FIG 1.
Ebola virus delta peptide. (A) EBOV glycoprotein editing. The GP gene can produce multiple versions of the protein due to differential RNA editing at a conserved site containing seven uridine residues. All versions share the first 295 amino acid residues. Full-length GP, which is 676 amino acids, is produced from mRNAs containing an additional nontemplated adenosine. Full-length GP is converted to GP1 and GP2 by cleavage at a furin-like protease-sensitive site and is membrane anchored by a C-terminal membrane-spanning helix domain. sGP, which is produced from the unedited RNA transcript, contains only the first 364 to 372 amino acids of the GP1 domain. The delta peptide, which is the frame-shifted C terminus of sGP, is cleaved from sGP in cells at an alternate furin-like protease site created by the RNA editing. (B) Delta peptide sequences from Ebola virus and related filoviruses. The conserved C terminus is green, except basic residues are blue, acidic residues are red, and cysteine is yellow. Aromatic residues are underlined. In the consensus sequence, a residue letter indicates that at least 6 of the 7 sequences have the same residue, while a plus sign indicates a conserved physical chemistry. (C) HPLC chromatograms of tryptophan fluorescence of Ebola virus delta peptide incubated in human serum for 1 to 24 h at 37°C. At each time point, C-terminal biotinylated delta peptide was extracted from serum using bead-immobilized streptavidin and released with 1 mM free biotin after extensive washing. The released peptide was eluted from a reverse-phase C18 column developed in water/acetonitrile. The data are not corrected for the number of tryptophans; therefore, the 8- and 24-h intensities per molecule are halved by the loss of tryptophan 18 by proteolysis. (D) Serum stability of E40ox after incubation with 10% human serum at 37°C for the indicated times. The bead wash solution was subjected to HPLC and MALDI-time of flight (TOF) mass spectrometry, where only fragments of E40 containing the C terminus were observed. The blue star marks the full-length peptide, and the red stars mark the 15-residue fragment that persists for at least 24 h. (E) Delta peptide fragments observed after incubation with 10% human serum for 1, 8, and 24 h. The blue star indicates full-length E40ox, and the red stars and arrows indicate the stable 15-residue C-terminal fragment. The black arrows represent the most prevalent fragments, and the gray arrows represent less abundant fragments that were detected by mass spectroscopy.