Abstract: A method of producing a protein of interest (POI) by culturing a recombinant eukaryotic cell line comprising an expression construct comprising a regulatable promoter and a nucleic acid molecule encoding a POI under the transcriptional control of said promoter, comprising the steps a) cultivating the cell line with a basal carbon source repressing the promoter, b) cultivating the cell line with a limited amount of a supplemental carbon source de-repressing the promoter to induce production of the POI at a transcription rate of at least 15% as compared to the native pGAP promoter, and c) producing and recovering the POI; and further an isolated regulatable promoter and a respective expression system.

Abstract: The invention refers to a method of producing a recombinant polypeptide of interest (POI) in a cell culture, comprising genetically engineering a eukaryotic cell line—to specifically cause prolongation of the G2+M cell cycle phase in a pre-culture phase, and—to produce the POI in a producing phase following the pre-culture phase, a high producer cell line and cell culture as well as a method of increasing the yield of a recombinant POI production in a cell culture.

Abstract: Invention relates to a eukaryotic host cell comprising a recombinant nucleotide sequence encoding an expression enhancer, which is selected from the group consisting of cLC52, RPL33 and cLC61, and its use in a method of producing a protein of interest (POI).

Abstract: The present invention relates to methods for increasing the secretion of a protein of interest (POI) from a eukaryotic cell comprising co-expression of a POI and of at least one protein that enhances protein secretion, said enhancing protein being selected from the group consisting of BMH2, BFR2, C0G6, C0Y1, CUP5, IMH 1, KIN2, SEC31, SSA4 and SSE1. The invention further relates to a yeast promoter sequence, in particular to a promoter sequence of the PET9 gene of P. pastoris, having, under comparable conditions, an increased promoter activity relative to a promoter sequence of the GAP protein. The invention further relates to an expression vector comprising such a promoter sequence and to the use of such an expression vector for expression of a POI in a host cell. The invention further relates to new yeast promoter sequences of genes from P. pastoris, which are useful for expression of a POI in yeast.

Abstract: A method of producing an organic acid by staining a yeast population with a stain capable of internal pH (pHi)-dependent fluorescence, to yield a stained yeast population; determining a gate pH and a corresponding fluorescence parameter of the stained yeast population; and sorting the cells of the stained yeast population such that the cells having a pHi above the gate pH are retained and the cells having a pHi below the gate pH are discarded, to yield a yeast population for the production of the organic acid. Also, a method of producing an organic acid by performing the above steps, followed by isolating individual cells of the yeast population, to yield individual yeast cells for the production of an organic acid; culturing an individual yeast cell, to yield a cloned yeast population for the production of an organic acid; and incubating the cloned yeast population for the production of an organic acid in a medium containing an organic acid precursor, to produce the organic acid.

Abstract: The present invention relates to a method for the manufacture and purification of recombinant trypsinogen and trypsin in E. coli and yeast, using high yield expression vectors with and without secretion leader sequences. The invention further relates to an improved method and apparatus for carrying out protein refolding specifically useful for processing trypsinogen that has accumulated intracellularly in the form of inclusion bodies.

Abstract: A method of increasing stress tolerance in recombinant organisms that have been engineered for industrial production is described. Stress tolerance is increased by making L-ascorbic acid available to the recombinant organism, either by exogenous addition to the culture medium or by endogenous production from D-glucose by the recombinant organism. To enable endogenous production, the recombinant organism is transformed with a coding region encoding a mannose epimerase (ME), a coding region encoding an L-galactose dehydrogenase (LGDH), and a D-arabinono-1,4-lactone oxidase (ALO). The recombinant organism may be further transformed with a myoinositol phosphatase (MIP).

Abstract: A method of producing an organic acid by staining a yeast population with a stain capable of internal pH (pHi)-dependent fluorescence, to yield a stained yeast population; determining a gate pH and a corresponding fluorescence parameter of the stained yeast population; and sorting the cells of the stained yeast population such that the cells having a pHi above the gate pH are retained and the cells having a pHi below the gate pH are discarded, to yield a yeast population for the production of the organic acid. Also, a method of producing an organic acid by performing the above steps, followed by isolating individual cells of the yeast population, to yield individual yeast cells for the production of an organic acid; culturing an individual yeast cell, to yield a cloned yeast population for the production of an organic acid; and incubating the cloned yeast population for the production of an organic acid in a medium containing an organic acid precursor, to produce the organic acid.

Abstract: Herein is disclosed a method of generating ascorbic acid from yeast transformed with a mannose epimerase. In a further embodiment, the yeast can be further transformed with a myoinositol phosphatase. In the method, the transformed yeast can produce L-ascorbic acid from D-glucose. The transformed yeast has been observed to have increased growth rate, cell density, or survival when cultured on appropriate media.

Abstract: The present invention relates to a method for the manufacture and purification of recombinant trypsinogen and trypsin in E. coli and yeast, using high yield expression vectors with and without secretion leader sequences. The invention further relates to an improved method and apparatus for carrying out protein refolding specifically useful for processing trypsinogen that has accumulated intracellularly in the form of inclusion bodies.