Abstract: A recombinant host cell comprising an endogenous gene encoding a FLO8 protein comprising the amino acid sequence identified as SEQ ID NO:1 or a homologue thereof, which host cell is engineered by one or more genetic modifications to reduce expression of said gene compared to the host cell prior to said one or more genetic modifications, and which host cell comprises a heterologous expression cassette comprising a gene of interest (GOI) under the control of an expression cassette promoter (ECP) which ECP is repressible by a non-methanol carbon source, and a method of producing a protein of interest using said recombinant host cell.

Abstract: A recombinant eukaryotic host cell expressing a gene of interest (GOI) which is engineered by genetic modifications to increase expression of two or more genes encoding translation initiation factors (TIF genes) of the messenger ribonucleoprotein (mRNP), compared to the host cell prior to said one or more genetic modifications, wherein said TIF genes comprise at least a gene encoding eIF4A and a gene encoding eIF4G, and wherein expression of at least one of said TIF genes is under transcriptional control of a promoter different from the promoter controlling expression of said GOI.

Abstract: A recombinant host cell comprising an endogenous gene encoding a FLO8 protein comprising the amino acid sequence identified as SEQ ID NO:1 or a homologue thereof, which host cell is engineered by one or more genetic modifications to reduce expression of said gene compared to the host cell prior to said one or more genetic modifications, and which host cell comprises a heterologous expression cassette comprising a gene of interest (GO!) under the control of an expression cassette promoter (ECP) which ECP is repressible by a non-methanol carbon source, and a method of producing a protein of interest using said recombinant host cell.

Abstract: A method for producing of a protein of interest (POI) in a yeast host cell that is modified to comprise within one or more expression cassettes heterologous nucleic acid molecules encoding helper factors and a gene of interest (GOI) encoding the POI, wherein: a) a first helper factor comprises at least 90% sequence identity to SEQ ID NO:1; b) a second helper factor comprises at east 90% sequence identity to SEQ ID NO:3; and c) a third helper factor comprises at least 90% sequence identity to SEQ ID NO:5; which method comprises (i) culturing said host cell in a culture medium under conditions to co-express said heterologous nucleic acid molecules and to secrete said POI into the host cell culture; and (ii) recovering the POI from the host cell culture.

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: 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: An isolated nucleic acid encoding a leader, which has a specific sequence, an isolated leader peptide encoded by such nucleic acid, an expression cassette comprising such nucleic acid encoding a leader operably linked to a nucleic acid sequence encoding a POI a recombinant yeast host cell or a vector comprising such expression cassette, a method of producing a POI in such yeast host cell, and further the use of the specific nucleic acid for the secretion of a POT from a host cell and/or to increase the secretion of a POT from a host cell.

Abstract: A recombinant methanol utilization pathway deficient methylotrophic yeast (Mut-) host cell which is engineered: a) by one or more genetic modifications to reduce expression of a first and a second endogenous gene compared to the host cell prior to said one or more genetic modifications, wherein i. the first endogenous gene encodes alcohol oxidase 1 (AOX1) comprising the amino acid sequence identified as SEQ ID NO:1 or a homologue thereof, and ii. the second endogenous gene encodes alcohol oxidase 2 (AOX2) comprising the amino acid sequence identified as SEQ ID NO:3 or a homologue thereof, and b) by one or more genetic modifications to increase expression of an alcohol dehydrogenase (ADH2) gene compared to the host cell prior to said one or more genetic modifications, wherein the ADH2 gene encodes an alcohol dehydrogenase (ADH2).

Abstract: A recombinant host cell comprising an endogenous gene encoding a FLO8 protein comprising the amino acid sequence identified as SEQ ID NO:1 or a homologue thereof, which host cell is engineered by one or more genetic modifications to reduce expression of said gene compared to the host cell prior to said one or more genetic modifications, and which host cell comprises a heterologous expression cassette comprising a gene of interest (GO!) under the control of an expression cassette promoter (ECP) which ECP is repressible by a non-methanol carbon source, and a method of producing a protein of interest using said recombinant host cell.

Abstract: The invention relates to an isolated nucleic acid sequence comprising a promoter, which is a native sequence of Pichia pastoris comprising the nucleic acid sequence of pCS1 of SEQ ID 1, or a functionally active variant thereof which is a size variant, a mutant or hybrid of SEQ ID 1, or a combination thereof, expression constructs and recombinant host cells comprising the promoter, and a method of producing a protein of interest under the control of the promoter. It further relates to a method to identify a constitutive promoter from eukaryotic cells, and an isolated nucleic acid sequence comprising a promoter which when operatively linked to a nucleotide sequence encoding a protein of interest directs the expression thereof in a host cell at an expression level that is higher than under control of the native pGAP promoter at high and low growth rates.

Abstract: The present invention is in the field of recombinant biotechnology, in particular in the field of protein expression. The invention generally relates to a method of increasing the yield of a protein of interest (POI) in a eukaryotic host cell, preferably a yeast, by overexpressing at least one polynucleotide encoding at least one transcription factor of the present invention, preferably Msn4/2. The invention relates further to a recombinant eukaryotic host cell for manufacturing a POI, wherein the host cell is engineered to overexpress at least one polynucleotide encoding at least one transcription factor as well as the use of the host cell for manufacturing a POI.

Abstract: The present invention is in the field of recombinant biotechnology, in particular in the field of protein expression. The invention generally relates to a method of expressing a protein of interest (POI) from a host cell. The invention relates particularly to improving a host cell's capacity to express and/or secrete a protein of interest and use of the host cell for protein expression. The invention also relates to cell culture technology, and more specifically to culturing cells to produce desired molecules for medical purposes or food products.

Abstract: An isolated and/or artificial pG1-x promoter, which is a functional variant of the carbon source regulatable pG1 promoter of Pichia pastoris identified by SEQ ID 1, which pG1-x promoter consists of or comprises at least a part of SEQ ID 1 with a length of at least 293 bp, characterized by the following promoter regions: a) at least one core regulatory region comprising the nucleotide sequences SEQ ID 2 and SEQ ID 3; and b) a non-core regulatory region, which is any region within the pG1-x promoter sequence other than the core regulatory region; wherein the pG1-x promoter comprises at least one mutation in any of the promoter regions and a sequence identity of at least 80% in SEQ ID 2 and SEQ ID 3, and a sequence identity of at least 50% in any region other than SEQ ID 2 or SEQ ID 3; and further wherein the pG1-x promoter is characterized by the same or an increased promoter strength and induction ratio as compared to the pG1 promoter, wherein the promoter strength is at least 1.

Abstract: An isolated and/or artificial pG1-x promoter, which is a functional variant of the carbon source regulatable pG1 promoter of Pichia pastoris identified by SEQ ID 1, which pG1-x promoter consists of or comprises at least a part of SEQ ID 1 with a length of at least 293 bp, characterized by the following promoter regions: a) at least one core regulatory region comprising the nucleotide sequences SEQ ID 2 and SEQ ID 3; and b) a non-core regulatory region, which is any region within the pG1-x promoter sequence other than the core regulatory region; wherein the pG1-x promoter comprises at least one mutation in any of the promoter regions and a sequence identity of at least 80% in SEQ ID 2 and SEQ ID 3, and a sequence identity of at least 50% in any region other than SEQ ID 2 or SEQ ID 3; and further wherein the pG1-x promoter is characterized by the same or an increased promoter strength and induction ratio as compared to the pG1 promoter, wherein the promoter strength is at least 1.

Abstract: A yeast comprising a nucleotide sequence expression system expressing a synthetic Calvin cycle comprising heterologous genes, which include at least a) a gene encoding an enzyme from the class of the ribulose-bisphosphate carboxylases (EC number: 4.1.1.39) (RuBisCO gene); and b) a gene encoding an enzyme from the class of the ribulose phosphate kinases (EC number: 2.7.1.19) (PRK gene), which is expressing; wherein the yeast optionally comprises a heterologous expression construct expressing a gene of interest (GOI) and/or wherein each of said RuBisCO gene and said PRK gene, is fused with a nucleotide sequence encoding a peroxisomal targeting signal (PTS).

Abstract: The present invention is in the field of recombinant biotechnology, in particular in the field of protein expression. The invention generally relates to a method of expressing a protein of interest (POI) from a host cell. The invention relates particularly to improving a host cell's capacity to express and/or secrete a protein of interest and use of the host cell for protein expression. The invention also relates to cell culture technology, and more specifically to culturing cells to produce desired molecules for medical purposes or food products.

Abstract: The invention relates to an isolated nucleic acid sequence comprising a promoter, which is a native sequence of Pichia pastoris comprising the nucleic acid sequence of pCS1 of SEQ ID 1, or a functionally active variant thereof which is a size variant, a mutant or hybrid of SEQ ID 1, or a combination thereof, expression constructs and recombinant host cells comprising the promoter, and a method of producing a protein of interest under the control of the promoter. It further relates to a method to identify a constitutive promoter from eukaryotic cells, and an isolated nucleic acid sequence comprising a promoter which when operatively linked to a nucleotide sequence encoding a protein of interest directs the expression thereof in a host cell at an expression level that is higher than under control of the native pGAP promoter at high and low growth rates.

Abstract: The invention relates to an isolated nucleic acid sequence comprising a promoter, which is a native sequence of Pichia pastoris comprising the nucleic acid sequence of pCS1 of SEQ ID 1, or a functionally active variant thereof which is a size variant, a mutant or hybrid of SEQ ID 1, or a combination thereof, expression constructs and recombinant host cells comprising the promoter, and a method of producing a protein of interest under the control of the promoter. It further relates to a method to identify a constitutive promoter from eukaryotic cells, and an isolated nucleic acid sequence comprising a promoter which when operatively linked to a nucleotide sequence encoding a protein of interest directs the expression thereof in a host cell at an expression level that is higher than under control of the native pGAP promoter at high and low growth rates.

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: 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 or 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.