TY - JOUR
T1 - Marburg virus glycoprotein GP2
T2 - PH-dependent stability of the ectodomain α-helical bundle
AU - Harrison, Joseph S.
AU - Koellhoffer, Jayne F.
AU - Chandran, Kartik
AU - Lai, Jonathan R.
PY - 2012/3/27
Y1 - 2012/3/27
N2 - Marburg virus (MARV) and Ebola virus (EBOV) constitute the family Filoviridae of enveloped viruses (filoviruses) that cause severe hemorrhagic fever. Infection by MARV requires fusion between the host cell and viral membranes, a process that is mediated by the two subunits of the envelope glycoprotein, GP1 (surface subunit) and GP2 (transmembrane subunit). Upon viral attachment and uptake, it is believed that the MARV viral fusion machinery is triggered by host factors and environmental conditions found in the endosome. Next, conformational rearrangements in the GP2 ectodomain result in the formation of a highly stable six-helix bundle; this refolding event provides the energetic driving force for membrane fusion. Both GP1 and GP2 from EBOV have been extensively studied, but there is little information available for the MARV glycoproteins. Here we have expressed two variants of the MARV GP2 ectodomain in Escherichia coli and analyzed their biophysical properties. Circular dichroism indicates that the MARV GP2 ectodomain adopts an α-helical conformation, and one variant sediments as a trimer by equilibrium analytical ultracentrifugation. Denaturation studies indicate the α-helical structure is highly stable at pH 5.3 (unfolding energy, ΔGunf,H 2O, of 33.4 ± 2.5 kcal/mol and melting temperature, T m, of 75.3 ± 2.1 °C for one variant). Furthermore, we found the α-helical stability to be strongly dependent on pH, with higher stability under lower-pH conditions (T m values ranging from ∼92 °C at pH 4.0 to ∼38 °C at pH 8.0). Mutational analysis suggests two glutamic acid residues (E579 and E580) are partially responsible for this pH-dependent behavior. On the basis of these results, we hypothesize that the pH-dependent folding stability of the MARV GP2 ectodomain provides a mechanism for controlling conformational preferences such that the six-helix bundle "postfusion" state is preferred under conditions of appropriately matured endosomes.
AB - Marburg virus (MARV) and Ebola virus (EBOV) constitute the family Filoviridae of enveloped viruses (filoviruses) that cause severe hemorrhagic fever. Infection by MARV requires fusion between the host cell and viral membranes, a process that is mediated by the two subunits of the envelope glycoprotein, GP1 (surface subunit) and GP2 (transmembrane subunit). Upon viral attachment and uptake, it is believed that the MARV viral fusion machinery is triggered by host factors and environmental conditions found in the endosome. Next, conformational rearrangements in the GP2 ectodomain result in the formation of a highly stable six-helix bundle; this refolding event provides the energetic driving force for membrane fusion. Both GP1 and GP2 from EBOV have been extensively studied, but there is little information available for the MARV glycoproteins. Here we have expressed two variants of the MARV GP2 ectodomain in Escherichia coli and analyzed their biophysical properties. Circular dichroism indicates that the MARV GP2 ectodomain adopts an α-helical conformation, and one variant sediments as a trimer by equilibrium analytical ultracentrifugation. Denaturation studies indicate the α-helical structure is highly stable at pH 5.3 (unfolding energy, ΔGunf,H 2O, of 33.4 ± 2.5 kcal/mol and melting temperature, T m, of 75.3 ± 2.1 °C for one variant). Furthermore, we found the α-helical stability to be strongly dependent on pH, with higher stability under lower-pH conditions (T m values ranging from ∼92 °C at pH 4.0 to ∼38 °C at pH 8.0). Mutational analysis suggests two glutamic acid residues (E579 and E580) are partially responsible for this pH-dependent behavior. On the basis of these results, we hypothesize that the pH-dependent folding stability of the MARV GP2 ectodomain provides a mechanism for controlling conformational preferences such that the six-helix bundle "postfusion" state is preferred under conditions of appropriately matured endosomes.
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U2 - 10.1021/bi3000353
DO - 10.1021/bi3000353
M3 - Article
C2 - 22369502
AN - SCOPUS:84859180550
SN - 0006-2960
VL - 51
SP - 2515
EP - 2525
JO - Biochemistry
JF - Biochemistry
IS - 12
ER -
