A5009248613- Viral Infectious Diseases and Gene Expression in Insects
- CRISPR and Genetic Engineering
- Virus-based gene therapy research
- RNA and protein synthesis mechanisms
- Cellular transport and secretion
- Microbial Metabolic Engineering and Bioproduction
- Protein Kinase Regulation and GTPase Signaling
- Ubiquitin and proteasome pathways
- Endoplasmic Reticulum Stress and Disease
- Monoclonal and Polyclonal Antibodies Research
- Fungal and yeast genetics research
- bioluminescence and chemiluminescence research
- Melanoma and MAPK Pathways
- Viral Infections and Immunology Research
- Bacterial Genetics and Biotechnology
- Biotin and Related Studies
- Biochemical Acid Research Studies
- Insect Resistance and Genetics
- Microtubule and mitosis dynamics
- Peptidase Inhibition and Analysis
- Lipid Membrane Structure and Behavior
- RNA Research and Splicing
- Cytokine Signaling Pathways and Interactions
- Epigenetics and DNA Methylation
- Bacterial biofilms and quorum sensing
Novo Nordisk Foundation
2013-2025
Technical University of Denmark
2019-2025
University of Copenhagen
2013-2014
Thermo Fisher Scientific (Denmark)
2006
Novo Nordisk (Denmark)
1998
SickKids Foundation
1989
University of Toronto
1989
Hospital for Sick Children
1989
ABSTRACT Use of the green fluorescent protein (Gfp) from jellyfish Aequorea victoria is a powerful method for nondestructive in situ monitoring, since expression fluorescence does not require any substrate addition. To expand use Gfp as reporter protein, new variants have been constructed by addition short peptide sequences to C-terminal end intact Gfp. This rendered susceptible action indigenous housekeeping proteases, resulting with half-lives ranging 40 min few hours when synthesized...
The widely used pESC vector series (Stratagene, La Jolla, CA, USA) with the bidirectional GAL1/GAL10 promoter provides possibility of simultaneously expressing two different genes from a single in Saccharomyces cerevisiae. This system can be induced by galactose and is repressed glucose. Since S. cerevisiae prefers glucose as carbon source, since its growth rate higher than galactose-containing media, we compared evaluated seven promoters expressed during on (pTEF1, pADH1, pTPI1, pHXT7,...
BubR1 is a central component of the spindle assembly checkpoint that inhibits progression into anaphase in response to improper kinetochore-microtubule interactions. In addition, also helps stabilize interactions by counteracting Aurora B kinase but mechanism behind this not clear. Here we show directly binds B56 family protein phosphatase 2A (PP2A) regulatory subunits through conserved motif phosphorylated cyclin-dependent 1 (Cdk1) and polo-like (Plk1). Two highly hydrophobic residues...
Abstract Allosteric transcription factors (aTFs) have proven widely applicable for biotechnology and synthetic biology as ligand-specific biosensors enabling real-time monitoring, selection regulation of cellular metabolism. However, both the biosensor specificity correlation between ligand concentration output signal, also known transfer function, often needs to be optimized before meeting application needs. Here, we present a versatile high-throughput method evolve prokaryotic aTF...
Abstract Oligoclonal mixtures of broadly-neutralizing antibodies can neutralize complex compositions similar and dissimilar antigens, making them versatile tools for the treatment e.g., infectious diseases animal envenomations. However, these biotherapeutics are complicated to develop due their nature. In this work, we describe application various strategies discovery cross-neutralizing nanobodies against key toxins in coral snake venoms using phage display technology. We prepare two...
The PRP4 (RNA4) gene product is involved in nuclear mRNA processing yeast cells; we have previously cloned the by complementation of a temperature-sensitive mutation. Sequence and transcript analyses predicted to be 52-kilodalton protein, which was confirmed with antibodies raised against product. These inhibited precursor splicing vitro, demonstrating direct role splicing. Immunoprecipitations indicated that protein associated U4/U6 small ribonucleoprotein particle.
The metabolism of gluconate is well characterized in prokaryotes where it known to be degraded following phosphorylation by gluconokinase. Less humans. Human gluconokinase activity was recently identified proposing questions about the metabolic role Here we report recombinant expression, purification and biochemical characterization isoform I human alongside substrate specificity kinetic assays enzyme catalyzed reaction. enzyme, shown a dimer, had ATP dependent strict towards out 122...
The PI3-kinase/Akt pathway is an important cell survival that deregulated in the majority of human cancers. Despite apparent druggability several kinases pathway, no specific catalytic inhibitors have been reported literature. authors describe development a fluorometric imaging plate reader (FLIPR)-based Akt1 translocation assay to discover activation. Screening diverse chemical library 45,000 compounds resulted identification classes inhibitors. Using combination classical vitro assays and...
Redistribution® (BioImage® A/S, Søborg, Denmark) is a novel high-throughput screening technology that monitors translocation of specific protein components intracellular signaling pathways within intact mammalian cells, using green fluorescent as tag. A single Redistribution assay can be used to identify multiple classes compounds act at, or upstream of, the level target in primary assay. Such may include both conventional and allosteric enzyme inhibitors, well protein-protein interaction...
Abstract Viral contamination in biopharmaceutical manufacturing can lead to shortages the supply of critical therapeutics. To facilitate protection bioprocesses, we explored basis for susceptibility CHO cells RNA virus infection. Upon infection with certain ssRNA and dsRNA viruses, fail generate a significant interferon (IFN) response. Nonetheless, downstream machinery generating IFN responses its antiviral activity is intact these cells: treatment exogenously-added type I or poly I:C prior...
Green fluorescent protein-assisted readout for interacting proteins (GRIP) is a universal protein interaction discovery system that can be used to generate truly high throughput screening-compatible cellular assays screen inhibitors of protein-protein interactions. The technology uses "bait and prey" principle based on the distinct translocation behavior human cyclic AMP phosphodiesterase 4A4. Here we use p53-Hdm2 Redistribution assay (Fisher BioImage ApS, Søborg, Denmark) as an example...
Abstract Allosteric transcription factors (aTFs) have proven widely applicable for biotechnology and synthetic biology as ligand-specific biosensors enabling real-time monitoring, selection regulation of cellular metabolism. However, both the biosensor specificity correlation between ligand concentration output signal, also known transfer function, often needs to be optimized before meeting application needs. Here, we present a versatile high-throughput method evolve functionalize...
Chinese hamster ovary (CHO) cell lines are widely used in industry for biological drug production. During culture development, considerable effort is invested to understand the factors that greatly impact growth, specific productivity and product qualities of biotherapeutics. While high-throughput omics approaches have been increasingly utilized reveal cellular mechanisms associated with line phenotypes guide process optimization, comprehensive data analysis management a challenge. Here we...
The Warburg effect is ubiquitous in proliferative mammalian cells, including cancer but poses challenges for biopharmaceutical production, as lactate accumulation inhibits cell growth and protein production. Previous efforts to eliminate production via knockout have failed bioprocessing since dehydrogenase has proven essential. However, here we eliminated the Chinese hamster ovary (CHO) HEK293 cells by simultaneously knocking out regulators involved a negative feedback loop that typically...
The RAS-mitogen-activated protein kinase (MAPK) signaling pathway has a central role in regulating the proliferation and survival of both normal tumor cells. This been 1 focus area for development anticancer drugs, resulting several compounds, primarily inhibitors, clinical testing. authors have undertaken cell-based, high-throughput screen using novel ERF1 Redistribution assay to identify compounds that modulate pathway. hit were subsequently tested activity functional cell designed...
Chemical modification of proteins has numerous applications, but it been challenging to achieve the required high degree selectivity on lysine amino groups. Recently, we described highly selective acylation with an N-terminal Gly-His6 segment. This tag promoted Nα -amine resulting in stable conjugates. Herein, report peptide sequences Hisn -Lys-Hism , which term Lys-His tags. In combination simple acylating agents, they facilitate designated Lys Nϵ under mild conditions and over native...
This chapter describes the design and development of cell-based assays, in which quantitation intracellular translocation a target protein--rather than binding or catalytic activity--provides primary assay readout. These are inherently high content they provide feedback on cellular response at systems level, rather data activities individual, purified molecules. Multiple protein assays can be used to profile signaling pathways play key role determination mechanism action for novel classes...
Abstract Alpha-1-antitrypsin (A1AT) is a multifunctional, clinically important, high value therapeutic glycoprotein that can be used for the treatment of many diseases such as alpha-1-antitrypsin deficiency, diabetes, graft-versus-host-disease, cystic fibrosis and various viral infections. Currently, only FDA-approved A1AT disorders intravenous augmentation therapy with human plasma-derived A1AT. In addition to its limited supply, this approach poses risk infection transmission, since it...
Abstract Despite their therapeutic potential, many protein drugs remain inaccessible to patients since they are difficult secrete. Each recombinant has unique physicochemical properties and requires different machinery for proper folding, assembly, posttranslational modifications (PTMs). Here we aimed identify the supporting secretion by measuring protein–protein interaction (PPI) networks of four proteins (SERPINA1, SERPINC1, SERPING1, SeAP) with various PTMs structural motifs using...
Abstract Viral contamination in biopharmaceutical manufacturing can lead to shortages the supply of critical therapeutics. To facilitate protection bioprocesses, we explored basis for susceptibility CHO cells, most commonly used cell line biomanufacturing, RNA virus infection. Upon infection with certain ssRNA and dsRNA viruses, cells fail generate a significant interferon (IFN) response. Nonetheless, downstream machinery generating IFN responses its antiviral activity is intact these cells:...
Glycosylation is fundamentally important to the activity, stability and pharmacokinetics of biopharmaceuticals. Move on future, glycoengineering a promising way improve protein drug efficacy. Recently, several major advances in engineering efforts have successfully led desired glycoforms, but some desirable designs proven toxic host cells (e.g., CHO cell lines). Up date, critical barrier controlling glycan structures diversity complexity glycosylation. Moreover, there has not been systematic...
Abstract Despite their therapeutic potential, many protein drugs remain inaccessible to patients since they are difficult secrete. Each recombinant has unique physicochemical properties and requires different machinery for proper folding, assembly, post-translational modifications (PTMs). Here we aimed identify the supporting secretion by measuring protein-protein interaction (PPI) networks of four proteins (SERPINA1, SERPINC1, SERPING1 SeAP) with various PTMs structural motifs using...