Thermal stability and energy of deactivation of immobilized cell wall invertase in natural and synthetic hydrogel polymers

[https://doi.org/10.1111/febs.12340

Thermal stability and energy of deactivation of immobilized cell wall invertase in natural and synthetic hydrogel polymers

[https://doi.org/10.1111/febs.12340

Immobilization of cell wall invertase in a polyacrylamide hydrogel for invert sugar production

Cell wall invertase (CWI) from Saccharomyces cerevisiae was immobilized in polyacrylamide hydrogels. The aim of the development of a new biocatalyst was to obtain an improved enzyme for invert sugar production. The monomer concentration and enzyme amount in the immobilizate were optimized, and the obtained biocatalyst had an enzyme activity of 138 +/- 6 IU g(-1). The pH and temperature optima were 4.0 and 70 degrees C, respectively. The stability of immobilized enzyme was determined at several temperatures in the absence of substrate and the half-life obtained at 50 degrees C was 81 days, at 60 degrees C, 128 min and at 70 degrees C, 1.24 min. The biocatalyst was tested at low pH values, 3.0 and 3.5 were tested, and showed great stability. The KM values were 34.1 +/- 1.7 and 126.2 +/- 6.3 mM for free and immobilized CWI, respectively. The activation energies were 37.7 and 23.0 kJ mol(-1) for free and immobilized CWI, respectively. Cell wall invertase immobilized in polyacrylamide hydrogel (CWI-PAA) was tested for the production of high concentrated invert sugar in a batch and a packed bed reactor. After five days of continuous process, the quality and characteristics of the produced invert sugar remained unchanged

Thermal stability and energy of deactivation of immobilized cell wall invertase in natural and synthetic hydrogel polymers

[https://doi.org/10.1111/febs.12340

Antioxidant potential and protein interactions of four tea plant extracts

Polyphenols are a large group of natural organic compounds mainly found in plants with diverse protective and metabolic functions. Phenolic compounds are well known for their antioxidant properties and the name “antioxidant” is mostly associated with them1. It’s also known that phenolic compounds, especially tannins, interact with proteins in various significant and distinct ways. This study was focused on examining these characteristics on aqueous extracts of tea plants local to Serbia (freshly picked Satureja montana, Mentha spicata, Salvia officinalis and Matricaria chamomilla), which are known to have various amounts and types of phenolic compounds. We examined total concentration of phenolic compounds, tannins, flavonoids, antioxidant activity and interactions with bovine serum albumin and whey protein. It was concluded that S. montana extract had the highest concentration of polyphenols and tannins. Flavonoid concentration was measured using the aluminium-chloride method and it was concluded that S. officinalis had the highest flavonoid content. Antioxidant activities were measured using DPPH, FRAP and ABTS, which are Single Electron Transfer (SET) mechanism-based antioxidant tests and ORAC which is a Hydrogen Atom Transfer mechanism-based antioxidant test. It was concluded that SET mechanism-based antioxidant activities correspond to the total concentration of polyphenols and tannins, which meant that S. montana extract had the highest antioxidant activity. S. officinalis extract exhibited the highest antioxidant activity measured by the ORAC assay, which corresponded to the highest Flavonoid concentration. Interactions between plant extracts and bovine serum albumin were measured via spectrophotometric and spectrofluorimetric titrations. It was concluded that S. montana exhibited most pronounced interactions with the protein. Nature of such interactions is still unknown but using SDS-PAGE it was concluded that proteins exhibit significant structural changes after interacting with plant extracts

Production of polyglutamic acid by Bacillus sp: Strains selection, optimization, batch fermentation and characterization

Polyglutamic acid (PGA) is an anionic, non-toxic natural polymer that consists of D- and L-glutamic acid. Glutamic acids that makeup PGA create bonds between α-amino and γ-carboxyl groups. PGA can be found on the surface of many different bacteria, usually as a part of their capsule. Bacteria use polyglutamic acid to survive adverse environmental conditions. PGA has found multiple potential applications as a thickener, drug carrier, biological adhesive, heavy metal and basic dye adsorber, etc. Its biodegradability is especially useful in the fields of food, cosmetics, medicine and water treatment1. The aim of this study was to find the best polyglutamic acid producer from the selection of 50 different Bacillus sp strains originating from Serbia, as well as the optimal medium composition. It was discovered that the best PGA producing strain was 17B and it was selected for further fermentation medium optimization. Optimization was carried out using Design of Experiment, specifically Box Behnken design. Results were analyzed using response surface methodology. PGA that was produced during this process was analyzed using SDS PAGE and basic dye adsorption was attempted as well. The best PGA producer, Bacillus sp strain 17B, was used for PGA production in batch fermenter 2. PGA isolated from the fermentation broth was purified using gel filtration and further characterized using SDS PAGE, FTIR spectroscopy and direct-infusion MS

Guanylate Cyclase-Activating Protein-2 Undergoes Structural Changes upon Binding to Detergent Micelles and Bicelles

GCAPs are neuronal Ca2+-sensors playing a central role in light adaptation. GCAPs are N-terminally myristoylated. membrane-associated proteins. Although, the myristoylation of GCAPs plays an important role in light adaptation its structural and physiological roles are not yet clearly understood. The crystal-structure of GCAP-1 shows the myristoyl moiety inside the hydrophobic core of the protein, stabilizing the protein structure; but H-2-solid-state NMR investigations on the deuterated myristoyl moiety of GCAP-2 in the presence of liposomes showed that it is inserted into the lipid bilayer. In this study, we address the question of the localization of the myristoyl group of Ca2+-bound GCAP-2, and the influence of CHAPS-, DPC-micelles and DMPC/DHPC-bicelles on the structure, and on the localization of the myristoyl group, of GCAP-2 by solution-state NMR. We also carried out the backbone assignment. Characteristic chemical shift differences have been observed between the myristoylated and the non-myristoylated forms of the protein. Our results support the view that in the absence of membrane forming substances the myristoyl moiety is buried inside a hydrophobic pocket of GCAP-2 similar to the crystal structure of GCAP-1. Addition of CHAPS-micelles and DMPC/DHPC-bicelles cause specific structural changes localized in and around the myristoyl binding pocket. We interpret these changes as an indication for the extrusion of the myristoyl moiety from its binding pocket and its insertion into the hydrophobic interior of the membrane mimic. On the basis of the backbone chemical shifts, we propose a structural model of myristoylated GCAP-2 in the presence of Ca2+ and membrane mimetics

Diffusion screening method for estimation potential fungal producers of xylanase responsible for xylooligosaccharides production

Wild-type microorganisms from the environment represent a wide source of potential enzyme producers. In order to determine whether an isolated microorganism produces an enzyme of interest, various screen tests have been developed. A new screening method for detection of endo and exo-xylanase activity including short time growth of fungal strains on a minimal medium containing xylan (inducible substrate) as a carbon source is developed and used for testing 58 fungal isolates from genus Aspergillus. The test is based on the diffusion of samples (fermentation extracts) in polyacrylamide gel incorporated by xylan. Endoxylanase activity is detected as enlightenment in the gel after staining of xylan with Congo Red. Exoxylanase activity was visualized as a precipitate after staining of reduction oligosaccharide ends with NBT. Selected isolate A. tubigensis was grown on SSF where corn cob served as an inducible substrate. In order to examine the influence of nitrogen sources on endoxylanase production and fungal growth, two sources (peptone and urea) were varied in 3 concentrations (1, 5 and 10 g/L). There were statistically significant differences in the obtained activities. The increase in activity compared to the screening medium was ~250 times. The obtained enzymes with high specific activity were further used for the production of xylooligosaccharides in high yield which showed that the selection of strain A. tubingensis was good

In situ production of xylooligosaccharides by Aspergillus tubingensis from corn cob

Xylooligosaccharides (XOS) are prebiotic, functional food ingredients, with biological benefits such as immunomodulatory and anti-inflammatory properties, anticancer and antioxidant activity. Usually, XOS are produced from xylan by the combination of pre-treatment, enzymatic, chemical and/or auto-hydrolytic methods. Fungal xylanases are the most suitable for XOS production. Xylan rich agro-industrial wastes (corn cob is one of them) are used as a substrate for fungal xylanase production and as start material for xylan extraction. In situ XOS production by fungal growth on xylan rich medium represents an attractive and advantageous approach but insufficient described till now. This method has many advantages over the others because bypasses the extraction and purification of xylan and enzymes, it is environmentally friendly, low cost and time-consuming. This study demonstrated A. tubingensis FAT35 has a great capacity for the synthesis of XOS using corn cob as a substrate in solid state fermentation (SSF). Obtained XOS, during the fungal growth, were characterized by TLC and HPLC. Significant antioxidant potential was shown by antioxidant tests (ORAC, DPPH and FRAP). The obtained XOS are suitable to be a functional food additive and are obtained in the simplest way that is both environmentally and economically suitable

The Supplementary data for: "Fungal oxidative and hydrolyzing enzymes as designers in the biological production of dietary fibers from triticale"

FTIR spectra of IDFT treatment with fungal enzymatic complex.The Supplementary data for: Margetić, A., Stojanović, S., Ristović, M., & Vujčić, Z. M. (2021). Fungal oxidative and hydrolyzing enzymes as designers in the biological production of dietary fibers from triticale. LWT - Food Science and Technology, Elsevier., 145, 111291. [https://doi.org/10.1016/j.lwt.2021.111291]The published version of the article: [https://cer.ihtm.bg.ac.rs/handle/123456789/4540