In vitro translation of luciferase mRNAs harboring mutated HIV-2 5′ leaders. (A) A schematic of the four mRNA constructs used in the S35 in vitro translation assays. The mutated nucleotides are in bold lowercase letters. (B) A schematic of the three HIV-2 Gag-luciferase fusion protein isoforms that are synthesized in vitro. The 50-, 42-, and 37-kDa isoforms are synthesized when translation initiates from methionine codons located at positions 546, 744, and 897, respectively. (C) Duplicate S35 in vitro translation assays of the wild-type, C-box119, G-box119, and CLS-119 mRNAs. (Lane 1) A mock where no mRNA was added to the translation reaction. RNAs used in each translation reaction are indicated below each lane. (D) Quantification of protein isoform expression levels. The RNAs used are indicated on the x-axis, along with the protein isoforms produced and quantified. The level of protein isoforms in arbitrary units is indicated on the y-axis. The y-axis error bars represent the standard error of the duplicate experiments.
