A5074129234- Fungal and yeast genetics research
- Viral Infectious Diseases and Gene Expression in Insects
- Microbial Metabolic Engineering and Bioproduction
- Protein purification and stability
- Endoplasmic Reticulum Stress and Disease
- Monoclonal and Polyclonal Antibodies Research
- Biofuel production and bioconversion
- Gene expression and cancer classification
- Transgenic Plants and Applications
- Cellular transport and secretion
- Skin Protection and Aging
- Advanced MEMS and NEMS Technologies
- Circadian rhythm and melatonin
- Analog and Mixed-Signal Circuit Design
- Telomeres, Telomerase, and Senescence
- Mechanical and Optical Resonators
- Acoustic Wave Resonator Technologies
- Enzyme Catalysis and Immobilization
- Neonatal Respiratory Health Research
- Fermentation and Sensory Analysis
- Fungal Biology and Applications
- Advanced Proteomics Techniques and Applications
- Microfluidic and Bio-sensing Technologies
- Microbial Inactivation Methods
- COVID-19 Impact on Reproduction
FH Campus Wien
2011-2025
Reid Health
2022-2023
Austrian Centre of Industrial Biotechnology (Austria)
2011-2020
University of Freiburg
2007-2015
BOKU University
2003-2011
Universitat Autònoma de Barcelona
2009
Abstract The expression of heterologous proteins may exert severe stress on the host cells at different levels. Depending specific features product, steps be rate‐limiting. For secretion recombinant from yeast cells, folding and disulfide bond formation were identified as rate‐limiting in several cases induction chaperone BiP (binding protein) is described. During development Pichia pastoris strains secreting human trypsinogen, a limitation amount secreted product was identified. Strains...
Pichia pastoris is widely used as a production platform for heterologous proteins and model organism organelle proliferation. Without published genome sequence available, strain process development relied mainly on analogies to other, well studied yeasts like Saccharomyces cerevisiae. To investigate specific features of growth protein secretion, we have sequenced the 9.4 Mb type DSMZ 70382 analyzed secretome sugar transporters. The computationally predicted consists 88 ORFs. When grown...
The impact of environmental factors on the productivity yeast cells is poorly investigated so far. Therefore, it a major concern to improve understanding cellular physiology microbial protein production hosts, including methylotrophic Pichia pastoris. Two-Dimensional Fluorescence Difference Gel electrophoresis and identification via mass spectrometry were applied analyze cultivation temperature heterologous secreting P. pastoris strain. Furthermore, specific was monitored fluxes through...
Abstract The methylotrophic yeast Pichia pastoris has been used for the expression of many proteins, including antibody fragments. However, limitations became obvious especially when secreting heterodimeric Fab Up‐to‐date, fragments have only expressed under control strong inducible alcohol oxidase 1 (AOX1) promoter, which may stress cells by excessive transcription. Here, we examined secretion characteristics single chain and two different monoclonal anti‐HIV1 antibodies (2F5 2G12) with...
ABSTRACT Efficient production of heterologous proteins with yeasts and other eukaryotic hosts is often hampered by inefficient secretion the product. Limitation protein has been attributed to a low folding rate, rational solution overexpression supporting folding, like disulfide isomerase (Pdi), or unfolded response transcription factor Hac1. Assuming that factors which are not directly involved in may also support proteins, we set out analyze differential transcriptome Pichia pastoris...
The production of recombinant proteins is frequently enhanced at the levels transcription, codon usage, protein folding and secretion. Overproduction heterologous proteins, however, also directly affects primary metabolism producing cells. By incorporation a into genome scale metabolic model yeast Pichia pastoris, effects overproduction were simulated gene targets for deletion or overexpression productivity predicted. Overexpression localized in pentose phosphate pathway TCA cycle, while...
Abstract Background Inducible high-level expression is favoured for recombinant protein production in Pichia pastoris . Therefore, novel regulated promoters are desired, ideally repressing heterologous gene during initial growth and enabling it the phase. In a typical large scale fed-batch culture repression desired batch phase where cells grow on surplus of e.g. glycerol, while should be active feed under carbon (e.g. glucose) limitation. Results DNA microarray analysis P. wild type growing...
High cell density cultivation of Pichia pastoris has to cope with several technical limitations, most importantly the transfer oxygen. By applying hypoxic conditions chemostat cultivations P. expressing an antibody Fab fragment under GAP promoter, a 2.5-fold increase specific productivity q(P) at low oxygen supply was observed. At same time biomass decreased and ethanol produced, indicating shift from oxidative oxidofermentative conditions. Based on these results we designed feedback control...
To determine whether multiple doses of bovine surfactant would improve neonatal mortality in very premature neonates, we conducted two multicenter controlled trials under identical protocols; the results were combined for analysis. Four hundred and thirty neonates born between 23 29 weeks gestation weighing 600 to 1250 g at birth assigned randomly receive either 100 mg phospholipids/kg Survanta, a modified (n = 210), or sham air placebo 220) within 15 minutes birth. Neonates who developed...
Abstract Background It has become evident that host cells react to recombinant protein production with a variety of metabolic and intrinsic stresses such as the unfolded response (UPR) pathway. Additionally, environmental conditions growth temperature may have strong impact on cell physiology specific productivity. However, there is little information about molecular reactions genomic level, especially in context secretion. For first time, we monitored transcriptional regulation subset...
Abstract The methylotrophic yeast Pichia pastoris has gained much attention during the last decade as a platform for producing heterologous recombinant proteins of pharmaceutical importance, due to its ability reproduce post‐translational modification similar higher eukaryotes. With recent release full genome sequence P. , in‐depth study functions become feasible. Here we present first reconstruction genome‐scale metabolic model eukaryote type strain DSMZ 70382, PpaMBEL1254, consisting 1254...
Secretion of heterologous proteins depends both on biomass concentration and the specific product secretion rate, which in turn is not constant at varying growth rates. As fed batch processes usually do maintain a steady state throughout feed phase, it trivial to model optimize such process by mathematical means.We have developed for accumulation based iterative calculation Microsoft Excel spreadsheets, used Solver software time course media order maximize volumetric productivity. The...
Protein production in yeasts is related to the specific growth rate μ. To elucidate on this correlation, we studied transcriptome of Pichia pastoris at different rates by cultivating a strain secreting human serum albumin μ = 0.015 0.15 h(-1) glucose-limited chemostats. Genome-wide regulation revealed that translation-related as well mitochondrial genes were upregulated with increasing μ, while autophagy and other proteolytic processes, carbon source-responsive targets TOR pathway many...
Abstract Background The effect of osmolarity on cellular physiology has been subject investigation in many different species. High is importance for biotechnological production processes, where high cell densities and product titers are aspired. Several studies indicated that increased the growth medium can have a beneficial recombinant protein host organisms. Thus, Pichia pastoris , prominent production, was studied carbon limited chemostat cultures at osmolarities. Transcriptome proteome...
Chinese hamster ovary (CHO) cells are currently the workhorse of biopharmaceutical industry. However, yeasts such as Pichia pastoris about to enter this field. To compare their capability for recombinant protein secretion, P. strains and CHO cell lines producing human serum albumin (HSA) 3D6 single chain Fv-Fc anti-HIV-1 antibody (3D6scFv-Fc) were cultivated in comparable fed batch processes. In pastoris, mean biomass-specific secretion rate (qp ) was 40-fold lower 3D6scFv-Fc compared HSA....
Microorganisms encounter diverse stress conditions in their native habitats but also during fermentation processes, which have an impact on industrial process performance. These environmental stresses and the physiological reactions they trigger, including changes protein folding/secretion machinery, are highly interrelated. Thus, investigation of factors, influence expression secretion is still great importance. Among all possible stresses, temperature appears particularly important for...
The methylotrophic yeast Pichia pastoris is widely used for production of recombinant proteins. Here we characterize a vitamin-sensitive regulatory sequence, which can be controlled independently the main culture medium compounds such as carbon, nitrogen, or phosphor source. THI11 promoter (PTHI11 ) sequence derives from gene involved in biosynthesis thiamine. For characterization, P. strain expressing human serum albumin under control PTHI11 was grown environment bioreactor. thiamine...
Senolytic and senomorphic therapies gain more attention in the last decade. This kind of therapy is based on killing cellular senescent cells without harming “normal” cells. Ageing not a disease. Clinical studies healthy people will be difficult to conduct. Therefore, one possibility draw large repertoire medicinal plants use their sneolytic properties provide mild, anti-ageing therapies. Chamomile, goldenrod, reishi green tea were tested for ability trigger senolysis quercetin was used as...
BACKGROUND: Pichia pastoris is a well established yeast host for heterologous protein expression, however, the physiological and genetic information about this remains scanty. The lack of published genome sequence renders DNA arrays unavailable, thereby hampering more global investigations P. from beginning. Here, we examine suitability Saccharomyces cerevisiae microarrays hybridisation with cDNA. RESULTS: We could show that it possible to obtain new valuable transcriptomic regulation in by...
Abstract Background The budding yeast Pichia pastoris is widely used for protein production. To determine the best suitable strategy strain improvement, especially high secretion, quantitative data of intracellular fluxes recombinant are very important. Especially balance between formation, degradation and secretion defines major bottleneck production system. Because these parameters different unlimited growth (shake flask) carbon-limited (bioreactor) conditions, they should be determined...