Fold promoters and their use for the production and stabilization of polypeptides
AssigneeMax-Planck-Gesellschaft zur Forderung der Wissenschaften e.V.
InventorsDirk GÖRLICH, Metin AKSU, Thomas GÜTTLER, Oleh RYMARENKO, Renate REES, Kathrin GREGOR, Waltraud TAXER
Life of this patent
Estimating the term…
Abstract
The present invention relates to the recombinant production of a protein of interest in a prokaryotic host cell or eukaryotic host cell wherein the protein of interest is obtained in a correctly folded and stable form. The protein of interest may be a difficult-to-make polypeptide for use as a vaccine or a pharmaceutical. The protein of interest is co-expressed with or fused to a ‘fold promoter’, which may be a VHH antibody recognizing the said protein.
Claims15
A method for the recombinant production of a polypeptide of interest in a host cell, e.g. a prokaryotic host cell, or a eukaryotic host cell comprising the steps:
- (a) providing a host cell comprising (i) a nucleic acid molecule encoding a fusion polypeptide comprising the polypeptide of interest and a fold promoter, or (ii) a first nucleic acid molecule encoding a polypeptide of interest and a second nucleic acid molecule encoding a fold promoter for the polypeptide of interest, wherein the fold promoter is a VHH antibody directed against the polypeptide of interest;
- and
- (b) cultivating the host cell under conditions where (i) the fusion polypeptide comprising the polypeptide of interest and the fold promoter is expressed or (ii) the polypeptide of interest and the fold promoter are co-expressed, and
- (c) obtaining the polypeptide of interest in correctly folded form from the host cell.
A nucleic molecule encoding a fusion polypeptide comprising a polypeptide of interest and a fold promoter for the polypeptide of interest, wherein the fold promoter is a VVH antibody directed against the polypeptide of interest.
A set of nucleic molecules comprising a first nucleic acid molecule encoding a polypeptide of interest and a second nucleic acid molecule encoding a fold promoter for the polypeptide of interest, wherein the fold promoter is a VVH antibody directed against the polypeptide of interest.
An RBD of the SARS-CoV-2 spike protein, particularly having an amino acid sequence as shown in SEQ ID NO. 1 or SEQ ID NO. 55 or a sequence identity of at least 80%, at least 90% or at least 95% thereto, in correctly folded form, particularly produced in a prokaryotic host cell, which binds to the non-competing VHH antibody Re5D06 having an amino acid sequence as shown in SEQ ID NO. 34 or 42.
A non-covalent complex comprising an RBD of a coronavirus spike protein, particularly the RBD of the SARS-CoV-2 spike protein, more particularly having an amino acid sequence as shown in SEQ ID NO. 1 or SEQ ID NO. 55 or a sequence identity of at least 80%, at least 90% or at least 95% thereto, and a fold-enhancing VHH antibody.
A covalent fusion protein comprising an RBD of a coronavirus spike protein, particularly the SARS-CoV-2 spike protein, more particularly having an amino acid sequence as shown in SEQ ID NO. 1 or SEQ ID NO. 55 or a sequence identity of at least 80%, at least 90% or at least 95% thereto, and a fold-enhancing VHH antibody
A polypeptide for use as a vaccine, which is attached to a heterologous amino acid sequence having a length of at least about 5 amino acids and up to about 100 amino acids, and which has a negative net charge of at least −5 and particularly a content of at least about 15%, at least about 20% or least about 25% of negatively charged amino acids selected from Asp and Glu.
Dependencies are inferred from the claim text.