Browsing by keyword "Virions"
Now showing items 1-2 of 2
-
Conformational changes in the Ebola virus membrane fusion machine induced by pH, Ca2+, and receptor bindingThe Ebola virus (EBOV) envelope glycoprotein (GP) is a membrane fusion machine required for virus entry into cells. Following endocytosis of EBOV, the GP1 domain is cleaved by cellular cathepsins in acidic endosomes, removing the glycan cap and exposing a binding site for the Niemann-Pick C1 (NPC1) receptor. NPC1 binding to cleaved GP1 is required for entry. How this interaction translates to GP2 domain-mediated fusion of viral and endosomal membranes is not known. Here, using a bulk fluorescence dequenching assay and single-molecule Forster resonance energy transfer (smFRET)-imaging, we found that acidic pH, Ca2+, and NPC1 binding synergistically induce conformational changes in GP2 and permit virus-liposome lipid mixing. Acidic pH and Ca2+ shifted the GP2 conformational equilibrium in favor of an intermediate state primed for NPC1 binding. Glycan cap cleavage on GP1 enabled GP2 to transition from a reversible intermediate to an irreversible conformation, suggestive of the postfusion 6-helix bundle; NPC1 binding further promoted transition to the irreversible conformation. Thus, the glycan cap of GP1 may allosterically protect against inactivation of EBOV by premature triggering of GP2.
-
Select gp120 V2 domain specific antibodies derived from HIV and SIV infection and vaccination inhibit gp120 binding to alpha4beta7The GI tract is preferentially targeted during acute/early HIV-1 infection. Consequent damage to the gut plays a central role in HIV pathogenesis. The basis for preferential targeting of gut tissues is not well defined. Recombinant proteins and synthetic peptides derived from HIV and SIV gp120 bind directly to integrin alpha4beta7, a gut-homing receptor. Using both cell-surface expressed alpha4beta7 and a soluble alpha4beta7 heterodimer we demonstrate that its specific affinity for gp120 is similar to its affinity for MAdCAM (its natural ligand). The gp120 V2 domain preferentially engages extended forms of alpha4beta7 in a cation -sensitive manner and is inhibited by soluble MAdCAM. Thus, V2 mimics MAdCAM in the way that it binds to alpha4beta7, providing HIV a potential mechanism to discriminate between functionally distinct subsets of lymphocytes, including those with gut-homing potential. Furthermore, alpha4beta7 antagonists developed for the treatment of inflammatory bowel diseases, block V2 binding to alpha4beta7. A 15-amino acid V2 -derived peptide is sufficient to mediate binding to alpha4beta7. It includes the canonical LDV/I alpha4beta7 binding site, a cryptic epitope that lies 7-9 amino acids amino terminal to the LDV/I, and residues K169 and I181. These two residues were identified in a sieve analysis of the RV144 vaccine trial as sites of vaccine -mediated immune pressure. HIV and SIV V2 mAbs elicited by both vaccination and infection that recognize this peptide block V2-alpha4beta7 interactions. These mAbs recognize conformations absent from the beta- barrel presented in a stabilized HIV SOSIP gp120/41 trimer. The mimicry of MAdCAM-alpha4beta7 interactions by V2 may influence early events in HIV infection, particularly the rapid seeding of gut tissues, and supports the view that HIV replication in gut tissue is a central feature of HIV pathogenesis.
