. An amine oxidase gene from mud crab, Scylla paramamosain, regulates the neurotransmitters serotonin and dopamine in vitro. PLoS One. 2018;13(9):e0204325. PubMed, PubMedCentral, CrossRef
  60. Setini A, Pierucci F, Senatori O, Nicotra A. Molecular characterization of monoamine oxidase in zebrafish (Danio rerio). Comp Biochem Physiol B Biochem Mol Biol. 2005;140(1):153-161. PubMed, CrossRef
  61. Sugimoto H, Taguchi YD, Shibata K, Kinemuchi H. Molecular characteristics of a single and novel form of carp (Cyprinus carpio) monoamine oxidase. Comp Biochem Physiol B Biochem Mol Biol. 2010;155(3):266-271. PubMed, CrossRef
  62. Edmondson DE, Binda C, Mattevi A. The FAD binding sites of human monoamine oxidases A and B. Neurotoxicology. 2004;25(1-2):63-72.
    PubMed, CrossRef
  63. Bach AW, Lan NC, Johnson DL, Abell CW, Bembenek ME, Kwan SW, Seeburg PH, Shih JC. cDNA cloning of human liver monoamine oxidase A and B: molecular basis of differences in enzymatic properties. Proc Natl Acad Sci USA. 1988;85(13):4934-4938. PubMed, PubMedCentral, CrossRef
  64. Grimsby J, Lan NC, Neve R, Chen K, Shih JC. Tissue distribution of human monoamine oxidase A and B mRNA. J Neurochem. 1990;55(4):1166-1169. PubMed, CrossRef
  65. Rodriguez-Navarro C, Cizer Ö, Kudłacz K, Ibañez-Velasco A, Ruiz-Agudo C, Elert K, Burgos-Cara A, Ruiz-Agudo E. The multiple roles of carbonic anhydrase in calcium carbonate mineralization. Cryst Eng Comm. 2019;21(48):7407-7423. CrossRef
  66. Wong DL, Yuan T, Korkola NC, Stillman MJ. Interplay between Carbonic Anhydrases and Metallothioneins: Structural Control of Metalation. Int J Mol Sci. 2020;21(16):5697. PubMed, PubMedCentral, CrossRef
  67. Le Roy N, Jackson DJ, Marie B, Ramos-Silva P, Marin F. Carbonic anhydrase and metazoan biocalcification: a focus on molluscs. Key Eng Mater. 2016;672:151-157. CrossRef
  68. Ozensoy Guler O, Capasso C, Supuran CT. A magnificent enzyme superfamily: carbonic anhydrases, their purification and characterization. J Enzyme Inhib Med Chem. 2016;31(5):689-694. PubMed, CrossRef
  69. Perfetto R, Del Prete S, Vullo D, Sansone G, Barone C, Rossi M, Supuran CT, Capasso C. Biochemical characterization of the native α-carbonic anhydrase purified from the mantle of the Mediterranean mussel, Mytilus galloprovincialis. J Enzyme Inhib Med Chem. 2017;32(1):632-639. PubMed, PubMedCentral, CrossRef
  70. Bertucci A, Moya A, Tambutte S, Allemand D, Supuran CT, Zoccola D. Carbonic anhydrases in anthozoan corals-A review. Bioorg Med Chem. 2013;21(6):1437-1450. PubMed, CrossRef
  71. Leggat W, Dixon R, Saleh S, Yellowlees D. A novel carbonic anhydrase from the giant clam Tridacna gigas contains two carbonic anhydrase domains. FEBS J. 2005;272(13):3297-3305. PubMed, CrossRef
  72. Kuzmina NV, Ostapiv DD, Vlizlo VV. Activity of superoxide dismutase and glutathione peroxidase in different organs and blood of cows. Biol Animals. 2008;10(1-2):128-132. (In Ukrainian).

Creative CommonsThis work is licensed under a Creative Commons Attribution 4.0 International License.