Showing papers on "Pichia pastoris published in 1987"

Expression of the lacZ gene from two methanol-regulated promoters in Pichia pastoris

Abstract: Two DNA fragments containing putative control regions regulating the expression of the alcohol oxidase (AOX) and dihydroxy-acetone synthase (DAS) genes from the methylotrophic yeast Pichia pastoris were used in the construction of vectors for the expression of the Escherichia coli lacZ gene. These vectors were transformed into P. pastoris host cells and employed in experiments to measure the control mechanisms employed by each promoter in the production of beta-galactosidase fusion products. Results in P. pastoris suggest that the processes used to regulate the expression of these gene fusions involve both repression/derepression and induction mechanisms. Expression of the AOX-lacZ and DAS-lacZ fusions was examined in Saccharomyces cerevisiae as well. Interestingly, beta-galactosidase was expressed in a regulated manner in the heterologous host.

High–Level Expression and Efficient Assembly of Hepatitis B Surface Antigen in the Methylotrophic Yeast, Pichia Pastoris

Abstract: The methylotrophic industrial yeast, Pi–chia pastoris, has been developed as a host system for the large–scale production of heterologous proteins. As an example of the systems, the synthesis of hepatitis B surface antigen (HBsAg) is described. Expression of the HBsAg gene is controlled by a methanol–regulated promoter derived from the primary alcohol oxidase gene, AOX1, of P. pastoris. The highest level of HBsAg is observed in a novel mutant strain of P. pastoris in which the AOX1 structural gene is deleted and replaced by a construction composed of the AOX1 promoter–HBsAg gene expression cassette. When grown on methanol, 2–3% of the soluble protein in this strain is HBsAg. Characterization of the recombinant HBsAg indicates that virtually all of the P. pastoris–synthesized HBsAg exists in a form which is similar to human serum–derived HBsAg 22 nm particles. Scale–up of the HBsAg production process from shake–flask to high cell–density fermentor cultures is described as well. A single culture of the P. pastoris HBsAg expression strain in a volume of 240 liters and at a density of 60 grams/liter dry weight of cells produced 0.4 grams/liter, or a total of 90 grams of HBsAg 22 nm particles, an amount sufficient for approximately 9 million doses of vaccine.

High-Level Secretion of Glycosylated Invertase in the Methylotrophic Yeast, Pichia Pastoris

Abstract: High-Level Secretion of Glycosylated Invertase in the Methylotrophic Yeast, Pichia Pastoris

β-Oxidation Systems in Eukaryotic Microorganisms

Abstract: In yeasts and fungi capability to degrade fatty acids is not limited to some specialized species of Candida. For example, the methylotropic yeasts Hansenula polymorpha and Pichia pastoris, the methylamine-oxidizing yeast Trichosporon cutaneum, the filamentous fungus Neurospora crassa, and even Saccharomyces cerevisiae (baker’s yeast) possess an inducible, non-mitochondrial s-oxidation system. The induction by oleate is associated with marked proliferation of microbodies.

Process for preparing a catalase-free oxidase and a catalase-free oxidase-containing yeast, and use thereof

Abstract: The invention provides the preparation of an oxidase or oxidase-containing mixture by aerobic fermentation of a methylotrophic yeast, wherein a catalase-negative yeast is allowed to grow in a nutritive medium in the presence of: (a) a so-called inducing substrate that induces expression of the oxidase gene and may also be substrate for the oxidase, and (b) another source of carbon that is suitable for the yeast species chosen, the molar ratio of inducing substrate to other source of carbon being such that the yeast formed and the oxidase formed do not suffer any harmful effects from the oxidation of the inducing substrate. For example, methanol oxidase is formed by induction with methanol in the presence of glucose. Examples of yeasts are Hansenula polymorpha and Pichia pastoris . Specific ratios of inducing substrate and glucose as other carbon source are (0.025-3):1 and preferably (1-­1.8):1. The yeasts are not capable of producing catalase, so that the oxidase produced can be used for producing hydrogen peroxide, e.g. when an alkanol is combined with methanol oxidase in a washing or bleaching process. The oxidase can also be used for qualitative and/or quantitative determination of a substrate for the oxidase, e.g. ethanol or amines, or be used as a catalyst for the oxidation of a substrate in chemical synthesis.

The Methylotrophic Yeast Pichia Pastoris as a Host for Heterologous Protein Production

Abstract: Yeasts have held great promise as hosts for production of economically important proteins and peptides using recombinant DNA technology. Among the heralded features of the bakers yeast, Saccharomyces cerevisiae, are its safety (lack of toxicity, pyrogenicity and allergenicity), ability to be grown on an industrial scale, and the vast wealth of information regarding its genetics and molecular biology which has led to the development of a number of expression vector systems. Invariably these vectors are based on the endogenous multicipy plasmid, 2 micron circle, and use a variety of promoter systems, either constitutive or regulated, to drive the expression of the desired gene [1] [2].

Microbial growth on C[1] compounds : proceedings of the 5th international symposium

Abstract: An eightieth anniversary of the study of microbial C1 metabolism.- Physiology and Biochemistry of Autotrophic Bacteria.- Physiology and Biochemistry of Autotrophic Bacteria.- Metabolic and Molecular Regulation of the CO2-Assimilating Enzyme System in Aerobic Chemoautotrophs.- CO2 fixation in Rhodopseudomonas blastica.- Autotrophic CO2 Fixation in Chemotrophic Anaerobic Bacteria.- Aerobic and Anaerobic Extremely Thermophilic Autotrophs.- Physiology and Biochemistry of Methanogens and Fermentative C1 Utilizers.- Biochemical Aspects of Methane Formation in Methanobacterium thermoautotrophicum.- Intermediary Steps in Methanogenesis.- Recent Developments on the Biochemistry of Methanogenesis from Acetate.- C1-Utilizing Acetogens.- Ecology of C1-Metabolizing Anaerobes.- Physiology and Biochemistry of Methylotrophic Bacteria.- Physiology and Biochemistry of Methylotrophic Bacteria.- Metabolic Regulation in Facultative Methylotrophs.- Quinoproteins in the Dissimilation of C1 Compounds.- NMR Spectroscopy in the Study of C1-Metabolism.- Comparative Biochemistry of C1-Carriers.- Physiology, Biochemistry and Genetics of Methylotrophic Yeasts.- Methylotrophic Yeasts - 1986.- Metabolic Regulation in Methylotrophic Yeasts.- Biochemistry of Alcohol Oxidase.- Genetics of Methylotrophic Yeasts.- Protein Topogenesis in Methylotrophic Yeasts.- Energetics of C1-Metabolism.- Energetics of C1-Compound Metabolism.- Energy Metabolism in Chemolithotrophs.- Energy Metabolim of Aerobic, Methylotrophic Bacteria.- Energy Metabolism in Methanogens.- Theoretical Limits of Growth Yields and an Analysis of Experimental data.- Genetics and Molecular Biology of C1-Utilizers.- The Molecular Genetics of C1 Utilizing Microorganisms - An Overview.- Genetics of Aerobic Lithoautotrophs.- Molecular Biology and Regulation of Microbial CO2 fixation.- Molecular Biology of Methanol Oxidation.- Divergence of Methanogens, Conservation of the Hisl Gene Sequence in All Three Biological Kingdoms and the Status of Methanobacterium thermoautotrophicum.- Applied Aspects of One-Carbon Metabolism.- The Potential of One-Carbon Compounds as Fermentation Feedstocks.- Chemical Synthesis from C1 Compounds.- Microbiological Synthesis from C1-Compounds: Application of Some Methylotrophic Functions to Synthesis of Useful Chemicals.- The Methylotrophic Yeast Pichia pastoris as a Host for Heterologous Protein Production.- Authors index.