Method for producing dimerized polypeptide fusions

A method for producing a secreted, biologically active dimerized polypeptide fusion, comprising:

• introducing into a eukaryotic host cell a first DNA construct comprising a transcriptional promoter operatively linked to a first secretory signal sequence followed downstream by and in proper reading frame with a first DNA sequence encoding a non-immunoglobulin polypeptide requiring dimerization for biological activity joined to an immunoglobulin light chain constant region;
• introducing into said host cell a second DNA construct comprising a transcriptional promoter operatively linked to a second secretory signal sequence followed downstream by and in proper reading frame with a second DNA sequence encoding an immunoglobulin heavy chain constant region domain selected from the group consisting of CH1, CH2, CH3, and CH4;
• growing said host cell in an appropriate growth medium under physiological conditions to allow the secretion of a biologically active dimerized polypeptide fusion encoded by said first and second DNA sequences;
• and
• isolating said biologically active dimerized polypeptide fusion from said host cell.

A method for producing a secreted, biologically active dimerized polypeptide fusion, comprising:

• introducing into a eukaryotic host cell a first DNA construct comprising a transcriptional promoter operatively linked to a first secretory signal sequence followed downstream by and in proper reading frame with a first DNA sequence encoding a non-immunoglobulin polypeptide requiring dimerization for biological activity joined to an immunoglobulin heavy chain constant region domain selected from the group consisting of CH1, CH2, CH3, and CH4;
• introducing into said host cell a second DNA construct comprising a transcriptional promoter operatively linked to a second secretory signal sequence followed downstream by and in proper reading frame with a second DNA sequence encoding an immunoglobulin light chain constant region;
• growing said host cell in an appropriate growth medium under physiological conditions to allow the secretion of a biologically active dimerized polypeptide fusion encoded by said first and second DNA sequences;
• and
• isolating said biologically active dimerized polypeptide fusion from said host cell.

A method for producing a secreted receptor analog, comprising:

• introducing into a eukaryotic host cell a DNA construct comprising a transcriptional promoter operatively linked to at least one secretory signal sequence followed downstream by and in proper reading frame with a DNA sequence encoding a ligand-binding domain of a receptor requiring dimerization for biological activity joined to a dimerizing protein;
• growing said host cell in an appropriate growth medium under physiological conditions to allow the secretion of a receptor analog encoded by said DNA sequence;
• and
• isolating said receptor analog from said host cell.

A method for producing a secreted PDGF receptor analog, comprising:

• introducing into a eukaryotic host cell a DNA construct comprising a transcriptional promoter operatively linked to a secretory signal sequence followed downstream in proper reading frame by a DNA sequence encoding a ligand-binding domain of a PDGF receptor;
• growing said host cell in an appropriate growth medium under physiological conditions to allow the secretion of a PDGF receptor analog encoded by said DNA sequence;
• and
• isolating said PDGF receptor analog from said host cell.

A method for producing a secreted PDGF receptor analog, comprising:

• introducing into a cultured rodent myeloma cell a DNA construct comprising a transcriptional promoter operatively linked to a PDGF receptor signal sequence followed downstream by and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIG. 11 (Sequence ID Numbers 35 and 36) from glutamine, number 24, to glutamic acid, number 524, joined to a dimerizing protein, wherein said dimerizing protein is an immunoglobulin constant region selected from the group consisting of CH1, CH2, CH3, CH4 and Cκ joined to an immunoglobulin hinge region;
• growing said cultured rodent myeloma cell in an appropriate growth medium under physiological conditions to allow the secretion of a PDGF receptor analog encoded by said DNA sequence;
• and
• isolating the PDGF receptor analog from said cultured rodent myeloma cell.

A method for producing a secreted PDGF receptor analog, comprising:

• introducing into a cultured rodent myeloma cell a first DNA construct comprising a transcriptional promoter operatively linked to a PDGF receptor signal sequence followed downstream by and in proper reading frame with a first DNA sequence encoding the amino acid sequence of FIG. 1 (Sequence ID Numbers 1 and 2) from isoleucine, number 29, to lysine, number 531 joined to an immunoglobulin light chain constant region;
• introducing into said cultured rodent myeloma cell a second DNA construct comprising a transcriptional promoter operatively linked to a PDGF receptor signal sequence followed downstream by and in proper reading frame with a second DNA sequence encoding the amino acid sequence of FIG. 1 (sequence ID Numbers 1 and 2) from isoleucine number 29, to lysine, number 531 joined to an immunoglobulin heavy chain constant region domain selected from the group consisting of CH1, CH2, CH3, and CH4 joined to an immunoglobulin hinge region;
• growing said cultured rodent myeloma cell in an appropriate growth medium under physiological conditions to allow the secretion of a PDGF receptor analog encoded by said first and second DNA sequences;
• and
• isolating said PDGF receptor analog from said cultured rodent myeloma cell.

A method for producing a secreted PDGF receptor analog, comprising:

• introducing into a cultured rodent myeloma cell a first DNA construct comprising a mouse VH promoter operatively linked to a PDGF receptor signal sequence followed downstream of and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIG. 11 (Sequence ID Numbers 35 and 36) from glutamine, number 24 to glutamic acid, number 524, joined to an immunoglobulin heavy chain constant region domain selected from the group consisting of CH1, CH2, CH3 and CH4 joined to an immunoglobulin hinge region;
• introducing into said cultured rodent myeloma cell a second DNA construct comprising a mouse Vκ promoter operatively linked to a PDGF receptor signal sequence followed downstream of and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIG. 11 (Sequence ID Numbers 35 and 36) from glutamine, number 24 to glutamic acid, number 524, joined to an immunoglobulin light chain constant region;
• growing said cultured rodent myeloma cell in an appropriate growth medium under physiological conditions to allow the secretion of a PDGF receptor analog encoded by said first and second DNA sequences;
• and
• isolating said PDGF receptor analog from said cultured myeloma cell.

A method for producing a secreted PDGF receptor analog, comprising:

• introducing into a cultured rodent myeloma cell a first DNA construct comprising a mouse VH promoter operatively linked to a PDGF receptor signal sequence followed downstream of and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIG. 1 (Sequence ID Numbers 1 and 2) from isoleucine number 29, to lysine, number 531, joined to an immunoglobulin heavy chain constant region domain selected from the group consisting of CH1, CH2, CH3 and CH4 joined to an immunoglobulin hinge region;
• introducing into said cultured rodent myeloma cell a second DNA construct comprising a mouse Vκ promoter operatively linked to a PDGF receptor signal sequence followed downstream of and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIG. 11 (Sequence ID Numbers 35 and 36) from glutamine, number 24 to glutamic acid, number 524, joined to an immunoglobulin light chain constant region;
• growing said cultured rodent myeloma cell in an appropriate growth medium under physiological conditions to allow the secretion of a PDGF receptor analog encoded by said first and second DNA sequences;
• and
• isolating said PDGF receptor analog from said cultured myeloma cell.

A method for producing a secreted PDGF receptor analog, comprising:

• introducing into a cultured rodent myeloma cell a first DNA construct comprising a mouse VH promoter operatively linked to a PDGF receptor signal sequence followed downstream of and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIG. 11 (Sequence ID Numbers 35 and 36) from glutamine, number 24 to glutamic acid, number 524, joined to an immunoglobulin heavy chain constant region domain selected from the group consisting of CH1, CH2, CH3 and CH4 joined to an immunoglobulin hinge region;
• introducing into said cultured rodent myeloma cell a second DNA construct comprising a mouse Vκ promoter operatively linked to a PDGF receptor signal sequence followed downstream of and in proper reading frame with a DNA sequence encoding the amino acid sequence of FIG. 1 (Sequence ID Numbers 1 and 2) from isoleucine number 29, to lysine, number 531, joined to an immunoglobulin light chain constant region;
• growing said cultured rodent myeloma cell in an appropriate growth medium under physiological conditions to allow the secretion of a PDGF receptor analog encoded by said first and second DNA sequences;
• and
• isolating said PDGF receptor analog from said cultured myeloma cell.

A method for purifying PDGF or an isoform thereof from a sample, comprising:

• immobilizing a polypeptide comprising a PDGF receptor analog fused to a dimerizing protein on a substrate;
• contacting a sample comprising PDGF or an isoform thereof with the immobilized polypeptide under conditions such that the PDGF or isoform thereof binds to the polypeptide;
• and
• eluting the PDGF or isoform thereof from the polypeptide.