Preparative Biochemistry & Biotechnology

Transmembrane serine protease 2 (TMPRSS2) is a membrane-bound protease belonging to the type II transmembrane serine protease (TTSP) family. It is a multidomain protein, including a serine protease domain responsible for its self-activation. The protein has been implicated as an oncogenic transcription factor and for its ability to cleave (prime) the SARS-CoV-2 spike protein. In order to characterize the TMPRSS2 biochemical properties, we expressed the serine protease domain (rTMPRSS2_SP) in Komagataella phaffii using the pPICZαA vector and purified it using immobilized metal affinity (Ni Sepharose™ excel) and size exclusion (Superdex 75) chromatography...

Oleaginous organisms accrue more than twenty percent of their biomass as lipids and hence are promising feedstocks for biodiesel production. In this study, lipid accumulating bacteria were isolated from diesel-contaminated soils and screened with Sudan black B stain. The most oleaginous was done using 16s rRNA gene sequencing. Lipid production was initially optimized based on media, nitrogen source, pH and temperature. Response surface methodology (RSM) was then employed for the enhancement of lipid weight and content...

α-Galactosidase (α-GAL) is a class of hydrolase that releases galactose from galacto-oligosaccharides and synthetic substrates such as pNPG. In this study, the production of α-GAL by Actinoplanes utahensis B1 in submerged fermentation was enhanced by using statistical methods. The effects of temperature, pH, and inoculum percentage on enzyme secretion were optimized using BBD of RSM. The optimized process was scaled up from the shake flask to the laboratory scale (5 L) and to pilot scale (30 L) using KL a based scale-up strategy...

The effect of gradients of elevated glucose and low dissolved oxygen in the addition zone of fed-batch E. coli thermoinduced recombinant high cell density cultures can be evaluated through two-compartment scale-down models. Here, glucose was fed in the inlet of a plug flow bioreactor (PFB) connected to a stirred tank bioreactor (STB). E. coli cells diminished growth from 48.2 ± 2.2 g/L in the stage of RP production if compared to control (STB) with STB-PFB experiments, when residence time inside the PFB was 25 s (34...

Klebsiella oxytoca JCM 1665 was subjected to extracellular rhodanese production using a submerged fermentation technique. The organism was further engineered for higher cyanide tolerance and rhodanese yield using ethylmethanesulfonate as a mutagen. Mutagenesis resulted in an improved mutant with high cyanide tolerance (100 mM) and rhodanese yield (26.7 ± 0.67 U/mL). This yield was 4.34-fold higher than the wild strain (6.15 ± 0.65 U/mL). At temperatures ranging from 30 to 80 °C, the first-order thermal denaturation constant ( Kd ) for free enzyme increases from 0...

Coprinus comatus is an edible mushroom and its fermented product possesses antioxidant activity. In this study, to further enhance the antioxidant activity and improve the reusability of the strain, calcium alginate hydrogel was used as the carrier for embedding and immobilizing Coprinus comatus . The effects of CaCl2 concentration, sodium alginate concentration, microsphere diameter, and the amount of magnetic particle on the antioxidant activity of fermented products were investigated. The results showed that the magnetic immobilized microsphere prepared by 2...

We established an efficient method using high-speed countercurrent chromatography (HSCCC) combined with preparative high-performance liquid chromatography (prep-HPLC) for isolating and purifying Gels emium elegans ( G. elegans ) alkaloids. First, the two-phase solvent system composed of 1% triethylamine aqueous solution/ n -hexane/ethyl acetate/ethanol (volume ratio 4:2:3:2) was employed to separate the crude extract (350 mg) using HSCCC. Subsequently, the mixture that resulted from HSCCC was further separated by Prep-HPLC, resulting in seven pure compounds including: 14-hydroxygelsenicine ( 1 , 12...

The human palate can discern multiple tastes; however, it predominantly perceives five fundamental flavors: sweetness, saltiness, sourness, bitterness, and umami. Sweetness is primarily mediated through the sweet taste receptor, a membrane-bound heterodimeric structure comprising T1R2-T1R3. However, unraveling the structural and mechanistic intricacies of the sweet taste receptor has proven challenging. This study aimed to address this knowledge gap by expressing an extracellular N-terminal domain encompassing the cysteine-rich domain of human hT1R3 (hT1R3-TMD) in Escherichia coli ...

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Synthetic dyes such as azo dyes are significant pollutants in the wastewater released from various textile industries. The low biodegradability and production from synthetic sources with high shelf life make azo dyes a challenging material for degradation. This study used chemically mutated Aspergillus terrus in the laccase production under solid-state fermentation using sugarcane bagasse. Initially, the wild-type strain produced a laccase activity of 4.12 U/mL. Later, the alkaline pretreatment of sugarcane bagasse showed a significant increase in laccase activity by 38...

To create tissue-engineered vascular grafts (TEVGs) in vitro, vascular smooth muscle cells (VSMCs) must function effectively and produce sufficient extracellular matrix (ECM) in a three-dimensional space. In this study, we investigated whether the addition of insulin-transferrin-selenium (ITS), a medium supplement, could enhance TEVG formation. PGA fabric was used as the scaffold, and 1% ITS was added to the medium. After two weeks, the tissues were examined using electron microscopy and staining. The ITS group exhibited a denser structure and increased collagen production...

The removal of hexavalent chromium [Cr (VI)] using non-living cells of Yarrowia lipolytica was investigated. Batch and continuous studies on removal of Cr (VI) achieved 97% and 99% removal from aqueous phase, respectively. The specific uptake values at pH of 2 in batch process were 40.73 ± 1.3 mg/g and 30.09 ± 0.23 mg/g on non-living cells, when 100 and 200 mg/L of metal Cr (VI) concentrations were used. In order to investigate the regulation of Cr (VI) under continuous operation based on reaction volume numerically a new class of feedback controller from structure polynomial was designed...

Traditional Chineae medicine (TCM) is often composed of a variety of natural medicines. Its composition is complex, and many of its components can not be analyzed and identified. The first step in the rational application of TCM is to successfully separate the effective components which is also a great inspiration for the development of new drugs. Among the many separation technologies of TCM, the traditional heating concentration separation technology has high energy consumption and low efficiency. As a new separation technology, membrane separation technology has the characteristics of simple operation, high efficiency, environment-friendly and so on...

In the [Bmim]Cl reaction medium, five different acidic ionic liquids were used as catalysts to study the effects of reaction time, reaction temperature, system water content, catalyst dosage, microwave power, and other factors on cellulose hydrolysis under microwave irradiation. The results showed that in the [Bmim]Cl reaction system, using N-methylpyrrolidone methylsulfonic acid salt as a catalyst, controlling the microwave reaction time at 10 min, reaction temperature at 130 °C, catalyst dosage at 1 g/g (cellulose), water addition at 0...

Since cytoplasmic expression of heterologous proteins with disulfide bonds leads to the formation of inclusion bodies in E. coli , periplasmic production is preferable. The N-terminal signal peptide attached to the secreted protein determines the type of secretory pathway through which the target protein is secreted; Sec, Tat, or SRP. The aim of this study was to design and compare two novel signal peptides for the secretion of recombinant neurturin (as a model) via the Sec and Tat pathways. For this purpose, we aligned the natural signal peptides from E...

An integrated approach involving response surface methodology (RSM) and artificial neural network-ant-colony hybrid optimization (ANN-ACO) was adopted to develop a bioprocess medium to increase the yield of Bacillus cereus neutral protease under submerged fermentation conditions. The ANN-ACO model was comparatively superior (predicted r 2 = 98.5%, mean squared error [MSE] = 0.0353) to RSM model (predicted r 2 = 86.4%, MSE = 23.85) in predictive capability arising from its low performance error...

An efficient method of solid-state fermentation (SSF) is reported for producing bioactive phenolic compounds using soil-isolated fungi. Antioxidant activity using a rapid DPPH (1,1-diphenyl-2-picryl hydrazyl), was employed to screen the 120 fungal isolates from soil. Aspergillus terreus 1, Aspergillus fumigatus, Aspergillus terreus 2, Penicillium citrinum, Aspergillus wentii1, Aspergillus wentii 2, Penicillium expansum and Penicillium granulatum were chosen, concerning their antioxidant activity and total phenolic content...

α-L-rhamnosidase (Rha) is ubiquitous in nature and has high feasibility in the food and biotechnology industries. A green and environmentally friendly method was used to improve the activity of Rha. Here, we show that the effects of ultrasound treatment on the Rha. Ultrasonic treatment at 80 W for 10 min yielded the highest enzyme activity. Treatment increased enzyme activity by 26.3% and half-life by 124 min. Further, treatment increased the catalytic efficiency of Rha and increased the substrate conversion rate by 33...

Food falsification is a pressing issue in today's food industry, with fraudulent substitution of costly ingredients with cheaper alternatives occurring globally. Consequently, developing straightforward and efficient diagnostic systems to detect such fraud is a top priority in scientific research. The aim of the work was to develop a test system and protocol for polymerase chain reaction (PCR) to detect in food products of animal origin the substitution of expensive meat raw materials for by-products of poultry processing...

To investigate the biocatalytic potential of Amazonian actinomycetes for monoterpenes biotransformation. To carry out the present study, eleven actinomycetes of the genus Streptomyces isolated from inga-cipó ( Inga edulis Mart.) rhizospheres were tested for their ability to bioconvert the substrates R -(+)-limonene, S -(-)-limonene, 1 S -(-)-α-pinene, and (-)-β-pinene as sole carbon and energy source. According to gas chromatography-mass spectrometry analysis, three strains, LabMicra B270, LaBMicrA B310, and LaBMicrA B314, were able to biotransform 1 S -(-)-α-pinene after 96 h of growth...