Temperature-dependent mycotoxins production investigation in Alternaria infected cherry by ultra-high performance liquid chromatography and Orbitrap high resolution mass spectrometry.

For temperature-dependent Alternaria mycotoxins production analysis, cherry samples were inoculated with Alternaria sp. and incubated at two different temperatures (4 °C and 25 °C). Six Alternaria mycotoxins, including altenuene (ALT), alternariol monomethyl ether (AME), alternariol (AOH), altertoxin-I (ATX-I), tenuazonic acid (TeA), and tentoxin (TEN), in cherries were detected with integrated visible data-processing tools.

[Expression of a SARS-CoV-2 neutralizing nanobody in ].

Nanobodies derived from camelid single-chain antibodies have the advantages of being small, simple, highly soluble and stable. Nanobodies can be administered by inhalation and therefore is potentially valuable for the prevention and control of respiratory viruses. is a food-grade protein expression host with a cellulase production capacity of up to 80 g/L, which can be employed for low-cost production of therapeutic proteins.

Streamlined Subclass-Specific Absolute Quantification of Serum IgG Glycopeptides Using Synthetic Isotope-Labeled Standards.

Absolute glycoproteomics quantification has drawn tremendous attention owing to its prospects in biomarker discovery and clinical implementation but is impeded by a general lack of suitable heavy isotope-labeled glycopeptide standards. In this study, we devised a facile chemoenzymatic strategy to synthesize a total of 36 human IgG glycopeptides attached with well-defined glycoforms, including 15 isotope-labeled ones with a mass increment of 6 Da to their native counterparts. Spiking of these standards into human sera enabled simplified, robust, and precise absolute quantification of IgG glycopeptides in a subclass-specific fashion.

Penicillium oxalicum putative methyltransferase Mtr23B has similarities and differences with LaeA in regulating conidium development and glycoside hydrolase gene expression.

Putative methyltranferase LaeA and LaeA-like proteins, which are conserved in many filamentous fungi, regulate the sporogenesis and biosynthesis of secondary metabolites. In this study, we reported the biological function of a LaeA-like methyltransferase, Penicillium oxalicum Mtr23B, which contains a methyltransf_23 domain and an S-adenosylmethionine binding domain, in controlling spore pigment formation and in the expression of secondary metabolic gene cluster and glycoside hydrolase genes. Additionally, we compared Mtr23B and LaeA, and determined their similarities and differences in terms of their roles in regulating the above biological processes.

Comparative glycosylation mapping of plasma-derived and recombinant human factor VIII.

Human coagulation factor VIII (FVIII) is a key co-factor in the clotting cascade, the deficiency of which leads to Hemophilia A. Human plasma-derived (pdFVIII) and recombinant FVIII (rFVIII) had been used as effective products to prevent and treat bleeding episodes. Both FVIII products share identical amino acid sequences and appear to be equivalent as of clinical efficiency.

Glucose oxidase and polydopamine functionalized iron oxide nanoparticles: combination of the photothermal effect and reactive oxygen species generation for dual-modality selective cancer therapy.

Cancer cells possess some inherent characteristics, such as glucose-dependence and intolerance to heat and exogenous reactive oxygen species (ROS). In this study, a strategy has been developed to target these vulnerable weaknesses of cancer cells using glucose oxidase (GOx) and polydopamine (PDA) functionalized iron oxide nanoparticles (FeO@PDA/GOx NPs). PDA is first deposited on the surfaces of iron oxide NPs through self-polymerization, and then GOx is covalently linked with PDA upon mixing the enzyme and FeO@PDA under alkaline conditions.

Enzymatic rhamnosylation of anticancer drugs by an α-L-rhamnosidase from Alternaria sp. L1 for cancer-targeting and enzyme-activated prodrug therapy.

The synthesis of rhamnosylated compounds has gained great importance since these compounds have potential therapeutic applications. The enzymatic approaches for glycosylation of bioactive molecules have been well developed; however, the enzymatic rhamnosylation has been largely hindered by lacking of the glycosyl donor for rhamnosyltransferases. Here, we employed an α-L-rhamnosidase from Alternaria sp.

Three glycoside hydrolase family 12 enzymes display diversity in substrate specificities and synergistic action between each other.

PoCel12A, PoCel12B, and PoCel12C are genes that encode glycoside hydrolase family 12 (GH12) enzymes in Penicillium oxalicum. PoCel12A and PoCel12B are typical GH12 enzymes that belong to fungal subfamilies 12-1 and 12-2, respectively. PoCel12C contains a low-complexity region (LCR) domain, which is not found in PoCel12A or PoCel12B and independent of fungal subfamily 12-1 or 12-2.

Identification of Key Components for the Optimization of Cellulase Mixtures Using a Proteomic Strategy.

Efficient degradation of complex lignocellulosic materials requires the synergistic action of different types of enzymes. Characterizing the compositions of lignocellulolytic enzyme mixtures could provide comprehensive understandings about the enzymatic degradation of lignocelluloses. In this chapter, we present a proteomic strategy for the analysis of enzyme mixtures produced by lignocellulolytic fungi.

Rational designed mutagenesis of levansucrase from Bacillus licheniformis 8-37-0-1 for product specificity study.

Levansucrases, which belong to the glycoside hydrolase family 68 (GH68), synthesize β (2-6)-linked fructan levan with sucrose as substrate. We described the use of a levansucrase (Bl_SacB) from Bacillus licheniformis 8-37-0-1 for catalysis of fructosyl transfer to obtain high levan yield previously. In the present study, six variants (Y246A, N251A, K372A, R369A, R369S, and R369K) were constructed through sequence alignment and structural analysis to explore the synthesis mechanism of Bl_SacB.

A general strategy for the synthesis of homogeneous hyaluronan conjugates and their biological applications.

Here, we developed a general strategy for synthesizing homogeneous HA conjugates, and generated homogeneous HA-pNP, HA-biotin, and HA-oroxylin conjugates to investigate the relationships between HA chain length and its diverse biological functions.

Diethylaminoethyl Sepharose (DEAE-Sepharose) microcolumn for enrichment of glycopeptides.

N-Glycosylation is one of the most prevalent protein post-translational modifications and is involved in many biological processes, such as protein folding, cellular communications, and signaling. Alteration of N-glycosylation is closely related to the pathogenesis of diseases. Thus, the investigation of protein N-glycosylation is crucial for the diagnosis and treatment of disease.

An OGA-Resistant Probe Allows Specific Visualization and Accurate Identification of O-GlcNAc-Modified Proteins in Cells.

O-linked β-N-acetyl-glucosamine (O-GlcNAc) is an essential and ubiquitous post-translational modification present in nucleic and cytoplasmic proteins of multicellular eukaryotes. The metabolic chemical probes such as GlcNAc or GalNAc analogues bearing ketone or azide handles, in conjunction with bioorthogonal reactions, provide a powerful approach for detecting and identifying this modification. However, these chemical probes either enter multiple glycosylation pathways or have low labeling efficiency.

Chemoenzymatic synthesis of the bacterial polysaccharide repeating unit undecaprenyl pyrophosphate and its analogs.

Polysaccharides are essential and immunologically relevant components of bacterial cell walls. These biomolecules can be found covalently attached to lipids (e.g.

Improvement of core-fucosylated glycoproteome coverage via alternating HCD and ETD fragmentation.

Unlabelled: Core-fucosylation (CF) plays important roles in regulating biological processes in eukaryotes. Alterations of CF-glycosites or CF-glycans in bodily fluids correlate with cancer development. Therefore, global research of protein core-fucosylation with an emphasis on proteomics can explain pathogenic and metastasis mechanisms and aid in the discovery of new potential biomarkers for early clinical diagnosis.

Biochemical characterization of an α1,2-colitosyltransferase from Escherichia coli O55:H7.

Colitose, also known as 3,6-dideoxy-L-galactose or 3-deoxy-L-fucose, is one of only five naturally occurring 3,6-dideoxyhexoses. Colitose was found in lipopolysaccharide of a number of infectious bacteria, including Escherichia coli O55 & O111 and Vibrio cholera O22 & O139. To date, no colitosyltransferase (ColT) has been characterized, probably due to the inaccessibility of the sugar donor, GDP-colitose.

Characterizing non-hydrolyzing Neisseria meningitidis serogroup A UDP-N-acetylglucosamine (UDP-GlcNAc) 2-epimerase using UDP-N-acetylmannosamine (UDP-ManNAc) and derivatives.

Neisseria meningitidis serogroup A non-hydrolyzing uridine 5'-diphosphate-N-acetylglucosamine (UDP-GlcNAc) 2-epimerase (NmSacA) catalyzes the interconversion between UDP-GlcNAc and uridine 5'-diphosphate-N-acetylmannosamine (UDP-ManNAc). It is a key enzyme involved in the biosynthesis of the capsular polysaccharide [-6ManNAcα1-phosphate-]n of N. meningitidis serogroup A, one of the six serogroups (A, B, C, W-135, X, and Y) that account for most cases of N.

Efficient Chemoenzymatic Synthesis of an N-glycan Isomer Library.

Quantification, characterization and biofunctional studies of N-glycans on proteins remain challenging tasks due to complexity, diversity and low abundance of these glycans. The availability of structurally defined N-glycans (especially isomers) libraries is essential to help on solving these tasks. We reported herein an efficient chemoenzymatic strategy, namely Core Synthesis/Enzymatic Extension (CSEE), for rapid production of diverse N-glycans.

Convenient and Precise Strategy for Mapping N-Glycosylation Sites Using Microwave-Assisted Acid Hydrolysis and Characteristic Ions Recognition.

N-glycosylation is one of the most prevalence protein post-translational modifications (PTM) which is involved in several biological processes. Alternation of N-glycosylation is associated with cellular malfunction and development of disease. Thus, investigation of protein N-glycosylation is crucial for diagnosis and treatment of disease.

Efficient chemoenzymatic synthesis of novel galacto-N-biose derivatives and their sialylated forms.

Galacto-N-biose (GNB) derivatives were efficiently synthesized from galactose derivatives via a one-pot two-enzyme system containing two promiscuous enzymes from Bifidobacterium infantis: a galactokinase (BiGalK) and a d-galactosyl-β1-3-N-acetyl-d-hexosamine phosphorylase (BiGalHexNAcP). Mono-sialyl and di-sialyl galacto-N-biose derivatives were then prepared using a one-pot two-enzyme system containing a CMP-sialic acid synthetase and an α2-3-sialyltransferase or an α2-6-sialyltransferase.

Improved one-pot multienzyme (OPME) systems for synthesizing UDP-uronic acids and glucuronides.

Arabidopsis thaliana glucuronokinase (AtGlcAK) was cloned and shown to be able to use various uronic acids as substrates to produce the corresponding uronic acid-1-phosphates. AtGlcAK or Bifidobacterium infantis galactokinase (BiGalK) was used with a UDP-sugar pyrophosphorylase, an inorganic pyrophosphatase, with or without a glycosyltransferase for highly efficient synthesis of UDP-uronic acids and glucuronides. These improved cost-effective one-pot multienzyme (OPME) systems avoid the use of nicotinamide adenine dinucleotide (NAD(+))-cofactor in dehydrogenase-dependent UDP-glucuronic acid production processes and can be broadly applied for synthesizing various glucuronic acid-containing molecules.

A Photobacterium sp. α2-6-sialyltransferase (Psp2,6ST) mutant with an increased expression level and improved activities in sialylating Tn antigens.

In order to improve the catalytic efficiency of recombinant Photobacterium sp. JT-ISH-224 α2-6-sialyltransferase Psp2,6ST(15-501)-His6 in sialylating α-GalNAc-containing acceptors for the synthesis of tumor-associated carbohydrate antigens sialyl Tn (STn), protein crystal structure-based mutagenesis studies were carried out. Among several mutants obtained by altering the residues close to the acceptor substrate binding pocket, mutant A366G was shown to improve the sialyltransferase activity of Psp2,6ST(15-501)-His6 toward α-GalNAc-containing acceptors by 21-115% without significantly affecting its sialylation activity to β-galactosides.

A precise approach in large scale core-fucosylated glycoprotein identification with low- and high-normalized collision energy.

Unlabelled: The core fucosylation (CF) of N-glycoproteins plays important roles in regulating protein functions during biological development, and it has also been shown to be up-regulated in several high metastasis cancer cell lines. Therefore, global profiling and quantitative characterization of CF-glycoproteins may reveal potent biomarkers for clinical applications. However, due to the complex fragmentation pattern of CF-glycopeptides, accurately identifying CF-glycosylation sites via mass spectrometry with high throughput remains a formidable challenge.

Synthetic disialyl hexasaccharides protect neonatal rats from necrotizing enterocolitis.

Two novel synthetic α2-6-linked disialyl hexasaccharides, disialyllacto-N-neotetraose (DSLNnT) and α2-6-linked disialyllacto-N-tetraose (DS'LNT), were readily obtained by highly efficient one-pot multienzyme (OPME) reactions. The sequential OPME systems described herein allowed the use of an inexpensive disaccharide and simple monosaccharides to synthesize the desired complex oligosaccharides with high efficiency and selectivity. DSLNnT and DS'LNT were shown to protect neonatal rats from necrotizing enterocolitis (NEC) and are good therapeutic candidates for preclinical experiments and clinical application in treating NEC in preterm infants.

Highly efficient one-pot multienzyme (OPME) synthesis of glycans with fluorous-tag assisted purification.

Oligo(ethylene glycol)-linked light fluorous tags have been found to be optimal for conjugating to glycans for both high-yield enzymatic glycosylation reactions using one-pot multienzyme (OPME) systems and quick product purification using fluorous solid-phase extraction (FSPE) cartridges. The combination of OPME glycosylation systems and the FSPE cartridge purification scheme provides a highly effective strategy for facile synthesis and purification of glycans.