title: Facile production and rapid purification of functional recombinant Qβ replicase heterotetramer complex.
abstract: We describe an improved method for the production of recombinant Qβ replicase heterotetramer. The successful expression of the soluble Qβ RNA polymerase complex depends on the EF-Ts and EF-Tu subunits being co-expressed prior to β-subunit expression. Efficient co-expression requires two different inducible operons to co-ordinate the expression of the heterotrimer. The complete heterotetramer enzyme complex is achieved by production of the recombinant S1-subunit of Qβ replicase in a separate host. This approach represents a facile way for producing and purifying large amounts of soluble and active recombinant Qβ replicase tetramer without the necessity of a His-tag for purification.
authors.affiliation: Department of Chemistry and Biomolecular Sciences, and Environmental Biotechnology CRC, Macquarie University, North Ryde, 2109 Sydney, NSW, Australia.
authors.forename: Karthikeyan
authors.initials: K
authors.lastname: Gunasekaran
authors.forename: Peter L
authors.initials: PL
authors.lastname: Bergquist
authors.forename: Anwar
authors.initials: A
authors.lastname: Sunna
mesh_terms: D019281:Dimerization; D004926:Escherichia coli / Q000502:physiology*; D015202:Protein Engineering / Q000379:methods*; D011777:Q beta Replicase / Q000096:biosynthesis* / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification*
chemical_list: D011994:Recombinant Proteins; D011777:Q beta Replicase