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1

Graphene oxide flake activation via divinylsulfone – a procedure for efficient β-galactosidase immobilization

Trusek Anna

Polish Journal of Chemical Technology

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2019

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Vol. 21, nr 1

27--32

EN

Flaky graphene oxide was activated with divinylsulfone followed by immobilization of the β-galactosidase enzyme. An active and stable preparation was obtained. β-galactosidase stability after immobilization was much higher than with the native enzyme. The half-life time of the immobilized enzyme was estimated as 165 hours, while for the native form, the estimate was only 5 hours. The developed procedure for the preparation of flaked graphene and its use in the chemical immobilization of enzymes can be used for any enzyme. A processing solution for continuous operation was proposed and verified using cow’s milk, with lactose as the hydrolysed substrate, as a dosing stream. Lactose, a milk sugar, was effectively hydrolysed. Product for allergy sufferers who cannot digest lactose has been obtained in this way.

2

Temperature effect on properties of β-galactosidase entrapped in alginate matrix: an experimental research supported by molecular modeling

Labus K., Radosiński Ł., Kuchta B., Trusek-Hołownia A.

Inżynieria i Aparatura Chemiczna

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2015

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Nr 3

98--100

PL

Badania składały się z dwu części: eksperymentalnego wyznaczenia wpływu temperatury na właściwości immobilizowanej β-galaktozydazy oraz wstępnego modelowania dynamiki zmian w strukturze matrycy alginian-Ca2+ w zależności od temperatury na poziomie molekularnym. Na podstawie uzyskanych wyników można stwierdzić, że takie połączenie badań eksperymentalnych z symulacjami komputerowymi na poziomie cząsteczkowym pozwala na uzyskanie bardziej szczegółowej wiedzy na temat dynamiki układu, co w konsekwencji stwarza możliwość lepszej kontroli i optymalizacji procesu prowadzonego z wykorzystaniem immobilizowanego enzymu.

EN

The current study was separated into two parts: experimental study of temperature impact on activity and stability of immobilized β-galactosidase and preliminary modeling of changes generated in alginate-Ca2+ matrix with respect to temperature using molecular dynamics approach. On the basis of obtained results it can be stated that combination of experimental research and computer simulations on molecular level provides an additional insight greatly enhancing capabilities to control and optimize the process using immobilized biocatalyst.

3

Influence of Ionic Liquids on growth of Escherichia coli strain possessing β-galactosidase activity

Stobińska M., Urbański D., Bartkowiak A.

Challenges of Modern Technology

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2013

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Vol. 4, no. 1

38--40

EN

Ionic liquids (ILs) have gained a significant attention as alternative solvents in many chemical and biological reactions. ILs have evolved as a new type of non-aqueous solvents for biocatalysis, because they can stabilize and activate enzymes. Ionic liquids to be considered as so called “green solvents” should possess only very low or no toxicity to living organism and the environment. Currently, very little is known about the biotoxicity of ionic liquids. Within this paper a relative toxicity of two ionic liquids 1,3 dimethylimidazolium methyl sulfate [MMIM][MeSO4] and 1-hexyl-3-methylimidazolium chloride [HMIM][Cl]) on the growth of Escherichia coli (E.coli) was evaluated. In this study we focused on ionic liquids, which potentially could increase activity of β-galactosidase produced by selected strain of E. coli. The inhibition of cell growth in the presence of various ionic liquids was determined using both, solid and liquid cultures. Liquid culture was incubated in culture medium with different concentrations of ionic liquids (0.1, 1, 5, 10% (v/v)). Determination of the minimal inhibitory concentration (MIC50) of ionic liquids was based on monitoring of E. coli growth by optical density measured at 600 nm using densitometer. In the second method the solid culture plate with addition of various amount of ILs has been used. The number of colonies formed on the solid culture plate was converted to colony forming units (CFUs). The obtained results have confirmed, that prescreening based on cytotoxicity tests is a key element in selection procedure of ILs for any biotechnological processes based on application of microorganisms.

4

Immobilization of Aspergillus oryzae Β galactosidase on concanavalin A-layered calcium alginate-cellulose beads and its application in lactose hydrolysis in continuous spiral bed reactors

Ansari S. A, Husain Q.

Polish Journal of Chemical Technology

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2011

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Vol. 13, nr 4

15-20

EN

In this study, Aspergillus oryzae Β galactosidase was immobilized on concanavalin A layered calcium alginate-cellulose beads as a bioaffi nity support. Immobilized enzyme showed a remarkable broadening in temperature-activity profi les as compared to the native enzyme and exhibited 65% activity in the presence of 5% galactose. Michaelis constant (Km) was 2.57 mM and 5.38 mM for the free and the immobilized Β galactosidase, respectively. Crosslinked Β galactosidase showed greater catalytic activity in the presence of Mg2+ and was more stable during storage at 4°C for 6 weeks. Immobilized enzyme hydrolyzed 67% lactose in milk in 8 h and 85% lactose in whey in 9 h in the stirred batch process at 50oC. The continuous hydrolysis of lactose by crosslinked Β galactosidase in spiral bed reactor exhibited 93% and 88% hydrolysis of lactose at flow rate of 20 ml/h and 30 ml/h, after 1 month operation, respectively.