yeast glycosylation

Terpenoids are important contributors to the aroma of grapes and wines. Grapes contain terpenoids in both volatile free form and non-volatile glycosidic form, with the latter being more abundant. Glycosylated terpenoids are deemed as latent aromatic potentials for their essential role in adding to the flowery and fruity bouquet of wines. However, the transcriptional regulatory mechanism underlying glycosylated terpenoid biosynthesis remains poorly understood. Our prior study identified an AP2/ERF transcription factor, VviERF003, through DNA pull-down screening using the promoter of terpenoid glycosyltransferase VviGT14 gene...

Plant cells are a capable system for producing economically and therapeutically important proteins for a variety of applications, and are considered a safer production system than some existing hosts such as bacteria or yeasts. However, plants do not perform protein modifications in the same manner as mammalian cells do. This can impact on protein functionality for plant-produced human therapeutics. This obstacle can be overcome by creating a plant-based system capable of 'humanising' proteins of interest resulting in a glycosylation profile of synthetic plant-produced proteins as it would occur in mammalian systems...

Bacterioruberin and its rare glycosylated derivatives are produced by Arthrobacter agilis as an adaptation strategy to low temperature conditions. The high antioxidant properties of bacterioruberin held great promise for different future applications like the pharmaceutical and food industries. Microbial production of bacterioruberin via a cost-effective medium will help increase its commercial availability and industrial use. The presented study aims to optimize the production of the rare C50 carotenoid bacterioruberin and its derivatives from the psychotrophic bacteria Arthrobacter agilis NP20 strain on a whey-based medium as a cost effective and readily available nutritious substrate...

The methylotrophic yeast, Pichia pastoris (P. pastoris; syn. Komagataella spp.), known for its ability to grow to high cell densities, its strong and tightly regulated promoters, and mammalian liked secretion pathway, has been widely used as a robust system to secrete heterologous proteins. The α-mating factor (MF) secretion signal leader from Saccharomyces cerevisiae (S. cerevisiae) is currently the most successfully used secretion signal sequence in the P. pastoris system. In this study, the secretion efficiency mediated by the α-MF secretion signal leaders from Komagataella pastoris (K...

Glycosyl-phosphatidylinositol (GPI) anchored proteins are implicated in remodeling of the yeast cell wall during growth and division. Schizosaccharomyces pombe proteins, Psu1 , Dfg501 , and Dfg502 are predicted GPI anchored proteins with likely cell wall modifying activity. Here, we isolated and characterized null and temperature-sensitive alleles that will allow further analysis of the function of these proteins and S. pombe cell wall formation. Our data confirm that Psu1 is necessary for cell separation, maintaining proper cell shape, and viability...

Hydrostatic pressure is a common mechanical stressor that modulates metabolism and reduces cell viability. Eukaryotic cells have genetic programs to cope with hydrostatic pressure stress and maintain intracellular homeostasis. However, the mechanism underlying hydrostatic pressure tolerance remains largely unknown. We have recently demonstrated that Maintenance of telomere capping protein 6 (Mtc6) plays a protective role in the survival of the budding yeast Saccharomyces cerevisiae under hydrostatic pressure stress by supporting the integrity of nutrient permeases...

The oligosaccharide needed for protein N -glycosylation is assembled on a lipid carrier via a multi-step pathway. Synthesis is initiated on the cytoplasmic face of the endoplasmic reticulum (ER) and completed on the luminal side after transbilayer translocation of a heptasaccharide lipid intermediate. More than 30 Congenital Disorders of Glycosylation (CDGs) are associated with this pathway, including CDG 1N which results from defects in the membrane protein Rft1. Rft1 is essential for the viability of yeast and mammalian cells and was proposed as the transporter needed to flip the heptasaccharide lipid intermediate across the ER membrane...

Biotransformation of minerals via glycosylation by microorganisms such as yeast and/or probiotics yields nutrients bound to a food matrix, resulting in increased bioavailability. The purpose of this study was to compare the effects of glycoprotein matrix-bound zinc (GPM) on absorption compared to inorganic zinc oxide. Sixteen participants ingested 11 mg of zinc as either GPM™ Soy-Free Zinc (GPM, Ashland, Kearny, NJ, USA) or zinc oxide (USP). Blood samples were taken at 0 (i.e., baseline), 30, 60, 90, 120, 180, 240, 300, 360, 420, and 480 min post-ingestion...

Mitogen-activated protein kinase (MAPK) pathways control the response to intrinsic and extrinsic stimuli. In the budding yeast Saccharomyces cerevisiae, cells undergo filamentous growth, which is regulated by the fMAPK pathway. To better understand the regulation of the fMAPK pathway, a genetic screen was performed to identify spontaneous mutants with elevated activity of an fMAPK-pathway dependent growth reporter (ste4 FUS1-HIS3). In total, 159 mutants were isolated and analyzed by secondary screens for invasive growth by the plate-washing assay, and filament formation by microscopy...

Brewer's spent yeast (BSY) hydrolysates are a source of antidiabetic peptides. Nevertheless, the impact of in vitro gastrointestinal digestion of BSY derived peptides on diabetes has not been assessed. In this study, two BSY hydrolysates were obtained ( H1 and H2 ) using β-glucanase and alkaline protease, with either 1 h or 2 h hydrolysis time for H1 and H2 , respectively. These hydrolysates were then subjected to simulated gastrointestinal digestion (SGID), obtaining dialysates D1 and D2 , respectively...

Sugar metabolism plays a pivotal role in sustaining life. Its dynamics within organisms is less understood compared to its intracellular metabolism. Galactose, a hexose stereoisomer of glucose, is a monosaccharide transported via the same transporters with glucose. Galactose feeds into glycolysis and regulates protein glycosylation. Defects in galactose metabolism are lethal for animals. Here, by transgenically implementing the yeast galactose sensing system into Drosophila, we developed a genetically encoded sensor, GALDAR, which detects galactose in vivo...

Glycosylation of biomolecules can greatly alter their physicochemical properties, cellular recognition, subcellular localization, and immunogenicity. Glycosylation reactions rely on the stepwise addition of sugars using nucleotide diphosphate (NDP)-sugars. Making these substrates readily available will greatly accelerate the characterization of new glycosylation reactions, elucidation of their underlying regulation mechanisms, and production of glycosylated molecules. In this work, we engineered Saccharomyces cerevisiae to heterologously express nucleotide sugar synthases to access a wide variety of uridine diphosphate (UDP)-sugars from simple starting materials (i...

UNLABELLED: Crocus sativus L., of the Iridaceae family, yields world's most prized spice, saffron. Saffron is well known for its distinctive aroma, odour and colour, which are imputed to the presence of some specific glycosylated apocarotenoids. Even though the main biosynthetic pathway and most of the enzymes leading to apocarotenoid production have been identified, the regulatory mechanisms that govern the developmental stage and tissue specific production of apocarotenoids in Crocus remain comparatively unravelled...

N-linked glycosylation is an essential and highly conserved co- and post-translational protein modification in all domains of life. In humans, genetic defects in N-linked glycosylation pathways result in metabolic diseases collectively called Congenital Disorders of Glycosylation. In this modification reaction, a mannose rich oligosaccharide is transferred from a lipid-linked donor substrate to a specific asparagine side-chain within the -N-X-T/S- sequence (where X ≠ Proline) of the nascent protein...

Peptidylglycine α-hydroxylating monooxygenase (PHM) is pivotal for C-terminal amidation of bioactive peptides in animals, offering substantial potential for customized protein synthesis. However, efficient PHM production has been hindered by the complexity of animal cell culture and the absence of glycosylation in bacterial hosts. Here, we demonstrate the recombinant expression of Caenorhabditis elegans PHM in the yeast Pichia pastoris, achieving a remarkable space-time yield of 28.8 U/L/day. This breakthrough surpasses prior PHM production rates and eliminates the need for specialized cultivation equipment or complex transfection steps...

The neuron-specific K+ /Cl- co-transporter-2, KCC2, which is critical for brain development, regulates γ-aminobutyric acid-dependent inhibitory neurotransmission. Consistent with its function, mutations in KCC2 are linked to neurodevelopmental disorders, including epilepsy, schizophrenia, and autism. KCC2 possesses 12 transmembrane spans and forms an intertwined dimer. Based on its complex architecture and function, reduced cell surface expression and/or activity have been reported when select disease-associated mutations are present in the gene encoding the protein, SLC12A5...

Previous data established 4'-deoxyflavone glycosides (4'-DFGs) as important pharmaceutical components in the roots of rare medical plants like Scutellaria baicalensis Georgi. Extracting these compounds from plants involves land occupation and is environmentally unfriendly. Therefore, a modular ("plug-and-play") yeast-consortium platform is developed to synthesize diverse 4'-DFGs de novo. By codon-optimizing glycosyltransferase genes from different organisms for Pichia pastoris, six site-specific glycosylation chassis are generated to be capable of biosynthesizing 18 different 4'-DFGs...

Immune checkpoint inhibitor (ICI) therapy improves outcomes in several cancers. Unfortunately, many patients experience grade 3-4 treatment-related adverse events, including gastrointestinal (GI) toxicities which are common. These GI immune-related adverse events (irAEs) induced by ICIs present significant clinical challenges, require prompt intervention, and result in treatment delays or discontinuations. The treatment for these potentially severe and even fatal GI irAEs which include enterocolitis, severe diarrhea, and hepatitis may interfere with the anti-cancer approach...

Mycobacterium marinum CAR (MmCAR) is one of the most widely used CARs as the key enzyme for the synthesis of aldehydes, alcohols and further products from the respective carboxylic acids. Herein, we describe the first functionally secreted 131kDa CAR and its isolated A-domain using Komagataella phaffii and a methanol-free constitutive expression strategy. Precipitated and lyophilized MmCAR (500µg) was isolated from the culture supernatant and showed no decrease in activity for piperonylic acid (80% conversion), even when stored for up to 3 weeks at 4°C...

Seryl-tRNA synthetases (SerRSs), members of the aminoacyl-tRNA synthetase family, interact with diverse proteins, enabling SerRSs to enhance their role in the translation of the genetic message or to perform alternative functions in cellular processes beyond translation. Atypical archaeal SerRS interacts with arginyl-tRNA synthetase and proteins of the ribosomal P-stalk to optimize translation through tRNA channeling. The complex between yeast SerRS and peroxin Pex21p provides a connection between translation and peroxisome function...