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Valuation of agro-industrial wastes as substrates for heterologous production of ?-galactosidase.

ABSTRACT: BACKGROUND:The recycling of agro-industrial wastes is at present limited by the availability of efficient and low-cost enzyme cocktails. The use of these materials as culture media to produce the enzymes can contribute to the profitability of the recycling process and to the circular economy. The aim of this work is the construction of a recombinant yeast strain efficient to grow in mixed whey (residue of cheese making) and beet molasses (residue of sugar manufacture) as culture medium, and to produce heterologous ?-galactosidase, an enzyme with varied industrial applications and wide market. RESULTS:The gene MEL1, encoding the ?-galactosidase of Saccharomyces cerevisiae, was integrated (four copies) in the LAC4 locus of the Kluyveromyces lactis industrial strain GG799. The constructed recombinant strain produces high levels of extracellular ?-galactosidase under the control of the LAC4 promoter, inducible by lactose and galactose, and the native MEL1 secretion signal peptide. K. lactis produces natively beta-galactosidase and invertase thus metabolizing the sugars of whey and molasses. A culture medium based on whey and molasses was statistically optimized, and then the cultures scaled-up at laboratory level, thus obtaining 19 U/mL of heterologous ?-galactosidase with a productivity of 0.158 U/L h, which is the highest value reported hitherto from a cheap waste-based medium. CONCLUSIONS:A K. lactis recombinant strain was constructed and a sustainable culture medium, based on a mixture of cheese whey and beet molasses, was optimized for high productivity of S. cerevisiae ?-galactosidase, thus contributing to the circular economy by producing a heterologous enzyme from two agro-industrial wastes.

SUBMITTER: Alvarez-Cao ME

PROVIDER: S-EPMC6122717 | biostudies-literature | 2018 Sep

REPOSITORIES: biostudies-literature

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Valuation of agro-industrial wastes as substrates for heterologous production of α-galactosidase.

Álvarez-Cao María-Efigenia ME Rico-Díaz Agustín A Cerdán María-Esperanza ME Becerra Manuel M González-Siso María-Isabel MI

Microbial cell factories 20180903 1

<h4>Background</h4>The recycling of agro-industrial wastes is at present limited by the availability of efficient and low-cost enzyme cocktails. The use of these materials as culture media to produce the enzymes can contribute to the profitability of the recycling process and to the circular economy. The aim of this work is the construction of a recombinant yeast strain efficient to grow in mixed whey (residue of cheese making) and beet molasses (residue of sugar manufacture) as culture medium, ...[more]

PMID: 30176892

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Project description:The microbial conversion of agro-industrial oil wastes into biosurfactants shows promise as a biomass refinery approach. In this study, Bacillus subtilis #309 was applied to produce surfactin using rapeseed and sunflower cakes, the most common oil processing side products in Europe. Studies of the chemical composition of the substrates were performed, to determine the feasibility of oil cakes for surfactin production. Initially, screening of proteolytic and lipolytic activity was performed to establish the capability of B. subtilis #309 for substrate utilization and hence effective surfactin production. B. subtilis #309 showed both proteolytic and lipolytic activity. The process of surfactin production was carefully analyzed by measurement of the surfactin concentration, pH, surface tension (ST) and emulsification index (E24). The maximal surfactin concentration in the sunflower and rapeseed cake medium reached 1.19 ± 0.03 and 1.45 ± 0.09 g/L, respectively. At the same time, a progressive decrease in the surface tension and increase in emulsification activity were observed. The results confirmed the occurrence of various surfactin homologues, while the surfactin C15 was the dominant one. Finally, the analysis of surfactin biological function exhibited antioxidant activity and significant angiotensin-converting enzyme (ACE)-inhibitory activity. The half-maximal inhibitory concentration (IC50) value for ACE inhibition was found to be 0.62 mg/mL for surfactin. Molecular docking of the surfactin molecule to the ACE domains confirmed its inhibitory activity against ACE. Several interactions, such as hydrophobic terms, hydrogen bonds and van der Waals interactions, were involved in the complex stabilization. To the best of our knowledge, this is the first report describing the effect of a lipopeptide biosurfactant, surfactin, produced by B. subtilis for multifunctional properties in vitro, namely the ACE-inhibitory activity and the antioxidant properties, using different assays, such as 2,2-azinobis (3-ethyl-benzothiazoline-6-sulfonic acid (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP). Thus, the ACE-inhibitory lipopeptide biosurfactant shows promise to be used as a natural antihypertensive agent.

Dataset Information

Valuation of agro-industrial wastes as substrates for heterologous production of ?-galactosidase.

Publications

Valuation of agro-industrial wastes as substrates for heterologous production of α-galactosidase.

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