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Secretory Expression of an Alkaline Alginate Lyase With Heat Recovery Property in Yarrowia lipolytica.

ABSTRACT: Alginate lyase possesses wide application prospects for the degradation of brown algae and preparation of alginate oligosaccharides, and its degradation products display a variety of biological activities. Although many enzymes of this type have been reported, alginate lyases with unique properties are still relatively rare. In the present work, an alginate lyase abbreviated as Alyw203 has been cloned from Vibrio sp. W2 and expressed in food-grade Yarrowia lipolytica. The Alyw203 gene consists of an open reading frame (ORF) of 1,566 bp containing 521 amino acids, of which the first 17 amino acids are considered signal peptides, corresponding to secretory features. The peak activity of the current enzyme appears at 45°C with a molecular weight of approximately 57.0 kDa. Interestingly, Alyw203 exhibits unique heat recovery performance, returning above 90% of its initial activity in the subsequent incubation for 20 min at 10°C, which is conducive to the recovery of current enzymes at low-temperature conditions. Meanwhile, the highest activity is obtained under alkaline conditions of pH 10.0, showing outstanding pH stability. Additionally, as an alginate lyase independent of NaCl and resistant to metal ions, Alyw203 is highly active in various ionic environments. Moreover, the hydrolyzates of present enzymes are mainly concentrated in the oligosaccharides of DP1-DP2, displaying perfect product specificity. The alkali suitability, heat recovery performance, and high oligosaccharide yield of Alyw203 make it a potential candidate for industrial production of the monosaccharide and disaccharide.

SUBMITTER: Liu L

PROVIDER: S-EPMC8381381 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundRecent trends in bioprocessing have underlined the significance of lignocellulosic biomass conversions for biofuel production. These conversions demand at least 90% energy upgradation of cellulosic sugars to generate renewable drop-in biofuel precursors (Heff/C ~ 2). Chemical methods fail to achieve this without substantial loss of carbon; whereas, oleaginous biological systems propose a greener upgradation route by producing oil from sugars with 30% theoretical yields. However, these oleaginous systems cannot compete with the commercial volumes of vegetable oils in terms of overall oil yields and productivities. One of the significant challenges in the commercial exploitation of these microbial oils lies in the inefficient recovery of the produced oil. This issue has been addressed using highly selective oil capturing agents (OCA), which allow a concomitant microbial oil production and in situ oil recovery process.ResultsAdsorbent-based oil capturing agents were employed for simultaneous in situ oil recovery in the fermentative production broths. Yarrowia lipolytica, a model oleaginous yeast, was milked incessantly for oil production over 380 h in a media comprising of glucose as a sole carbon and nutrient source. This was achieved by continuous online capture of extracellular oil from the aqueous media and also the cell surface, by fluidizing the fermentation broth over an adsorbent bed of oil capturing agents (OCA). A consistent oil yield of 0.33 g per g of glucose consumed, corresponding to theoretical oil yield over glucose, was achieved using this approach. While the incorporation of the OCA increased the oil content up to 89% with complete substrate consumptions, it also caused an overall process integration.ConclusionThe nondisruptive oil capture mediated by an OCA helped in accomplishing a trade-off between microbial oil production and its recovery. This strategy helped in realizing theoretically efficient sugar-to-oil bioconversions in a continuous production process. The process, therefore, endorses a sustainable production of molecular drop-in equivalents through oleaginous yeasts, representing as an absolute microbial oil factory.

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Secretory Expression of an Alkaline Alginate Lyase With Heat Recovery Property in Yarrowia lipolytica.

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