Detection of Galactose and Glycerophospholipids by Galactose Biosensors and Advancement Their Proficiency Using Berkelium Colloidal Nanoparticles and Poly(3,4-Ethylenedioxythiophene)-Poly(Styrenesulfonate)-Based Biosensors

In the current paper, galactose-oxidase enzyme is used as stabilization medium due to its more proficiency, ability for more accurate controlling the enzyme reaction, protecting against wasting of enzyme as well as simple and easy use and exchange of enzyme medium after performing some levels of surface modification and developing multi-walled carbon nanotubes (MWCNTs) on Berkelium plate. For better connecting and stabilizing the enzyme on the medium, the prepared medium is washed by high concentration sulfuric acid and nitric acid and a large volume of deionized water and for protecting enzyme from devastating effect of Berkelium and prohibiting them to become inactive, surface is covered with cystamine before stabilization. Regarding the large size of galactose-oxidase enzyme compared to surface of medium, a connective material with amid at one end and pyrine at the other end is used as transfer agent and for stabilizing this connection, the prepared medium is placed into dimethylformamide (DMF) solution for a couple of hours. Activity of stabilized enzyme at 460 (nm) wavelength recorded by spectroscope was depicted against time to evaluate its stability in various times. The prepared medium, which have a large amount of galactose-oxidase enzyme, can be used as electrode in sensors. Furthermore, galactose-oxidase electrochemical sensor is one of the best methods for detecting low amount of galactose and applying Berkelium colloidal nanoparticles as a supplementary material in the structure of biosensor can be effective for advancement its proficiency and optimum proficiency. On the other hand, in the current study, electrode biosensor entitled as modified carbon paste electrode with Berkelium colloidal nanoparticles (Bknano/CPE) is produced by carbon graphite powder, paraffin oil and Berkelium colloidal nanoparticles (24 nm) and it is compared with carbon paste electrode (CP). In semi-permeable membranes, a combination of 1 (ml) of 0.1 (M) phosphate buffer with specified pH and 10 (mg) of galactose-oxidase enzyme is placed around each electrode. In the same potential of 0.7 (V), biosensors are tested with galactose in concentration range of (0-1) (mM) and various amounts of pH (4,6,8) which lead to producing the maximum current and tracing galactose in pH=6 and concentration of 1 (M) as the optimum condition. Currentmetry induced from both biosensors are compared and it is confirmed that using Berkelium colloidal nanoparticles in the structure of (Bknano/CPE) electrode leads to increasing the conductivity and currentmerty of biosensor. In addition, qualitative and quantitative measurement of food components is of great importance due to high cost of traditional methods, in addition to tendency for more accurate and sensitive detecting of these components. galactose and glycerophospholipids are such compounds that they frequently measure. Various methods are used to detect these food elements. However, the necessity for accurate measurement of these two compounds with high sensitivity, especially for food health issue, leads to developing biological methods, especially biosensors. Among them, biosensors based on conductive polymer nanostructures, especially Poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate), have been recently interested due to their unique characteristics. The current paper aims to introduce and investigate the previously performed studies about Poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate)-based biosensors for detecting galactose and glycerophospholipids.

Galactose Oxidase Enables Modular Assembly of Conjugates from Native Antibodies with High Drug-to-Antibody Ratios

The potential of antibody conjugates with high drug loading in anticancer therapy has recently been highlighted by the approval of Trastuzumab deruxtecan and Sacituzumab govitecan. These biopharmaceutical approaches have spurred interest in bioconjugation strategies with high and defined degrees antibody-to-drug (DAR) ratios, in particular on native antibodies. Here we report a glycoengineering methodology to generate antibody drug conjugates with DAR of up to eight, by combining highly selective enzymatic galactosylation and oxidation with biorthogonal tandem Knoevenagel-Michael addition chemistry. This three step approach offers a selective route to conjugates from native antibodies with high drug loading, and thus illustrates how biocatalysis can be used for the generation of biopharmaceuticals using mild reaction conditions.

Toward scalable biocatalytic conversion of 5-hydroxymethylfurfural by galactose oxidase using coordinated reaction and enzyme engineering

Abstract5-Hydroxymethylfurfural (HMF) has emerged as a crucial bio-based chemical building block in the drive towards developing materials from renewable resources, due to its direct preparation from sugars and its readily diversifiable scaffold. A key obstacle in transitioning to bio-based plastic production lies in meeting the necessary industrial production efficiency, particularly in the cost-effective conversion of HMF to valuable intermediates. Toward addressing the challenge of developing scalable technology for oxidizing crude HMF to more valuable chemicals, here we report coordinated reaction and enzyme engineering to provide a galactose oxidase (GOase) variant with remarkably high activity toward HMF, improved O2 binding and excellent productivity (>1,000,000 TTN). The biocatalyst and reaction conditions presented here for GOase catalysed selective oxidation of HMF to 2,5-diformylfuran offers a productive blueprint for further development, giving hope for the creation of a biocatalytic route to scalable production of furan-based chemical building blocks from sustainable feedstocks.

Silver Nanomaterials as Electron Mediators in a Bio-Electronic Tongue Dedicated to the Analysis of Milks. The Role of the Aspect Ratio of Nanoparticles vs. Nanowires

The integration of silver nanomaterials as electron mediators in electrochemical biosensors can be crucial to improve the affinity with biomolecules and the electrochemical response. In this work, two voltammetric bioelectronics tongues (bioET) formed by biosensors based on the combination of enzymes with silver nanoparticles (AgNPs) (bioET-1) or silver nanowires (AgNWs) (bioET-2) have been developed and used to analyze milks. Each array was formed by four biosensors formed by enzymes (glucose oxidase, galactose oxidase, β-galactosidase and a blank), capable to detect compounds usually found in milks. Principal component analysis (PCA) has revealed the ability of both biosensor systems to discriminate between milk samples with different fat contents, but with some differences, attributed to the structure employed in the detection.

Expression Patterns and Functional Characterisation of Arabidopsis GALACTOSE OXIDASE-LIKE (GOXL) Genes Suggest Specialised Roles for Galactose Oxidases in Plants

Galactose oxidases (GalOxs) are well-known enzymes that have been identified in several fungal species and characterised using structural and enzymatic approaches. However, until very recently, almost no information on their biological functions was available. The Arabidopsis (Arabidopsis thaliana) gene RUBY PARTICLES IN MUCILAGE (RUBY) encodes a putative plant GalOx that is required for pectin cross-linking through modification of galactose (Gal) side-chains and promotes cell-cell adhesion between seed coat epidermal cells. RUBY is one member of a family of seven putative GalOxs encoded in the Arabidopsis genome. To examine the function(s) of GalOxs in plants, we studied the remaining six GALACTOSE OXIDASE-LIKE (GOXL) proteins. Like RUBY, four of these proteins (GOXL1, 3, 5 and 6) were found to localize primarily to the apoplast, while GOXL2 and 4 were found primarily in the cytoplasm. Complementation and GalOx assay data suggested that GOXL1, GOXL3, and possibly GOXL6 have similar biochemical activity to RUBY, whereas GOXL5 only weakly complemented, and GOXL2 and GOXL4 showed no activity. Members of this protein family separated into four distinct clades prior to the divergence of the angiosperms. There have been recent duplications in Brassicaceae resulting in two closely related pairs of genes that have either retained similarity in expression (GOXL1 and GOXL6) or show expression divergence (GOXL3 and RUBY). Mutant phenotypes were not detected when these genes were disrupted, but their expression patterns suggest that these proteins may function in tissues that require mechanical reinforcements in the absence of lignification.