title: Designed protein mimics of the Ebola virus glycoprotein GP2 α-helical bundle: stability and pH effects.
abstract: Ebola virus (EboV) belongs to the Filoviridae family of viruses that causes severe and fatal hemhorragic fever. Infection by EboV involves fusion between the virus and host cell membranes mediated by the envelope glycoprotein GP2 of the virus. Similar to the envelope glycoproteins of other viruses, the central feature of the GP2 ectodomain postfusion structure is a six-helix bundle formed by the protein's N- and C-heptad repeat regions (NHR and CHR, respectively). Folding of this six-helix bundle provides the energetic driving force for membrane fusion; in other viruses, designed agents that disrupt formation of the six-helix bundle act as potent fusion inhibitors. To interrogate determinants of EboV GP2-mediated membrane fusion, we designed model proteins that consist of the NHR and CHR segments linked by short protein linkers. Circular dichroism and gel filtration studies indicate that these proteins adopt stable α-helical folds consistent with design. Thermal denaturation indicated that the GP2 six-helix bundle is highly stable at pH 5.3 (melting temperature, T(m) , of 86.8 ± 2.0°C and van't Hoff enthalpy, ΔH(vH) , of -28.2 ± 1.0 kcal/mol) and comparable in stability to other viral membrane fusion six-helix bundles. We found that the stability of our designed α-helical bundle proteins was dependent on buffering conditions with increasing stability at lower pH. Small pH differences (5.3-6.1) had dramatic effects (ΔT(m) = 37°C) suggesting a mechanism for conformational control that is dependent on environmental pH. These results suggest a role for low pH in stabilizing six-helix bundle formation during the process of GP2-mediated viral membrane fusion.
authors: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
authors: Joseph S
authors: JS
authors: Harrison
authors: Chelsea D
authors: CD
authors: Higgins
authors: Kartik
authors: K
authors: Chandran
authors: Jonathan R
authors: JR
authors: Lai
mesh_terms: D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D002942:Circular Dichroism; D029043:Ebolavirus / Q000378:metabolism*; D006023:Glycoproteins / Q000737:chemistry* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008954:Models, Biological; D008969:Molecular Sequence Data; D055550:Protein Stability; D017433:Protein Structure, Secondary; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000378:metabolism*; D053586:Virus Internalization
chemical_list: D006023:Glycoproteins; D014759:Viral Envelope Proteins