Figure 1—figure supplement 1. | The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA

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The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA

Figure 1—figure supplement 1.

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University of California, San Francisco, United States
Figure 1—figure supplement 1.
Download figureOpen in new tabFigure 1—figure supplement 1. Assessment of the stoichiometry of Crm1, RanGTP, Rev, and the RRE.

(A) Size-exclusion chromatograms of the reconstituted complex (top), Rev-RRE (second from top), Crm1 (third from top), or RanGTP (bottom) with indicated molecular masses (MW) determined by multi-angle laser light scattering after size exclusion chromatography. The RRE has a mass of 80.172 kD and each Rev subunit has a mass of 13.179 kD, so the total 159 kD mass of the complex corresponds to six Rev subunits per RRE. (BC) Determination of stoichiometry based on protein standards. (B) Size-exclusion chromatograms from Figure 2E of human or murine Crm1 assembled with Rev-RRE and RanGTP with fractions used for determining stoichiometry marked with an asterisk. (C) An SDS-PAGE gel stained with Sypro-Ruby showing a dilution series of protein standards, which were quantified by amino acid analysis, and indicated fractions from the chromatograms in panel B. (D) Quantification of (C) and the calculated stoichiometry of the complex. The RRE concentration was determined by UV spectrometry and used to determine the molar ratios for proteins. These data indicate that this reconstitution had slightly sub-stoichiometric amounts of Crm1 and RanGTPand six subunits of Rev bound to the RRE, with a small population having slightly more Rev bound.

DOI: http://dx.doi.org/10.7554/eLife.04121.004