Enzyme and Microbial Technology

Prunin of desirable bioactivity and bioavailability can be transformed from plant-derived naringin by the key enzyme α-L-rhamnosidase. However, the production was limited by unsatisfactory properties of α-L-rhamnosidase such as thermostability and organic solvent tolerance. In this study, biochemical characteristics, and hydrolysis capacity of a novel α-L-rhamnosidase from Spirochaeta thermophila (St-Rha) were investigated, which was the first characterized α-L-rhamnosidase for Spirochaeta genus...

The solvent-free esterification of the free fatty acids (FFAs) obtained by the hydrolysis of castor oil (a non-edible vegetable oil) with 2-ethyl-1-hexanol (a branched fatty alcohol) was catalyzed by different free lipases. Eversa Transform 2.0 (ETL) features surpassed most commercial lipases. Some process parameters were optimized by the Taguchi method (L16'). As a result, a conversion over 95% of the FFAs of castor oil into esters with lubricants properties was achieved under optimized reaction conditions (15 wt% of biocatalyst content, 1:4 molar ratio (FFAs/alcohol), 30 °C, 180 rpm, 96 h)...

Alginate lyases with unique characteristics for degrading alginate into size-defined oligosaccharide fractions, were considered as the potential agents for disrupting Pseudomonas aeruginosa biofilms. In our study, a novel endolytic PL-7 alginate lyase, named AlyG2, was cloned and expressed through Escherichia coli. This enzyme exhibited excellent properties: it maintained more than 85% activity at low temperatures of 4 °C and high temperatures of 70 °C. After 1 h of incubation at 4 °C, it still retained over 95% activity, demonstrating the ability to withstand low temperature...

Due to the extreme living conditions, extremophiles have unique characteristics in morphology, structure, physiology, biochemistry, molecular evolution mechanism and so on. Extremophiles have superior growth and synthesis capabilities under harsh conditions compared to conventional microorganisms, allowing for unsterilized fermentation processes and thus better performance in low-cost production. In recent years, due to the development and optimization of molecular biology, synthetic biology and fermentation technology, the identification and screening technology of extremophiles has been greatly improved...

The chiral amine (R)-2-(1-aminoethyl)-4-fluorophenol has attracted increasing attentions in recent years in the field of pharmaceuticals because of its important use as a building block in the synthesis of novel anti-tumor drugs targeting tropomyosin receptor kinases. In the present study, a ω-transaminase (ωTA) library consisting of 21 (R)-enantioselective enzymes was constructed and screened for the asymmetric biosynthesis of (R)-2-(1-aminoethyl)-4-fluorophenol from its prochiral ketone. Using (R)-α-methylbenzylamine, D-alanine, or isopropylamine as amino donor, 18 ωTAs were identified with target activity and the enzyme AbTA, which was originally identified from Arthrobacter sp...

Commercially available cellulase cocktails frequently demonstrate high efficiency in hydrolyzing easily digestible pretreated biomass, which often lacks hemicellulose and/or lignin fractions. However, the challenge arises with enzymatic hydrolysis of mildly pretreated lignocellulosic biomasses, which contain cellulose, hemicellulose and lignin in high proportions. This study aimed to address this question by evaluating the supplementation of a commercial cellulolytic cocktail with accessory hemicellulases and two additives (H2 O2 and Tween® 80)...

Pectinase plays a crucial role in ramie degumming. A gene encoding a putative pectate lyase from Bacillus sp. strain B58-2 was cloned and heterologously expressed in Escherichia coli. The amplified gene BvelPL1 encoded a mature protein of 400 amino acids. BvelPL1 shared the highest amino acid sequence identity (78.75%) with the enzymatically characterized pectate lyase Pel from Bacillus subtilis strain RCK (GenBank: AFH66771.1). The purified recombinant enzyme rBvelPL1-Ec exhibited a maximum specific activity of 2433...

L-theanine is an amino acid with a unique flavor and many therapeutic effects. Its enzymatic synthesis has been actively studied and γ-Glutamylmethylamide synthetase (GMAS) is one of the promising enzymes in the biological synthesis of theanine. However, the theanine biosynthetic pathway with GMAS is highly ATP-dependent and the supply of external ATP was needed to achieve high concentration of theanine production. As a result, this study aimed to investigate polyphosphate kinase 2 (PPK2) as ATP regeneration system with hexametaphosphate...

The robustness of microbial consortia isolated from compost habitat encompasses the complementary metabolism that aids in consolidated bioprocessing (CBP) of lignocellulosic biomass (LCB) by division of labor across the symbionts. Composting of organic waste is deemed to be an efficient way of carbon recycling, where the syntrophic microbial population exerts a concerted action of lignin and polysaccharide (hemicellulose and cellulose) component of plant biomass. The potential of this interrelated microorganism could be enhanced through adaptive laboratory evolution (ALE) with LCB for its desired functional capabilities...

Lignin depolymerization, which enables the breakdown of a complex and heterogeneous aromatic polymer into relatively uniform derivatives, serves as a critical process in valorization of lignin. Enzymatic lignin depolymerization has become a promising biological strategy to overcome the heterogeneity of lignin, due to its mild reaction conditions and high specificity. However, the low solubility of lignin compounds in aqueous environments prevents efficient lignin depolymerization by lignin-degrading enzymes...

Organohalides are recalcitrant, toxic environmental pollutants. Reductive dehalogenase enzymes (RDases) found in organohalide respiring bacteria (OHRB) utilise organohalides as electron acceptors for cellular energy and growth, producing lesser-halogenated compounds. Consequently, microbial reductive dehalogenation via organohalide respiration represents a promising solution for clean-up of organohalide pollutants. Dehalobacter sp. UNSWDHB is an OHRB capable of respiring highly toxic chloroform (CF) and converting it to dichloromethane (DCM)...

The split-GFP system is a versatile tool with numerous applications, but it has been underutilized for the labeling of heterologous surface-displayed proteins. By inserting the 16 amino acid sequence of the GFP11-tag between a protein of interest and an autotransporter protein, it is possible to present a protein at the outer membrane of gram-negative bacteria and to fluorescently label it by complementation with externally added GFP1-10. The labeled cells could be clearly discerned from cells without the protein of interest using flow cytometry and the insertion of the GFP11-tag caused no significant alteration of the catalytic activity for the tested model enzyme CsBglA...

Cyanide is widely utilized in the extraction of precious metal extraction even though it has been deemed as the most toxic compound. Fusarium oxysporum has been shown to degrade cyanide through the activity of the Nitrilase enzyme. In this study, the coding sequence of nitrilase gene from F. oxysporum genomic DNA was optimized for cloning and expression in E. coli. The pUC57 containing synthetic optimized nitrilase gene was transferred into E. coli DH5α strain. This nitrilase gene was sub-cloned into pET26b (+) expression vector containing an in-built His-tag at the C-terminal end to facilitate its purification...

Indigo, an economically important dye, could be biosynthesized from indole by catalysis of the styrene monooxygenase StyAB. To enhance indigo biosynthesis, the styAB gene and its transcription regulator gene styS/styR in styrene catabolism were cloned from Pseudomonas putida and coexpressed in Escherichia coli. The presence of the intact regulator gene styS/styR dramatically increased the transcriptional levels of styA and styB by approximately 120-fold in the recombinant strain SRAB2 with coexpression of styS/styR and styAB compared to the control strain ABST with solo expression of styAB...

Sepsis is a serious disease caused by an impaired host immune response to infection, resulting in organ dysfunction, tissue damage and is responsible for high in-hospital mortality (approximately 20%). Recently, WHO documented sepsis as a global health priority. Nevertheless, there is still no effective and specific therapy for clinically detecting sepsis. Nanomaterial-based approaches have appeared as promising tools for identifying bacterial infections. In this review, recent biosensors are introduced and summarized as nanomaterial-based platforms for sepsis management and severe complications...

Xanthine is derived from hypoxanthine by xanthine oxidase (XOD), a flavoprotein containing molybdenum and non-haem iron, sulfur and from guanine by guanine deaminase enzyme. Xanthine is oxidized into uric acid by XOD. Xanthine is used as an indicator of fish freshness, based on the reactions in which ATP is degraded into xanthine and its quantity increases with time of fish death. Fresh fish meat is required in food industry for making high quality items. The determination of xanthine in biological fluids is also used in diagnosing and curing many diseases like renal failure, gout, xanthinuria, hyperuricemia...

The enzymes of the mevalonate pathway need to be improved to achieve high yields of isoprenoids in the yeast Saccharomyces cerevisiae. The red yeast Rhodosporidium toruloides produces high levels of carotenoids and may have evolved to carry a naturally high flux of isoprenoids. Enzymes from such yeasts are likely to be promising candidates for improvement. Towards this end, we have systematically investigated the various enzymes of the mevalonate pathway of R. toruloides and custom synthesized, expressed, and evaluated six key enzymes in S...

Immobilized enzymes exhibit favorable advantages in biocatalysis, such as high operation stability, feasible reusability, and improved organic solvents tolerance. Herein, an immobilized ω-amine transaminase AtATA@MWCNTs-NH2 is successfully prepared using amino modified multi-walled carbon nanotubes as carrier and glutaraldehyde as crosslinker. Under the optimum immobilization conditions, the activity recovery is 78.7%. Compared with purified enzyme AtATA, AtATA@MWCNTs-NH2 possesses superior stability, even in harsh conditions (e...

γ-Aminobutyric acid (GABA) has been widely used in the food, feed, pharmaceutical, and chemical industry fields. Previously, we developed a whole-cell catalyst capable of converting L-glutamate (L-Glu) into GABA by overexpressing the glutamate decarboxylase gene (gadz11) from Bacillus sp. Z11 in Escherichia coli BL21(DE3). However, to enhance cell permeability, a freeze-thaw treatment is required, and to enhance GADZ11 activity, pyridoxal 5'-phosphate (PLP) must be added to the reaction system. The aim of this study is to provide a more efficient approach for GABA production by engineering the recombinant E...

To understand the influence of family 3 Carbohydrate Binding Module (hereafter CBM3), single (GH5 cellulase; CelB, CelBΔCBM), bi-chimeric [GH26 endo-mannanase (ManB-1601) and GH11 endo-xylanase (XynB); ManB-XynB [1], ManB-XynB-CBM] and tri-chimeric [ManB-XynB-CelB [1], ManB-XynB-CelBΔCBM] enzyme variants (fused or deleted of CBM) were produced and purified to homogeneity. CBM3 did not alter the pH and temperature optima of bi- and tri-chimeric enzymes but improved the pH and temperature stability of ManB in CBM variants of bi-/tri-chimeric enzymes...