Assembly of wild-type and chimeric influenza virus-like particles (VLPs)
Influenza virus-like particles (VLPs) comprising the structural proteins HA, NA, M1 and M2 are described. VLPs are also generated containing M1 alone, as are VLPs with M1 and any one or two of HA, NA and M2. VLPs with HA from one influenza subtype and NA from a different influenza subtype are also described, as are VLPs in which a portion or all of HA or NA is replaced by a heterologous moiety not produced by influenza virus, so as to comprise chimeric VLPs.
1. A method of production of influenza virus-like particles (VLPs), said method comprising the steps of:
(i) constructing a recombinant DNA molecule encoding a protein consisting of an influenza virus matrix structural protein (M1);
(ii) transfecting, infecting or otherwise transforming a suitable host cell with said recombinant DNA molecule, culturing the host cell under conditions which permit the expression of said M1 structural protein, so that VLPs are assembled within the cells after expression of the M1 structural protein; and
(iii) purifying the VLPs from the culture supernatant,
wherein the method of production is in the absence of an influenza ribonucleoprotein complex (RNP), and
wherein the method does not comprise constructing one or more recombinant DNA molecules encoding any one of the influenza virus structural proteins selected from the group consisting of hemagglutinin (HA), neuraminidase (NA) and the spliced product of M1 mRNA (M2).
2. A method of production of influenza virus-like particles (VLPs), said method comprising the steps of:
(i) constructing a first recombinant DNA molecule encoding a protein consisting of an influenza virus matrix structural protein (M1);
(ii) transfecting, infecting or otherwise transforming a suitable host cell with said first recombinant DNA molecule, culturing the host cell under conditions which permit the expression of said M1 structural protein, so that VLPs are assembled within the cells after expression of the M1 structural protein; and co-transfecting, co-infecting or co-transforming the host cell with a second recombinant DNA molecule encoding a protein consisting of an influenza nucleocapsid protein (NP), so that NP is co-localized with the M1 structural protein, and;
(iii) purifying the VLPs from the culture supernatant,
wherein the method of production is in the absence of an influenza ribonucleoprotein complex (RNP), and
wherein the method does not comprise constructing one or more recombinant DNA molecules encoding any one of the influenza virus structural proteins selected from the group consisting of hemagglutinin (HA), neuraminidase (NA) and the spliced product of M1 mRNA (M2).