Thermal Stability of RNA Phage Virus-Like Particles Displaying Foreign Peptides

Abstract

BackgroundTo be useful for genetic display of foreign peptides a viral coat protein must tolerate peptide insertions without major disruption of subunit folding and capsid assembly. The folding of the coat protein of RNA phage MS2 does not normally tolerate insertions in its AB-loop, but an engineered single-chain dimer readily accepts them as long as they are restricted to one of its two halves.ResultsHere we characterize the effects of peptide insertions on the thermal stabilities of MS2 virus-like particles (VLPs) displaying a variety of different peptides in one AB-loop of the coat protein single-chain dimer. These particles typically denature at temperatures around 5-10°C lower than unmodified VLPs. Even so, they are generally stable up to about 50°C. VLPs of the related RNA phage PP7 are cross-linked with intersubunit disulfide bonds and are therefore significantly more stable. An AB-loop insertion also reduces the stability of PP7 VLPs, but they only begin to denature above about 70°C.ConclusionsVLPs assembled from MS2 single-chain dimer coat proteins with peptide insertions in one of their AB-loops are somewhat less stable than the wild-type particle, but still resist heating up to about 50°C. Because they possess disulfide cross-links, PP7-derived VLPs provide an alternate platform with even higher stability.

RETRACTED ARTICLE: Cell-specific delivery of messenger RNA and microRNA by recombinant MS2 virus-like particles carrying cell-penetrating peptide

“…This recombinant MS2 VLPs and CPP-based RNA delivery system has the following characteristics: (i) easily prepared by recombinant protein technology; (ii) high yield in a yeast expression system; (iii) strong cell-penetrating ability, which enhances in vivo delivery efficiency of mRNA or microRNA and lays the foundation for targeted delivery of RNA; (iv) low cytotoxicity; and (v) high expression levels of mRNA or microRNA in mammalian cells, which may be used in therapeutics. Hence, recombinant MS2 VLPs designed in this study are an efficient, targeted delivery strategy for both peptide and RNA, due to the abilities of MS2 VLPs to package specific exogenous RNA with pac site and display peptides (Caldeira and Peabody 2011). In addition, this system holds great potential in the development of mRNA/microRNA and peptide-based targeted therapeutics for the treatment of cancer and other diseases.…”

“…Insertion of DNA oligonucleotides at this site allows the production of chimeric MS2 coat proteins, which have foreign peptide sequences in the central part of the hairpin (Mastico et al 1993). Most importantly, the displayed peptides are highly immunogenic (Peabody et al 2008), and the MS2 VLPs with inserted peptides are heat-resistant up to about 50°C (Caldeira and Peabody 2011). To date, MS2 VLPs have been used as peptide vaccine carriers for HIV (Peabody et al 2008), human papillomavirus (Tumban et al 2012), influenza virus (Mastico et al 1993), hypersensitivity (Mastico et al 1993), and malaria (Heal et al 1999).…”

Engineering virus-like particles as vaccine platforms

“…In the case of bacteriophage MS2 VLPs, foreign peptides displayed in a surface-exposed loop (the AB-loop) on the wild-type viral coat protein are rarely compatible with protein folding and VLP assembly (Fig. 1) [11]. However, the physical proximity of N- and C- termini of subunits of the coat protein dimer facilitated their genetic fusion into a so-called single-chain dimer [12].…”

Engineered Mutations Change the Structure and Stability of a Virus-Like Particle

“…7–9 They can be highly stable toward extremes of temperature, 10,11 pH, and variations in solvent composition, 12 and thus are friendly toward chemical modification, while retaining many advantageous characteristics of biological molecules. VLPs have well-controlled composition and structure, 13 are biocompatible, 14,15 and can be mutated 16–18 and/or evolved.…”