Abstract
Laccase is a versatile enzyme widely used for the oxidation of environmental contaminants and exhibits great potential in many others applications; however, it undergoes photo-degradation when irradiated with UVB light. The photo-stability of this biomolecule can be improved by immobilization in different encapsulation media and reverse micelles have been employed with this purpose. The laccase activity using syringaldazine as substrate has been studied in the absence and in the presence of reverse micelles of 0.15 M of sodium 1,4-bis (2-ethylhexyl) sulfosuccinate (AOT) in isooctane at W0 ([H2O]/[AOT]) = 30, before and after irradiation of the enzyme with UVB light. The kinetic parameters, i.e., Michaelis–Menten constant (KM), catalytic constant (kCAT), and catalytic efficiency (kCAT/KM), were determined by spectroscopic measurements in the micellar system and in homogeneous aqueous medium. The distribution of the substrate in two pseudo-phases (micelle and organic solvent) was taking into account in the kinetic parameters’ determinations. The results obtained indicate that the nano-aggregate system confers a solubilization media in the water core of the micelle, both for the enzyme and the substrate, in which the catalytic function of the enzyme is preserved. On the other hand, in homogeneous aqueous medium kCAT/KM value, it is reduced by ~50% after UVB irradiation of the enzyme, while in micellar medium, less than 10% of the activity was affected. This mean that the enzyme achieves a considerably photo-protection when it is irradiated with UVB light in reverse micelles as compared with the homogeneous aqueous medium. This phenomenon can be mainly due to the confinement of the biomolecule inside the micelle. Physical properties of the nano-environment could affect photochemical reactions.
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References
Abd El-Gawad HS (2014) Aquatic environmental monitoring and removal efficiency of detergents. Water Sci 28:51–64. https://doi.org/10.1016/j.wsj.2014.09.001
Aguilar LF, Abuin E, Lissi E (2001) A procedure for the joint evaluation of substrate partitioning and kinetic parameters for reactions catalyzed by enzymes in reverse micellar solutions. Arch Biochem Biophys 388:231–236. https://doi.org/10.1006/abbi.2001.2289
Arregui L, Ayala M, Gómez-Gil X, Gutiérrez-Soto G, Carlos Hernández-Luna CE, Herrera de los Santos M, Levin L, Rojo-Domínguez A, Romero-Martínez D, Saparrat MCN, Trujillo-Roldán MA, Valdez-Cruz NA (2019) Laccases: structure, function, and potential application in water bioremediation. Microb Cell Fact 18:200. https://doi.org/10.1186/s12934-019-1248-0
Arsene ML, Răut I, Călin M, Jecu ML, Doni M, Gurban AM (2021) Versatility of reverse micelles: from biomimetic models to nano (bio)sensor design. Processes 9:345. https://doi.org/10.3390/pr9020345
Biasutti MA, Abuin EA, Silber JJ, Correa NM, Lissi EA (2008) Kinetics of reactions catalyzed by enzymes in solutions of surfactants. Adv Colloid Interface Sci 136:1–24. https://doi.org/10.1016/j.cis.2007.07.001
Bilkis I, Silman I, Weiner L (2018) Generation of reactive oxygen species by photosensitizers and their modes of action on proteins. Curr Med Chem 25:5528–5539
Biswas R, Rohman N, Pradhan T, Buchner R (2008) Intramolecular charge transfers reaction, polarity, and dielectric relaxation in AOT/water/heptane reverse micelles: pool size dependence. J Phys Chem B 112:9379–9388. https://doi.org/10.1021/jp8023149
Bohidar HB, Behboudnia M (2001) Characterization of reverse micelles by dynamic light scattering. Colloids Surf A Physicochem Eng Asp 178:313–323. https://doi.org/10.1016/S0927-7757(00)00736-6
Cacciari RD, Reynoso A, Sosa S, Parodi F, Goldbaum FA, Montejano HA, Biasutti MA, Reynoso E (2020) Effect of UVB solar irradiation on laccase enzyme: evaluation of the photooxidation process and its impact over the enzymatic activity for pollutants bioremediation. Amino Acids 52:925–939. https://doi.org/10.1007/s00726-020-02861-0
Chhaya U, Gupte A (2013) Possible role of laccase from Fusarium incarnatum UC-14 in bioremediation of Bisphenol A using reverse micelles system. J Hazard Mater 254:149–156. https://doi.org/10.1016/j.jhazmat.2013.03.054
Correa NM, Durantini EN, Silber JJ (1999) Catalysis in micellar media. Kinetics and mechanism for the reaction of 1-Fluoro-2,4-dinitrobenzene with n-Butylamine and Piperidine in n-Hexane and AOT/n-Hexane/Water Reverse Micelles. J Org Chem 64:5757–5763. https://doi.org/10.1021/jo9818632
Correa NM, Durantini EN, Silber JJ (2001) Substituent effects on binding constants of carotenoids to n-heptane/AOT reverse micelles. J Colloid Interface Sci 240:573–580. https://doi.org/10.1006/jcis.2001.7640
Correa NM, Zorzan DH, D’Anteo L, Lasta E, Chiarini M, Cerichelli G (2004) Reverse micellar aggregates: effect on ketone reduction. 2. Surfactant role. J Org Chem 69:8231–8238. https://doi.org/10.1021/jo049172v
Correa NM, Silber JJ, Riter RE, Levinger NE (2012) Nonaqueous polar solvents in reverse micelle systems. Chem Rev 112:4569–4602. https://doi.org/10.1021/cr200254q
Deska M, Kończak B (2019) Immobilized fungal laccase as “green catalyst” for the decolourization process—state of the art. Process Biochem 84:112–123. https://doi.org/10.1016/j.procbio.2019.05.024
Falcone RD, Biasutti MA, Correa NM, Silber JJ, Lissi E, Abuin E (2004) Effect of the addition of a nonaqueous polar solvent (glycerol) on enzymatic catalysis in reverse micelles. Hydrolysis of 2-naphthyl acetate by α-chymotrypsin. Langmuir 20:5732–5737. https://doi.org/10.1021/la036243x
Feng Y, Lu K, Gao S, Mao L (2017) The fate and transformation of tetrabromobisphenol A in natural waters, mediated by oxidoreductase enzymes. Environ Sci Process Imp 19:596–604. https://doi.org/10.1039/C6EM00703A
Fernández-Fernández M, Sanromán MÁ, Moldes D (2013) Recent developments and applications of immobilized laccase. Biotechnol Adv 31:1808–1825. https://doi.org/10.1016/j.biotechadv.2012.02.013
Iark D, dos Reis Buzzo AJ, Garcia JAA, Côrrea VG, Helm CV, Corrêa RCG, Peralta RA, Moreira RF, Bracht A, Peralta RM (2019) Enzymatic degradation and detoxification of azo dye Congo red by a new laccase from Oudemansiella canarii. Biores Technol 289:121655. https://doi.org/10.1016/j.biortech.2019.121655
Janusz G, Pawlik A, Swiderska-Burek U, Polak J, Sulej J, Jarosz-Wilkołazka A, Paszczynski A (2020) Laccase properties, physiological functions, and evolution. Int J Mol Sci 221:966. https://doi.org/10.3390/ijms21030966
Johnson KA, Goody RS (2011) The original Michaelis constant: translation of the 1913 Michaelis–Menten paper. Biochemistry 50:8264–8269. https://doi.org/10.1021/bi201284u
Jones SM, Solomon EI (2015) Electron transfer and reaction mechanism of laccases. Cell Mol Life Sci 72:869–883. https://doi.org/10.1007/s00018-014-1826-6
Kang C, Ren D, Zhang S, Zhang X, He X, Deng Z, Huang C, Guo H (2019) Effect of polyhydroxyl compounds on the thermal stability and structure of laccase. Pol J Environ Stud 28:3253–3259. https://doi.org/10.15244/pjoes/94839
Kelbert M, Senna Pereira C, Daronch NA, Cesca K, Michels C, de Oliveira D, Moreira Soares H (2021) Laccase as an efficacious approach to remove anticancer drugs. A study of doxorubicin degradation, kinetic parameters and toxicity assessment. J Hazard Mater 409:124520. https://doi.org/10.1016/j.jhazmat.2020.124520
Ketelaar JAA, Van de Stolpe C, Gersmann HR (1951) Spectrophotometric study of the solvation of iodine in dioxan solution. Rec Trav Chim 70:499–508. https://doi.org/10.1002/recl.19510700604
Kurniawati S, Nicell JA (2008) Characterization of Trametes versicolor laccase for the transformation of aqueous phenol. Biores Technol 99:7825–7834. https://doi.org/10.1016/j.biortech.2008.01.084
Lakowicz JR (2006) Principles of fluorescence spectroscopy, 3rd edn. Springer, Baltimore, MD
Lissi EA, Abuin EB (2000) A general treatment for meaningful comparison of rate parameters of enzyme-catalyzed reactions in aqueous and reverse micellar solutions. Langmuir 16:10084–10086. https://doi.org/10.1021/la000788z
Luiz M, Biasutti MA, García NA (2004) Effect of reverse micelles on the Rose Bengal-sensitized photo-oxidation of 1-and 2-hydroxynaphthalenes. Red Rep 9:199–205. https://doi.org/10.1179/135100004225005165
Manole A, Herea D, Chiriac H, Melnig V (2008) Laccase activity determination. Biomater Biophys Med Phys Ecol 1:17–24
Margot J, Bennati-Granier C, Maillard J, Blánquez P, Barry DA, Holliger C (2013) Bacterial versus fungal laccase: potential for micropollutant degradation. AMB Expr 3:1–14. https://doi.org/10.1186/2191-0855-3-63
Mena-Giraldo P, Orozco J (2022) Photosensitive polymeric Janus micromotor for enzymatic activity protection and enhanced substrate degradation. ACS Appl Mater Interfaces 14:5897–5907. https://doi.org/10.1021/acsami.1c14663
Michizoe J, Goto M, Furusaki S (2001) Catalytic activity of lactase hosted in reversed micelles. J Biosci Bioeng 92:67–71. https://doi.org/10.1016/S1389-1723(01)80201-2
Michizoe J, Ichinose H, Kamiya N, Maruyama T, Goto M (2005) Biodegradation of phenolic environmental pollutants by a surfactant-laccase complex in organic media. J Biosci Bioeng 99:642–647. https://doi.org/10.1263/jbb.99.642
Moyano F, Falcone RD, Mejuto JC, Silber JJ, Correa NM (2010) Cationic reverse micelles create water with super hydrogen-bond-donor capacity for enzymatic catalysis: hydrolysis of 2-naphthyl acetate by α-chymotrypsin. Chem Eur J 16:8887–8893. https://doi.org/10.1002/chem.201000437
Okazaki S, Michizoe J, Goto M, Furusaki S, Wariishi H, Hi T (2002) Oxidation of bisphenol A catalyzed by laccase hosted in reversed micelles in organic media. Enzyme Microb Technol 31:227–232. https://doi.org/10.1016/S0141-0229(02)00104-7
Pattison DI, Rahmanto AS, Davies MJ (2012) Photooxidation of proteins. Photochem Photobiol Sci 11:38–53. https://doi.org/10.1039/C1PP05164D
Prasetyo EN, Kudanga T, Steiner W, Murkovic M, Nyanhongo GS, Guebitz GM (2010) Laccase-generated tetramethoxy azobismethylene quinone (TMAMQ) as a tool for antioxidant activity measurement. Food Chem 118:437–444. https://doi.org/10.1016/j.foodchem.2009.04.102
Punekar NS (2018) Enzymes: catalysis, kinetics and mechanisms. Springer, New York
Rajendiran N, Balasubramanian T (2007) Dual fluorescence of syringaldazine. Spectrochim Acta A 68:894–904. https://doi.org/10.1016/j.saa.2007.01.004
Rao MA, Scelza R, Acevedo F, Diez MC, Gianfreda L (2014) Enzymes as useful tools for environmental purposes. Chemosphere 107:145–162. https://doi.org/10.1016/j.chemosphere.2013.12.059
Reynoso E, Biasutti MA, García NA (2008) Kinetics of photosensitized oxidation of chymotrypsin in different media. Amino Acids 34:61–68. https://doi.org/10.1007/s00726-007-0591-3
Reynoso E, Cacciari RD, Suchetti C, Montejano HA, Biasutti MA (2019) Influence of pH and micellar system on the sensitized photooxidation of bovine serum albumin. Luminescence 34:324–333. https://doi.org/10.1002/bio.3610
Rodakiewicz-Nowak J (2000) Phenols oxidizing enzymes in water-restricted media. Top Cat 11:419–434. https://doi.org/10.1023/A:1027291629302
Sar P, Saha B (2020) Potential application of micellar nanoreactor for electron transfer reactions mediated by a variety of oxidants: a review. Adv Colloid Interface Sci 284:102241. https://doi.org/10.1016/j.cis.2020.102241
Sar P, Ghosh A, Scarso A, Saha B (2019) Surfactant for better tomorrow: applied aspect of surfactant aggregates from laboratory to industry. Res Chem Intermed 45:6021–6041. https://doi.org/10.1007/s11164-019-04017-6
Shapovalova OE, Levy D, Avnitr D, Vinogradov VV (2016) Protection of enzymes from photodegradation by entrapment within alumina. Colloid Surf B 146:731–736. https://doi.org/10.1016/j.colsurfb.2016.07.020
Silber JJ, Biasutti A, Abuin E, Lissi E (1999) Interactions of small molecules with reverse micelles. Adv Colloid Interface Sci 82:189–252. https://doi.org/10.1016/S0001-8686(99)00018-4
Silva OF, Fernández MA, Silber JJ, Rossi RH, Correa NM (2012) Inhibited phenol ionization in reverse micelles: confinement effect at the nanometer scale. Chem Phys Chem 13:124–130. https://doi.org/10.1002/cphc.201100634
Solterman T, Luiz M, Biasutti MA, Carrascoso M, Amat-Guerri F, García NA (1999) Monosubstituted naphthalenes as quenchers and generators of singlet molecular oxygen. J Photochem Photobiol A 129:25–32. https://doi.org/10.1016/S1010-6030(99)00172-0
Tonova K, Lazarova Z (2008) Reversed micelle solvents as tools of enzyme purification and enzyme-catalyzed conversion. Biotechnol Adv 26:516–532. https://doi.org/10.1016/j.biotechadv.2008.06.002
Xu P, Du H, Peng X, Tang Y, Zhou Y, Chen X, Fei J, Meng Y, Yuan L (2020) Degradation of several polycyclic aromatic hydrocarbons by laccase in reverse micelle system. Sci Total Environ 708:134970. https://doi.org/10.1016/j.scitotenv.2019.134970
Zdarta J, Meyer AS, Jesionowski T, Pinelo M (2018) Developments in support materials for immobilization of oxidoreductases: a comprehensive review. Adv Colloid Interf Sci 258:1–20. https://doi.org/10.1016/j.cis.2018.07.004
Zhou W, Zhang W, Cai Y (2021) Laccase immobilization for water purification: a comprehensive review. Chem Eng J 403:126272. https://doi.org/10.1016/j.cej.2020.126272
Acknowledgements
Financial support from Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET) and Secretaría de Ciencia y Técnica de la Universidad Nacional de Río Cuarto (SECyT, UNRC), both from Argentina, is gratefully acknowledged. Eugenia Reynoso, M. Alicia Biasutti, and Hernán A. Montejano are permanent research staff of CONICET; Facundo Parodi would like to thank CONICET for doctoral scholarships. R. Daniel Cacciari would like to thank CONICET for post-doctoral scholarships. Jeremías N. Mazalú would like to thank SECyT-UNRC for undergraduate scholarships.
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FP, RDC, JNM, HAM, ER, and MAB: investigation; FP, RDC, and JNM: experimental and data analysis; HAM, ER, and MAB: original draft preparation; ER and MAB: writing and editing; ER and MAB: supervision; MAB: project administration. All authors reviewed the manuscript.
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Parodi, F., Cacciari, R.D., Mazalu, J.N. et al. UVB light influence on the laccase enzyme catalytic activity in reverse micelles and in homogeneous aqueous medium. Amino Acids 55, 469–479 (2023). https://doi.org/10.1007/s00726-023-03237-w
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DOI: https://doi.org/10.1007/s00726-023-03237-w