title: Characterization of Viral Capsid Protein Self-Assembly around Short Single-Stranded RNA.
abstract: For many viruses, the packaging of a single-stranded RNA (ss-RNA) genome is spontaneous, driven by capsid protein-capsid protein (CP) and CP-RNA interactions. Furthermore, for some multipartite ss-RNA viruses, copackaging of two or more RNA molecules is a common strategy. Here we focus on RNA copackaging in vitro by using cowpea chlorotic mottle virus (CCMV) CP and an RNA molecule that is short (500 nucleotides (nts)) compared to the lengths (≈3000 nts) packaged in wild-type virions. We show that the degree of cooperativity of virus assembly depends not only on the relative strength of the CP-CP and CP-RNA interactions but also on the RNA being short: a 500-nt RNA molecule cannot form a capsid by itself, so its packaging requires the aggregation of multiple CP-RNA complexes. By using fluorescence correlation spectroscopy (FCS), we show that at neutral pH and sufficiently low concentrations RNA and CP form complexes that are smaller than the wild-type capsid and that four 500-nt RNAs are packaged into virus-like particles (VLPs) only upon lowering the pH. Further, a variety of bulk-solution techniques confirm that fully ordered VLPs are formed only upon acidification. On the basis of these results, we argue that the observed high degree of cooperativity involves equilibrium between multiple CP/RNA complexes.
authors.affiliation: Department of Chemistry and Biochemistry, University of California , Los Angeles, California 90095, United States.
authors.forename: Mauricio
authors.initials: M
authors.lastname: Comas-Garcia
authors.affiliation: Department of Chemistry and Biochemistry, University of California , Los Angeles, California 90095, United States.
authors.forename: Rees F
authors.initials: RF
authors.lastname: Garmann
authors.affiliation: Department of Chemistry and Biochemistry, University of California , Los Angeles, California 90095, United States.|Department of Physical Chemistry, The Hebrew University , Jerusalem 91904, Israel.
authors.forename: Surendra W
authors.initials: SW
authors.lastname: Singaram
authors.affiliation: Department of Physical Chemistry, The Hebrew University , Jerusalem 91904, Israel.
authors.forename: Avinoam
authors.initials: A
authors.lastname: Ben-Shaul
authors.affiliation: Department of Chemistry and Biochemistry, University of California , Los Angeles, California 90095, United States.
authors.forename: Charles M
authors.initials: CM
authors.lastname: Knobler
authors.affiliation: Department of Chemistry and Biochemistry, University of California , Los Angeles, California 90095, United States.|California NanoSystems Institute, University of California , Los Angeles, California 90095, United States.
authors.forename: William M
authors.initials: WM
authors.lastname: Gelbart