Plasmids and methods for peptide display and affinity-selection on virus-like particles of RNA bacteriophages

摘要

The present invention relates to a system and method for controlling peptide display valency on virus-like particles (VLPs), especially including MS2 VLPs. In this method, large amounts of wild-type and low quantities of single-chain dimer coat proteins may be produced from a single RNA. Valency is controlled in immunogen (vaccine) production by providing a system that allows the production of large amounts of wild-type and low quantities of single-chain dimer coating proteins from a single RNA, allowing facile adjustment of display valency levels on VLPs, especially MS2 VLPS over a wide range, from few than one—on average—to as many as ninety per particle. This facilitates the production of immunogens and vaccines, including VLPs exhibiting low valency. Nucleic acid constructs useful in the expression of virus-like particles are disclosed, comprised of a coat polypeptide of MS2 modified by insertion of a heterologous peptide, wherein the heterologous peptide is displayed on the virus-like particle and encapsidates MS2 niRNA. Nucleic acid constructs are also disclosed which are useful in the expression of virus-like particles comprised of a coat polypeptide of PP7 modified by insertion of a heterologous peptide, wherein the heterologous peptide is displayed on the virus-like particle and encapsidates PP7 mRNA.

主权项

1. A nucleic acid construct comprising:
(a) a bacterial or bacteriophage promoter operably associated with a coding sequence of an RNA bacteriophage single chain coat polypeptide dimer, the RNA bacteriophage single chain coat polypeptide dimer comprising (i) a first upstream coat polypeptide coding sequence, (ii) a second downstream coat polypeptide coding sequence comprising a nucleic acid sequence insert encoding a heterologous peptide, and (iii) a translation stop codon between the upstream and downstream coat polypeptide encoding sequences; (b) a replication origin for replication in a prokaryote; and (c) an antibiotic resistance gene.