Abstract
Cross-linking enzymes such as transglutaminase, laccase, tyrosinase and peroxidase catalyse the formation of new covalent bonds between proteins and therefore lead to their polymerisation. This approach has been studied extensively as a way to modify physical and textural properties of (fermented) dairy products, or to create well-defined micro- and nanostructures from milk proteins that could serve as techno-functional additives in foods or as nanocarriers for molecules with nutritional benefits. This chapter gives an overview of the most prominent cross-linking enzymes in dairy science and technology, as well as approaches that are commonly applied for the analysis of cross-linking reactions and the resulting protein structures. Finally, different applications of cross-linking enzymes for milk protein modification are discussed.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Abbate RA, Raak N, Boye S et al (2019) Asymmetric flow field flow fractionation for the investigation of caseins cross-linked by microbial transglutaminase. Food Hydrocoll 92:117–124
Akal HC, Koçak C, Özer HB (2018) Transglutaminase applications in dairy technology. In: Öztürkoğlu Budak Ş, Akal HC (eds) Microbial cultures and enzymes in dairy technology, 1st edn. IGI Global, Hershey, pp 152–181
Alexander M, Dalgleish DG (2006) Dynamic light scattering techniques and their applications in food science. Food Biophys 1:2–13
Ando H, Adachi M, Umeda K et al (1989) Purification and characteristics of a novel transglutaminase derived from microorganisms. Agric Biol Chem 53:2613–2617
Anema SG, de Kruif CG (2012) Lactoferrin binding to transglutaminase cross-linked casein micelles. Int Dairy J 26:83–87
Bahri A, Martin M, Gergely C et al (2018) Topographical and nanomechanical characterization of casein nanogel particles using atomic force microscopy. Food Hydrocoll 83:53–60
Bönisch MP, Lauber S, Kulozik U (2004) Effect of ultra-high temperature treatment on the enzymatic cross-linking of micellar casein and sodium caseinate by transglutaminase. J Food Sci 69:E398–E404
Bönisch MP, Tolkach A, Kulozik U (2006) Inactivation of an indigenous transglutaminase inhibitor in milk serum by means of UHT-treatment and membrane separation techniques. Int Dairy J 16:669–678
Bönisch MP, Lauber S, Kulozik U (2007a) Improvement of enzymatic cross-linking of casein micelles with transglutaminase by glutathione addition. Int Dairy J 17:3–11
Bönisch MP, Huss M, Lauber S et al (2007b) Yoghurt gel formation by means of enzymatic protein cross-linking during microbial fermentation. Food Hydrocoll 21:585–595
Bönisch M, Heidebach T, Kulozik U (2008) Influence of transglutaminase protein cross-linking on the rennet coagulation of casein. Food Hydrocoll 22:288–297
Buchert J, Ercili Cura D, Ma H et al (2010) Crosslinking food proteins for improved functionality. Annu Rev Food Sci Technol 1:113–138
Casanova F, Nogueira Silva NF, Gaucheron F et al (2017) Stability of casein micelles cross-linked with genipin: a physicochemical study as a function of pH. Int Dairy J 68:70–74
Cozzolino A, Di Pierro P, Mariniello L et al (2003) Incorporation of whey proteins into cheese curd by using transglutaminase. Biotechnol Appl Biochem 38:289–295
Cragnell C, Choi J, Segad M et al (2017) Bovine β-casein has a polydisperse distribution of equilibrium micelles. Food Hydrocoll 70:65–68
de Jong GAH, Wijngaards G, Koppelman SJ (2003) Transglutaminase inhibitor from milk. J Food Sci 68:820–825
Dhayal SK, Gruppen H, de Vries R et al (2014) Controlled formation of protein nanoparticles by enzymatic cross-linking of α-lactalbumin with horseradish peroxidase. Food Hydrocoll 36:53–59
Dhayal SK, Sforza S, Wierenga PA et al (2015a) Peroxidase induced oligo-tyrosine cross-links during polymerization of α-lactalbumin. Biochim Biophys Acta 1854:1898–1905
Dhayal SK, Delahaije RJBM, de Vries RJ et al (2015b) Enzymatic cross-linking of α-lactalbumin to produce nanoparticles with increased foam stability. Soft Matter 11:7888–7898
Dinnella C, Gargaro MT, Rossano R et al (2002) Spectrophotometric assay using o-phtaldialdehyde for the determination of transglutaminase activity on casein. Food Chem 78:363–368
Djoullah A, Sok N, Djemaoune Y et al (2015) Monitoring of transglutaminase crosslinking reaction by 1H NMR spectroscopy on model substrates. Colloid Surf A 475:69–74
Duerasch A, Wissel J, Henle T (2018) Reassembling of alkali-treated casein micelles by microbial transglutaminase. J Agric Food Chem 66:11748–11756
Eissa AS, Bisram S, Khan SA (2004) Polymerization and gelation of whey protein isolates at low pH using transglutaminase enzyme. J Agric Food Chem 52:4456–4464
Ercili Cura D, Lantto R, Lille M et al (2009) Laccase-aided protein modification: effects on the structural properties of acidified sodium caseinate gels. Int Dairy J 19:737–745
Ercili Cura D, Lille M, Partanen R et al (2010) Effect of Trichoderma reesei tyrosinase on rheology and microstructure of acidified milk gels. Int Dairy J 20:830–837
Faccio G, Kruus K, Saloheimo M et al (2012) Bacterial tyrosinases and their applications. Process Biochem 47:1749–1760
Færgemand M, Sørensen MV, Jørgensen U et al (1999) Transglutaminase: effect on instrumental and sensory texture of set-style yoghurt. Milchwissenschaft 54:563–566
Folk JE, Cole PW (1965) Structural requirements of specific substrates for guinea pig liver transglutaminase. J Biol Chem 240:2951–2960
Folk JE, Cole PW (1966) Mechanism of action of guinea pig liver transglutaminase. I. Purification and properties of the enzyme: identification of a functional cysteine essential for activity. J Biol Chem 241:5518–5525
Gaspar ALC, de Góes-Favoni SP (2015) Action of microbial transglutaminase (MTGase) in the modification of food proteins: a review. Food Chem 171:315–322
Gharibzahedi SMT, Chronakis IS (2018) Crosslinking of milk proteins by microbial transglutaminase: utilization in functional yogurt products. Food Chem 245:620–632
Gharibzahedi SMT, Koubaa M, Barba FJ et al (2018a) Recent advances in the application of microbial transglutaminase crosslinking in cheese and ice cream products: a review. Int J Biol Macromol 107:2364–2374
Gharibzahedi SMT, Roohinejad S, George S et al (2018b) Innovative food processing technologies on the transglutaminase functionality in protein-based food products: trends, opportunities and drawbacks. Trends Food Sci Technol 75:194–205
Gharibzahedi SMT, George S, Greiner R et al (2018c) New trends in the microencapsulation of functional fatty acid-rich oils using transglutaminase catalyzed crosslinking. Compr Rev Food Sci Food Saf 17:274–289
Giosafatto CVL, Al-Asmar A, Mariniello L (2018) Transglutaminase protein substrates of food interest. In: Kuddus M (ed) Enzymes in food technology, 1st edn. Springer, Singapore, pp 293–317
Glantz M, Håkansson A, Lindmark Månsson H et al (2010) Revealing the size, conformation, and shape of casein micelles and aggregates with asymmetrical flow field-flow fractionation and multiangle light scattering. Langmuir 26:12585–12591
HadjSadok A, Pitkowski A, Nicolai T et al (2008) Characterisation of sodium caseinate as a function of ionic strength, pH and temperature using static and dynamic light scattering. Food Hydrocoll 22:1460–1466
Hagel L (2011) Gel filtration: size exclusion chromatography. In: Janson J-C (ed) Protein purification: principles, high resolution methods, and applications, 3rd edn. Wiley, Hoboken, pp 51–91
Heber A, Paasch S, Partschefeld C et al (2012) 31P NMR spectroscopic investigations of caseins treated with microbial transglutaminase. Food Hydrocoll 28:36–45
Heck T, Faccio G, Richter M et al (2013) Enzyme-catalyzed protein crosslinking. Appl Microbiol Biotechnol 97:461–475
Heijnis WH, Wierenga PA, Janssen AEM et al (2010a) In-line quantification of peroxidase-catalyzed cross-linking of α-lactalbumin in a microreactor. Chem Eng J 157:189–193
Heijnis WH, Wierenga PA, van Berkel WJH et al (2010b) Directing the oligomer size distribution of peroxidase-mediated cross-linked bovine α-lactalbumin. J Agric Food Chem 58:5692–5697
Heijnis WH, Dekker HL, de Koning LJ et al (2011) Identification of the peroxidase-generated intermolecular dityrosine cross-link in bovine α-lactalbumin. J Agric Food Chem 59:444–449
Hinz K, Huppertz T, Kelly AL (2012) Susceptibility of the individual caseins in reconstituted skim milk to cross-linking by transglutaminase: influence of temperature, pH and mineral equilibria. J Dairy Res 79:414–421
Huppertz T (2009) Novel applications of enzymes in the dairy sector: optimizing functional properties of milk proteins by enzymatic cross-linking. In: Corredig M (ed) Dairy-derived ingredients: food and nutraceutical uses, 1st edn. Woodhead, Cambridge, pp 394–416
Huppertz T (2014) Heat stability of transglutaminase-treated milk. Int Dairy J 38:183–186
Huppertz T, de Kruif CG (2007a) High pressure-induced solubilisation of micellar calcium phosphate from cross-linked casein micelles. Colloid Surf A 295:264–268
Huppertz T, de Kruif CG (2007b) Ethanol stability of casein micelles cross-linked with transglutaminase. Int Dairy J 17:436–441
Huppertz T, de Kruif CG (2007c) Rennet-induced coagulation of enzymatically cross-linked casein micelles. Int Dairy J 17:442–447
Huppertz T, de Kruif CG (2008) Structure and stability of nanogel particles prepared by internal cross-linking of casein micelles. Int Dairy J 18:556–565
Huppertz T, Smiddy MA, de Kruif CG (2007) Biocompatible micro-gel particles from cross-linked casein micelles. Biomacromolecules 8:1300–1305
Huppertz T, Gazi I, Luyten H et al (2017) Hydration of casein micelles and caseinates: implications for casein micelle structure. Int Dairy J 74:1–11
Isaschar-Ovdat S, Fishman A (2018) Crosslinking of food proteins mediated by oxidative enzymes – a review. Trends Food Sci Technol 72:134–143
Jacob M, Nöbel S, Jaros D et al (2011) Physical properties of acid milk gels: acidification rate significantly interacts with cross-linking and heat treatment of milk. Food Hydrocoll 25:928–934
Jaros D, Rohm H (2016) Enzymes exogenous to milk in dairy technology: transglutaminase. In: Smithers GW (ed) Reference module in food science. Elsevier, Amstersam
Jaros D, Partschefeld C, Henle T et al (2006a) Transglutaminase in dairy products: chemistry, physics, applications. J Texture Stud 37:113–155
Jaros D, Pätzold J, Schwarzenbolz U et al (2006b) Small and large deformation rheology of acid gels from transglutaminase treated milks. Food Biophys 1:124–132
Jaros D, Jacob M, Otto C et al (2010) Excessive cross-linking of caseins by microbial transglutaminase and its impact on physical properties of acidified milk gels. Int Dairy J 20:321–327
Jaros D, Schwarzenbolz U, Raak N et al (2014) Cross-linking with microbial transglutaminase: relationship between polymerisation degree and stiffness of acid casein gels. Int Dairy J 38:174–178
Kieliszek M, Misiewicz A (2014) Microbial transglutaminase and its application in the food industry. A review. Folia Microbiol 59:241–250
Kütemeyer C, Froeck M, Werlein H-D et al (2005) The influence of salts and temperature on enzymatic activity of microbial transglutaminase. Food Control 16:735–737
Labus K, Bryjak J, Polakovič M (2015) Kinetics of thermal inactivation of immobilized Agaricus bisporus tyrosinase. J Mol Catal B 120:136–140
Lam E, Holt C, Edwards P et al (2017) The effect of transglutaminase treatment on the physico-chemical properties of skim milk with added ethylenediaminetetraacetic acid. Food Hydrocoll 69:329–340
Lam E, McKinnon I, Marchesseau S et al (2018) The effect of transglutaminase on reconstituted skim milks at alkaline pH. Food Hydrocoll 85:10–20
Lauber S, Henle T, Klostermeyer H (2000) Relationship between the crosslinking of caseins by transglutaminase and the gel strength of yoghurt. Eur Food Res Technol 210:305–309
Loi M, Quintieri L, Fanelli F et al (2018) Application of a recombinant laccase-chlorogenic acid system in protein crosslink and antioxidant properties of the curd. Food Res Int 106:763–770
Lorenzen PC, Neve H, Mautner A et al (2002) Effect of enzymatic cross-linking of milk proteins on functional properties of set-style yoghurt. Int J Dairy Technol 55:152–157
Loveday SM, Sarkar A, Singh H (2013) Innovative yoghurts: novel processing technologies for improving acid milk gel texture. Trends Food Sci Technol 33:5–20
Lucey JA, Srinivasan M, Singh H et al (2000) Characterization of commercial and experimental sodium caseinates by multiangle laser light scattering and size-exclusion chromatography. J Agric Food Chem 48:1610–1616
Mautner A, Meisel H, Lorenzen PC et al (1999) Destimmung des Dipeptids ε-(γ-glutamyl)lysin aus Transglutaminase-vernetzten Proteinen mittels Aminosäureanalyse. Kieler Milchw Forsch 51:155–163
Miller ML, Johnson GVW (1999) Rapid, single-step procedure for the identification of transglutaminase-mediated isopeptide crosslinks in amino acid digests. J Chromatogr B 732:65–72
Mokoonlall A, Pfannstiel J, Struch M et al (2016a) Structure modification of stirred fermented milk gel due to laccase-catalysed protein crosslinking in a post-processing step. Innov Food Sci Emerg Technol 33:563–570
Mokoonlall A, Hippich M, Struch M et al (2016b) Antioxidant activity of milk suppresses laccase induced radicals and the subsequent modification of acidified milk protein gels. Int Dairy J 60:24–31
Mokoonlall A, Nöbel S, Hinrichs J (2016c) Post-processing of fermented milk to stirred products: reviewing the effects on gel structure. Trends Food Sci Technol 54:26–36
Mokoonlall A, Sykora L, Pfannstiel J et al (2016d) A feasibility study on the application of a laccase-mediator system in stirred yoghurt at the pilot scale. Food Hydrocoll 60:119–127
Monogioudi E, Permi P, Filpponen I et al (2011) Protein analysis by 31P NMR spectroscopy in ionic liquid: quantitative determination of enzymatically created cross-links. J Agric Food Chem 59:1352–1362
Moon J-H, Hong Y-H, Huppertz T et al (2009) Properties of casein micelles cross-linked by transglutaminase. Int J Dairy Technol 62:27–32
Mounsey JS, O’Kennedy BT, Kelly PM (2005) Influence of transglutaminase treatment on properties of micellar casein and products made therefrom. Lait 85:405–418
Nemes Z, Petrovski G, Fésüs L (2005) Tools for the detection and quantitation of protein transglutamination. Anal Biochem 342:1–10
Nilsson L (2013) Separation and characterization of food macromolecules using field-flow fractionation: a review. Food Hydrocoll 30:1–11
Nogueira Silva NF, Saint-Jalmes A, de Carvalho AF et al (2014) Development of casein microgels from cross-linking of casein micelles by genipin. Langmuir 30:10167–10175
Nogueira MH, Tavares GM, Nogueira Silva NF et al (2019) Physico-chemical stability of casein micelles cross-linked by transglutaminase as a function of acidic pH. Food Struct 19:100103
O’Mahony JA, Ardö Y, McSweeney PLH (2002) Analysis | electrophoresis. In: Roginski H, Fuquay JQ, Fox PF (eds) Encyclopedia of dairy sciences, 1st edn. Academic, New York, pp 67–74
Özrenk E (2006) The use of transglutaminase in dairy products. Int J Dairy Technol 59:1–7
Partschefeld C, Schwarzenbolz U, Richter S et al (2007) Crosslinking of casein by microbial transglutaminase and its resulting influence on the stability of micelle structure. Biotechnol J 2:456–461
Pecora R (2000) Dynamic light scattering measurement of nanometer particles in liquids. J Nanopart Res 2:123–131
Raak N, Abbate RA, Alkhalaf M et al (2020) Concentration-triggered liquid-to-solid transition of sodium caseinate suspensions as a function of temperature and enzymatic cross-linking. Food Hydrocoll 101:105464
Raak N, Rohm H, Jaros D (2017) Cross-linking with microbial transglutaminase: Isopeptide bonds and polymer size as drivers for acid casein gel stiffness. Int Dairy J 66:49–55
Raak N, Abbate RA, Lederer A et al (2018) Size separation techniques for the characterisation of cross-linked casein: a review of methods and their applications. Separations 5:14
Raak N, Brehm L, Abbate RA et al (2019a) Self-association of casein studied using enzymatic cross-linking at different temperatures. Food Biosci 28:89–98
Raak N, Schöne C, Rohm H et al (2019b) Acid-induced gelation of enzymatically cross-linked caseinate in different ionic milieus. Food Hydrocoll 86:43–49
Romeih E, Walker G (2017) Recent advances on microbial transglutaminase and dairy application. Trends Food Sci Technol 62:133–140
Sakamoto H, Kumazawa Y, Kawajiri H et al (1995) ε-(γ-glutamyl)lysine crosslink distribution in foods as determined by improved method. J Food Sci 60:416–420
Saricay Y, Dhayal SK, Wierenga PA et al (2012) Protein cluster formation during enzymatic cross-linking of globular proteins. Faraday Discuss 158:51–63
Saricay Y, Wierenga P, de Vries R (2013) Nanostructure development during peroxidase catalysed cross-linking of α-lactalbumin. Food Hydrocoll 33:280–288
Saricay Y, Wierenga PA, de Vries R (2014) Changes in protein conformation and surface hydrophobicity upon peroxidase-catalyzed cross-linking of apo-α-lactalbumin. J Agric Food Chem 62:9345–9352
Saricay Y, Wierenga PA, de Vries R (2016) Rheological properties of dispersions of enzymatically cross-linked apo-α-lactalbumin. Food Hydrocoll 56:344–351
Saricay Y, Wierenga PA, de Vries R (2017) Limited changes in physical and rheological properties of peroxidase-cross-linked apo-α-lactalbumin after heat treatment. Food Hydrocoll 66:326–333
Schäfer C, Schott M, Brandl F et al (2005) Identification and quantification of ε-(γ-glutamyl)lysine in digests of enzymatically cross-linked leguminous proteins by high-performance liquid chromatography−electrospray ionization mass spectrometry (HPLC-ESI-MS). J Agric Food Chem 53:2830–2837
Smiddy MA, Martin J-EGH, Kelly AL et al (2006) Stability of casein micelles cross-linked by transglutaminase. J Dairy Sci 89:1906–1914
Snyder SL, Sobocinski PZ (1975) An improved 2,4,6-trinitrobenzenesulfonic acid method for the determination of amines. Anal Biochem 64:284–288
Stachel I, Schwarzenbolz U, Henle T et al (2010) Cross-linking of type I collagen with microbial transglutaminase: identification of cross-linking sites. Biomacromolecules 11:698–705
Struch M, Linke D, Mokoonlall A et al (2015) Laccase-catalysed cross-linking of a yoghurt-like model system made from skimmed milk with added food-grade mediators. Int Dairy J 49:89–94
Struch M, Krahe N-K, Linke D et al (2016) Dose dependent effects of a milk ion tolerant laccase on yoghurt gel structure. LWT Food Sci Technol 65:1144–1152
Veitch NC (2004) Horseradish peroxidase: a modern view of a classic enzyme. Phytochemistry 65:249–259
Wahlund K-G, Nilsson L (2012) Flow FFF – basics and key applications. In: Williams SKR, Caldwell KD (eds) Field-flow fractionation in biopolymer analysis, 1st edn. Springer, Vienna, pp 1–21
Wang P, Chen C, Guo H et al (2018) Casein gel particles as novel soft Pickering stabilizers: the emulsifying property and packing behaviour at the oil-water interface. Food Hydrocoll 77:689–698
Weidendorfer K, Hinrichs K (2010) Online particle size measurement in microgel particle suspensions: principles and data analysis. Chem Ing Tech 82:1685–1691
Westermeier R (2011) Electrophoresis in gels. In: Janson J-C (ed) Protein purification: principles, high resolution methods, and applications, 3rd edn. Wiley, Hoboken, pp 365–377
Wyatt PJ (1993) Light scattering and the absolute characterization of macromolecules. Anal Chim Acta 272:1–40
Yokoyama K, Nio N, Kikuchi Y (2004) Properties and applications of microbial transglutaminase. Appl Microbiol Biotechnol 64:447–454
Zeeb B, Fischer L, Weiss J (2014) Stabilization of food dispersions by enzymes. Food Funct 5:198–213
Zeeb B, McClements DJ, Weiss J (2017) Enzyme-based strategies for structuring foods for improved functionality. Annu Rev Food Sci Technol 8:21–34
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2021 Springer Nature Switzerland AG
About this chapter
Cite this chapter
Raak, N., Rohm, H., Jaros, D. (2021). Enzymatic Protein Cross-Linking in Dairy Science and Technology. In: Kelly, A.L., Larsen, L.B. (eds) Agents of Change. Food Engineering Series. Springer, Cham. https://doi.org/10.1007/978-3-030-55482-8_17
Download citation
DOI: https://doi.org/10.1007/978-3-030-55482-8_17
Published:
Publisher Name: Springer, Cham
Print ISBN: 978-3-030-55481-1
Online ISBN: 978-3-030-55482-8
eBook Packages: Chemistry and Materials ScienceChemistry and Material Science (R0)