Immobilization of the glucose isomerase from Caldicoprobacter algeriensis on Sepabeads EC-HA and its efficient application in continuous High Fructose Syrup production using packed bed reactor

[EN] The glucose isomerase GICA from Caldicoprobacter algeriensis was immobilized by ionic adsorption on polymethacrylate carriers (Sepabeads EC-EA and EC-HA) or covalent attachment to glyoxal agarose. The Sepabeads EC-HA yielded the highest recovery of activity (89%). The optimum temperature and pH of immobilized GICA were 90 °C and 7.0, respectively, similar to the corresponding values of free enzyme. Nevertheless, the adsorbed enzyme displayed higher relative activity at acidic pH, greater thermostability, and better storage stability, compared to the free form. Moreover, the immobilized enzyme showed an excellent operational stability, in 15 successive 3 h reaction cycles at 85 °C under a batch reactor, preserving 83% of its initial activity. Interestingly, a continuous process for High Fructose Syrup (HFS) production was established with the adsorbed GICA using a packed bed reactor during eleven days at 70 °C. HPAEC-PAD analysis showed a maximum bioconversion rate of 49% after 48 h of operation.This work was supported by the Tunisian Ministry of Higher Education and Scientific Research and Technology (contract program LBMIE-CBS, code: LR15CBS06), University of Sfax (doctoral school of FSS) and the Spanish Ministry of Economy and Competitiveness (Grant BIO2016-76601-C3-1-R)

A three-step process for the bioconversion of whey permeate into a glucose-free D-tagatose syrup

We have developed a sustainable three-stage process for the revaluation of cheese whey permeate into D-tagatose, a rare sugar with functional properties used as sweetener. The experimental conditions (pH, temperature, cofactors, etc.) for each step were independently optimized. In the first step, concentrated whey containing 180–200 g/L of lactose was fully hydrolyzed by β-galactosidase from Bifidobacterium bifidum (Saphera®) in 3 h at 45 °C. Secondly, glucose was selectively removed by treatment with Pichia pastoris cells for 3 h at 30 °C. The best results were obtained with 350 mg of cells (previously grown for 16 h) per mL of solution. Finally, L-arabinose isomerase US100 from Bacillus stearothermophilus was employed to isomerize D-galactose into D-tagatose at pH 7.5 and 65 °C, in presence of 0.5 mM MnSO4. After 7 h, the concentration of D-tagatose was approximately 30 g/L (33.3% yield, referred to the initial D-galactose present in whey). The proposed integrated process takes place under mild conditions (neutral pH, moderate temperatures) in a short time (13 h), yielding a glucose-free syrup containing D-tagatose and galactose in a ratio 1:2 (w/w).This work was supported by grants from the Spanish Ministry of Economy and Competitiveness (Grants BIO2016-76601-C3-1-R and C3-2-R) and Fundación Ramón Areces (XIX Call of Research Grants in Life and Material Sciences). We thank Fundación Ramón Areces for an institutional grant to the Center of Molecular Biology Severo Ochoa. F.V. Cervantes thanks CONACYT (Mexico) for her Ph.D. fellowship (Ref. 440242)

Genome sequence and Carbohydrate Active Enzymes (CAZymes) repertoire of the thermophilic Caldicoprobacter algeriensis TH7C1T

11 p.-3 fig.-3 tab.Background: Omics approaches are widely applied in the field of biology for the discovery of potential CAZymes including whole genome sequencing. The aim of this study was to identify protein encoding genes including CAZymes in order to understand glycans-degrading machinery in the thermophilic Caldicoprobacter algeriensis TH7C1T strain.Results: Caldicoprobacter algeriensis TH7C1T is a thermophilic anaerobic bacterium belonging to the Firmicutes phylum, which grows between the temperatures of 55 °C and 75 °C. Next generation sequencing using Illumina technology was performed on the C. algeriensis strain resulting in 45 contigs with an average GC content of 44.9% and a total length of 2,535,023 bp. Genome annotation reveals 2425 protein-coding genes with 97 ORFs coding CAZymes. Many glycoside hydrolases, carbohydrate esterases and glycosyltransferases genes were found linked to genes encoding oligosaccharide transporters and transcriptional regulators; suggesting that CAZyme encoding genes are organized in clusters involved in polysaccharides degradation and transport. In depth analysis of CAZomes content in C. algeriensis genome unveiled 33 CAZyme gene clusters uncovering new enzyme combinations targeting specific substrates.Conclusions: This study is the first targeting CAZymes repertoire of C. algeriensis, it provides insight to the high potential of identified enzymes for plant biomass degradation and their biotechnological applications.This work was supported by the Tunisian Ministry of Higher Education and Scientific Research (contract program LBMIE-CBS, code: LR15CBS06) and the Algerian-Tunisian R&I Cooperation for the Mixed Laboratories of Scientific Excellence 2021–2024 (Hydro-BIOTECH, code LABEX/TN/DZ/21/01). The high throughput sequencing, assembly and annotation work was supported by the IBISBA1.0 H2020 project 730976 (https://www.ibisba.eu/) at its CSIC Cell Factory node.Peer reviewe