The molecular basis for dysfunctional bacterial lipopolysaccharide-mediated immune receptor activation by SARS-CoV-2 spike

SARS-CoV-2 has caused hundreds of millions of COVID-19 infections worldwide. While COVID-19 presents with various clinical manifestations, severe COVID-19 disease causes dysregulated host immune reactions that trigger onset of sepsis and acute respiratory distress syndrome. Accumulating evidence suggests that lipopolysaccharide (LPS) derived from the outer membranes of Gram-negative bacteria plays an instrumental role in the progression of such inflammatory states. Patients with pre-existing conditions such as diabetes, hypertension, and obesity are at higher risk to develop severe COVID-19 disease and tend to have significantly elevated blood LPS levels. LPS serves as a signal of bacterial infection upon recognition by Toll-like receptor (TLR4) of the innate immune system, but this can also result in over-amplified immune reactions and sepsis. Here, we present the results of molecular simulations and free-energy calculations, supported by in vitro and in vivo assays and hydrogen-deuterium exchange mass spectrometry experiments, which reveal the molecular mechanism by which the envelope spike glycoprotein of SARS-CoV-2 augments hyperinflammation by acting as a conduit in the TLR4 pathway. LPS was found to bind to several conserved pockets on spike across S1 and S2 subunits. S1 affinity for LPS was comparable to that of CD14, a co-receptor used by immune cells to transfer LPS to TLR4. Cell-based assays and reporter mice experiments showed that low concentrations of spike and LPS synergistically induce a strong pro-inflammatory response, thus pinpointing spike's capacity to “boost” innate immune activation. Finally, the loss of a high-affinity binding site in the Omicron spike led to a reduction of its “boosting” capacity, which may translate to the less severe inflammation observed in patients infected with this variant. Collectively, our findings highlight the potential impact of elevated LPS levels and Gram-negative bacterial coinfections in severe COVID-19 complications.