Abstract
LDLRAD3 is a recently defined attachment and entry receptor for Venezuelan equine encephalitis virus (VEEV)1, a New World alphavirus that causes severe neurological disease in humans. Here we present near-atomic-resolution cryo-electron microscopy reconstructions of VEEV virus-like particles alone and in a complex with the ectodomains of LDLRAD3. Domain 1 of LDLRAD3 is a low-density lipoprotein receptor type-A module that binds to VEEV by wedging into a cleft created by two adjacent E2–E1 heterodimers in one trimeric spike, and engages domains A and B of E2 and the fusion loop in E1. Atomic modelling of this interface is supported by mutagenesis and anti-VEEV antibody binding competition assays. Notably, VEEV engages LDLRAD3 in a manner that is similar to the way that arthritogenic alphaviruses bind to the structurally unrelated MXRA8 receptor, but with a much smaller interface. These studies further elucidate the structural basis of alphavirus–receptor interactions, which could inform the development of therapies to mitigate infection and disease against multiple members of this family.
Highlights
Alphaviruses enter host cells through receptor-mediated endocytosis[4]
Cryo-electron micrographs of Venezuelan equine encephalitis virus (VEEV) virus-like particles (VLPs)[17] with or without bound Low-density lipoprotein receptor class A domain-containing 3 (LDLRAD3)(D1) were acquired using a 300 kV Titan Krios system equipped with a Gatan K2 detector (Extended Data Fig. 1a and Supplementary Table 1)
Two-hundred and forty molecules of LDLRAD3(D1) bound to sites on VEEV VLP (100% saturation), each one wedged into a cleft formed between two adjacent E2–E1 heterodimers
Summary
This cleft widens slightly when D1 of LDLRAD3 is bound (Supplementary Video 1). Local resolution estimation performed in RELION revealed heterogenous resolution; the capsid proteins and membrane proximal regions of the E2–E1 heterodimers were best resolved (about 4 Å) and the membrane distal regions and LDLRAD3(D1) were less-well resolved (about 5–6 Å) (Fig. 1d). To avoid under- and over-sharpening of the reconstructions by conventional global B-factor correction, post-processing was performed using DeepEMhancer[18]. This resulted in improved continuity and reduced noise in the density (Extended Data Fig. 1c). The visibly clear tracing of the carbon backbone simplified subsequent model building
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