Microbial β‐Galactosidases of industrial importance: Computational studies on the effects of point mutations on the lactose hydrolysis reaction

Andrade, Bruna Coelho de; Timmers, Luís Fernando Saraiva Macedo; Renard, Gaby; Volpato, Giandra; Souza, Cláucia Fernanda Volken de

doi:10.1002/btpr.2982

Cited by 11 publications

(3 citation statements)

References 35 publications

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“…Besides enzyme source, substrate concentration and the composition of the reaction medium are key factors that affect the enzyme activity (Gaur, Pant, Jain, & Khare, 2006). Lactose hydrolysis is a key industrial process, especially in the food industry, that generates galactose and glucose as reaction products (Basso & Serban, 2019;Andrade, Timmers, Renard, Volpato, & Souza, 2020). However, some authors have highlighted that this reaction can be hindered by product inhibition, such as glucose and galactose, which reduces the reaction rates of lactose hydrolysis.…”

Section: Resultsmentioning

confidence: 99%

Cell permeabilization of Kluyveromyces and Saccharomyces species to obtain potential biocatalysts for lactose hydrolysis

Alves

Bosso

Morioka

et al. 2022

Acta Sci. Biol. Sci.

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Yeast’s beta-galactosidase is an intracellular enzyme, through which it is possible to determine in vivo its activity as a biocatalyst in the lactose hydrolysis. Permeabilization process was used for transforming the microorganisms cells into biocatalysts with an enhanced enzyme activity. The potential application of this enzyme technology in industrial process depends mainly on the enzyme activity. Beta-galactosidase enzyme that hydrolyzes lactose, for instance, is largely dependent on the reaction time and its stability under different physical conditions, such as pH, temperature and enzyme concentration. The objective of this study was to optimize the cellular permeabilization process of Kluyveromyces marxianus CCT 3172 and Saccharomyces fragilis CCT 7586 cultured in cheese whey for lactose hydrolysis. Box-Behnken design was carried out for cell permeabilization with three independent variables, ethanol concentration, permeabilization time and temperature. The best permeability conditions for K. marxianus CCT 3172 were 27% (v v-1) ethanol, 3 min at 20ºC, with specific enzymatic activity of 0.98 U mg-1. For S. fragilis CCT 7586, a specific enzymatic activity of 1.31 U mg-1 was achieved using 45% (v v-1) of ethanol, 17 min. of reaction under 17ºC. Thus, it was concluded that cellular permeabilization with ethanol is an efficient process to determine beta-galactosidase activity.

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Section: Resultsmentioning

confidence: 99%

Cell permeabilization of Kluyveromyces and Saccharomyces species to obtain potential biocatalysts for lactose hydrolysis

Alves

Bosso

Morioka

et al. 2022

Acta Sci. Biol. Sci.

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“…As an environment-friendly biocatalyst, β-galactosidase has been widely studied for its recognized advantages. The ratio between lactose hydrolysis and GOS synthesis crucially depends on the concentration of lactose, temperature of the reactor, and intrinsic properties of enzymes De [ 7 , 8 ]. Therefore, it is of importance to optimize the conditions for GOS synthesis and to discover β-galactosidases with high transgalactosylation capacity.…”

Section: Introductionmentioning

confidence: 99%

β-galactosidase GALA fromBacillus circulanswith high transgalactosylation activity

Yan

Guan

et al. 2021

Bioengineered

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β-galactosidase catalyzes lactose hydrolysis and transfers reactions to produce prebiotics such as galacto-oligosaccharides (GOS) with potential application in the food industry and pharmaceuticals. However, there is still a need for improved transgalactosylation activity of β-galactosidases and reaction conditions of GOS production in order to maximize GOS output and reduce production costs. In this study, a β-galactosidase gene, galA, from Bacillus circulans was expressed in Pichia pastoris, which not only hydrolyzed lactose but also had strong transgalactosylation activity to produce GOS. Response surface methodology was adopted to investigate the effects of temperature, enzyme concentration, pH, initial lactose concentration and reaction time on production of GOS and optimize the reaction conditions for GOS.The optimal pH for the enzyme was 6.0 and remained stable in neutral and basic conditions. Meanwhile, GALA showed most activity at 50℃ and retained considerable activity at lower temperature 30-40°C, indicating this enzyme could work under mild conditions. The enzyme concentration and temperature were found to be the critical parameters affecting the transgalactosylation activity. Response surface methodology showed that the optimal enzyme concentration, initial lactose concentration, temperature, pH, and reaction time were 3.03 U/mL, 500 g/L, 30℃, 5.08, and 4 h, respectively. Under such conditions, the maximum yield of GOS was 252.8 g/L, accounting for approximately 50.56% of total sugar. This yield can be considered relatively high compared to those obtained from other sources of β-galactosidases, implying a great potential for GALA in the industrial production and application of GOS.

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“…3.18 shows the concentrationt of fructose after isomerisation in acid whey permeate samples was within the range of 19±3 to 24±1 g L -1 . The use of β-galactosidase in this study was limited because it is difficult to achieve a complete hydrolysis of lactose due to the side reactions (Andrade et al, 2020). Whey is a complex mixture of many ingredients, each of which has a significant effect on the hydrolysis of lactose.…”

Section: Att Cukuru Saturs Biezpena Sūkalu Ultrafiltrātā Izmantojot Glikozes Izomerāzimentioning

confidence: 99%

Valorisation of whey for lactose recycling products production = Sūkalu valorizācija laktozes pārstrādes produktu ieguvei

Majore¹

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The doctoral thesis “Valorisation of whey for lactose recycling products production” was developed from 2016 to 2021. Experiments were carried out in the research laboratories of the Faculty of Food Technology, Latvia University of Life Sciences and Technologies; Dairy Innovation Institute, California Polytechnic State University (USA); Institute of Microbiology and Biotechnology, Latvia University; Faculty of Chemistry, Latvia University; Institute of Solid State Physics, Latvia University and J.S. Hamilton Baltic Ltd. The aim of the doctoral thesis was to improve the lactose hydrolysis process for obtaining glucose-galactose and oligosaccharide syrups. The hypothesis of the doctoral thesis – the two-stage fermentation increases the sweetness of glucose-galactose syrup. The hypothesis of the doctoral thesis has been confirmed by the defended thesis: 1. The presence of cations affects the β-galactosidase activity in the sweet and acid whey permeate. 2. The chemical composition and quality of whey affect the physical properties of lactose. 3. Enzymatic reactions affect the functional and sensory properties of syrups. The research objects – sweet and acid whey permeates, glucose isomerase, commercial β-galactosidases and glucose-galactose syrup. The following tasks were set to achieve the aim of the doctoral thesis: 1. To evaluate the effect of cation concentration to ensure the β-galactosidase activity in substrate. 2. To investigate the physical properties of whey lactose in order to better understand its behaviour. 3. To study the changes of monosaccharide concentration in the lactose hydrolysis, varying with the solids concentration of the substrates and enzyme units. 4. To assess the possibilities of glucose isomerase to increase the sweetness of glucose-galactose syrup. 5. To evaluate the sensory properties of the developed syrups. The novelty of the doctoral thesis: 1. The study of the relationship of lactose hydrolysis process in the formation of galacto-oligosaccharides and lactulose. 2. The combination of β-galactosidase and glucose isomerase increases the sweetness of glucose-galactose syrup. The economic significance of the doctoral thesis: 1. The studies have shown the possibility to obtain syrup that can be used as sugar and sweeteners replacer in the food industry and to produce functional products. 2. A technological solution for hydrolysis of lactose is proposed, comprehensively evaluating the physical properties of lactose, fermentation parameters and whey composition. The doctoral thesis consists of three chapters: Chapter 1 describes the composition of whey and the possibilities of using it. An overview of the chemical and physical properties of lactose, lactose hydrolysis methods, the application of β-galactosidases and the properties of glucose-galactose syrup are provided. Chapter 2 summarises the materials and methods used in the thesis. Chapter 3 provides a summary of the results obtained in the study, the properties of commercial enzymes in different cation concentrations, the stability of enzymes in the gastrointestinal tract model, methods for the determination of lactose, the properties of dehydrated permeates are evaluated. The influence of factors on the hydrolysis of permeates and the profile of the obtained sugars was analysed. Possibilities for lactulose synthesis are considered. Sensory analysis of glucose-galactose syrups and syrups obtained in the two-stage fermentation are given. During the PhD studies the author had an internship at the Dairy Innovation Institute California Polytechnic State University (USA), where the experimental work was done. Internship was provided by the Baltic – American Freedom Foundation (BAFF) and the Council on International Education Exchange (CIEE). The study was partly financed by the LLU programme “Strengthening Research Capacity at the Latvia University of Agriculture” grant (Contract No. 3.2.-10/2017/LLU/27) “The optimization of bioprocesses for lactose recycling products”. The study was partly financed by the doctoral studies grant “Transition to the new doctoral funding model at the Latvia University of Life Sciences and Technologies” (Contract No. 3.2.-10/90). The thesis is written in English, it consists of 111 pages, 32 tables, 41 figures, 3 appendixes, and 233 bibliographic sources.

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Microbial β‐Galactosidases of industrial importance: Computational studies on the effects of point mutations on the lactose hydrolysis reaction

Cited by 11 publications

References 35 publications

Cell permeabilization of Kluyveromyces and Saccharomyces species to obtain potential biocatalysts for lactose hydrolysis

Cell permeabilization of Kluyveromyces and Saccharomyces species to obtain potential biocatalysts for lactose hydrolysis

β-galactosidase GALA fromBacillus circulanswith high transgalactosylation activity

Valorisation of whey for lactose recycling products production = Sūkalu valorizācija laktozes pārstrādes produktu ieguvei

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