Striking antibody evasion of SARS-CoV-2 Omicron sub-lineages BQ.1.1, XBB.1 and CH.1.1.

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Omicron BQ.1.1 and XBB.1 unprecedentedly escape broadly neutralizing antibodies elicited by prototype vaccination.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariants have seriously attacked the antibody barrier established by natural infection and/or vaccination, especially the recently emerged BQ.1.1 and XBB.1. However, crucial mechanisms underlying the virus escape and the broad neutralization remain elusive. Here, we present a panoramic analysis of broadly neutralizing activity and binding epitopes of 75 monoclonal antibodies isolated from prototype inactivated vaccinees. Nearly all neutralizing antibodies (nAbs) partly or totally lose their neutralization against BQ.1.1 and XBB.1. We report a broad nAb, VacBB-551, that effectively neutralizes all tested subvariants including BA.2.75, BQ.1.1, and XBB.1. We determine the cryoelectron microscopy (cryo-EM) structure of VacBB-551 complexed with the BA.2 spike and perform detailed functional verification to reveal the molecular basis of N460K and F486V/S mutations mediating the partial escape of BA.2.75, BQ.1.1, and XBB.1 from the neutralization of VacBB-551. Overall, BQ.1.1 and XBB.1 raised the alarm over SARS-CoV-2 evolution with unprecedented antibody evasion from broad nAbs elicited by prototype vaccination.

Additional mutations based on Omicron BA.2.75 mediate its further evasion from broadly neutralizing antibodies.

SARS-CoV-2 Omicron BA.2.75 subvariant has evolved to a series of progeny variants carrying several additional mutations in the receptor-binding domain (RBD). Here, we investigated whether and how these single mutations based on BA.2.75 affect the neutralization of currently available anti-RBD monoclonal antibodies (mAbs) with well-defined structural information. Approximately 34% of mAbs maintained effective neutralizing activities against BA.2.75, consistent with those against BA.2, BA.4/5, and BA.2.12.1. Single additional R346T, K356T, L452R, or F486S mutations further facilitated BA.2.75-related progeny variants to escape from broadly neutralizing antibodies (bnAbs) at different degree. Only LY-CoV1404 (bebtelovimab) displayed a first-class neutralization potency and breadth against all tested Omicron subvariants. Overall, these data make a clear connection between virus escape and antibody recognizing antigenic epitopes, which facilitate to develop next-generation universal bnAbs against emerging SARS-CoV-2 variants.

Structures of a constitutively active mutant of human IDH3 reveal new insights into the mechanisms of allosteric activation and the catalytic reaction.

Human NAD-dependent isocitrate dehydrogenase or IDH3 (HsIDH3) catalyzes the decarboxylation of isocitrate into α-ketoglutarate in the tricarboxylic acid cycle. It consists of three types of subunits (α, ß, and γ) and exists and functions as the (αßαγ)2 heterooctamer. HsIDH3 is regulated allosterically and/or competitively by numerous metabolites including CIT, ADP, ATP, and NADH. Our previous studies have revealed the molecular basis for the activity and regulation of the αß and αγ heterodimers. However, the molecular mechanism for the allosteric activation of the HsIDH3 holoenzyme remains elusive. In this work, we report the crystal structures of the αß and αγ heterodimers and the (αßαγ)2 heterooctamer containing an α-Q139A mutation in the clasp domain, which renders all the heterodimers and the heterooctamer constitutively active in the absence of activators. Our structural analysis shows that the α-Q139A mutation alters the hydrogen-bonding network at the heterodimer-heterodimer interface in a manner similar to that in the activator-bound αγ heterodimer. This alteration not only stabilizes the active sites of both αQ139Aß and αQ139Aγ heterodimers in active conformations but also induces conformational changes of the pseudo-allosteric site of the αQ139Aß heterodimer enabling it to bind activators. In addition, the αQ139AICT+Ca+NADßNAD structure presents the first pseudo-Michaelis complex of HsIDH3, which allows us to identify the key residues involved in the binding of cofactor, substrate, and metal ion. Our structural and biochemical data together reveal new insights into the molecular mechanisms for allosteric regulation and the catalytic reaction of HsIDH3.

A key F27I substitution within HCDR1 facilitates the rapid maturation of P2C-1F11-like neutralizing antibodies in a SARS-CoV-2-infected donor.

Although thousands of anti-SARS-CoV-2 monoclonal neutralizing antibodies (nAbs) have been identified and well characterized, some crucial events in the development of these nAbs during viral infection remain unclear. Using deep sequencing, we explore the dynamics of antibody repertoire in a SARS-CoV-2-infected donor, from whom the potent and broad nAb P2C-1F11 (the parent version of Brii-196) was previously isolated. Further analysis shows a rapid clonal expansion of some SARS-CoV-2-specific antibodies in early infection. Longitudinal tracing of P2C-1F11 lineage antibodies reveals that these elite nAbs were rare. Using sequence alignment, structure modeling, and bioactivity analysis based on site-mutated assay, we demonstrate that a key substitution F27I in heavy chain contributes significantly to the maturation of P2C-1F11-like antibodies. Overall, our findings elucidate the developmental process and maturation pathway of P2C-1F11, providing some important information for the design of novel immunogens to elicit more potent nAbs against SARS-CoV-2 infection.

Identification and application of a pair of noncompeting monoclonal antibodies broadly binding to the nucleocapsid proteins of SARS-CoV-2 variants including Omicron.

The SARS-CoV-2 nucleocapsid protein (NP) is an important indicator for the virus infection, highlighting the crucial role of NP-specific monoclonal antibodies (mAbs) used in multiple biochemical assays and clinical diagnosis for detecting the NP antigen. Here, we reported a pair of noncompeting human NP-specific mAbs, named P301-F7 and P301-H5, targeting two distinct linear epitopes on SARS-CoV-2 or SARS-CoV. We evaluated the application of P301-F7 in the analysis of enzyme linked immunosorbent assay, western blot, flow cytometry, immunofluorescence, and focus reduction neutralization test. We for the first time report a broad mAb effectively recognizing various live viruses of SARS-CoV-2 variants including Alpha, Beta, Delta, and Omicron, indicating a wide range of application prospects.

Increased resistance of SARS-CoV-2 Lambda variant to antibody neutralization.

A recently identified SARS-CoV-2 variant, Lambda, has spread to many countries around the world. Here, we measured and evaluated the reduced sensitivity of Lambda variant to the neutralization by plasma polyclonal antibodies elicited by the natural SARS-CoV-2 infection and inactivated vaccine. The combination of two substitutions appearing in the RBD of spike protein (L452Q and F490S) resulted in noticeably reduced neutralization against Lambda variant. F490S contributed more than L452Q in affecting the neutralization. In addition, the neutralization test with 12 published nAbs binding to RBD of SARS-CoV-2 with defined structures suggested that Lambda variant resisted the neutralization by some antibodies from Class 2 and Class 3. Overall, these results suggest that pre-existing antibody neutralization established by natural infection from non-Lambda variants or immunization could be significantly decreased, re-emphasizing the importance of ongoing viral mutation monitoring.

Structure of Arabidopsis thaliana N6-methyl-AMP deaminase ADAL with bound GMP and IMP and implications for N6-methyl-AMP recognition and processing.

Arabidopsis thaliana aminohydrolase (AtADAL) has been shown to be involved in the metabolism of N6-methyl-AMP, a proposed intermediate during m6A-modified RNA metabolism, which can be subsequently incorporated into newly synthesized RNA by Pol II. It has been proposed that AtADAL will prevent N6-methyl-AMP reuse and catabolize it to inosine monophosphate (IMP). Here, we have solved the crystal structures of AtADAL in the apo form and in complex with GMP and IMP in the presence of Zn2+. We have identified the substrate-binding pocket of AtADAL and compared it with that for adenosine deaminase (ADA), adenine deaminase (ADE) and AMP deaminase (AMPD) from multiple species. The comparisons reveal that plant ADAL1 may have the potential ability to catalyze different alkyl-group substituted substrates.

Cbln1 and Cbln4 Are Structurally Similar but Differ in GluD2 Binding Interactions.

Unlike cerebellin 1 (Cbln1), which bridges neurexin (Nrxn) receptors and δ-type glutamate receptors in a trans-synaptic triad, Cbln4 was reported to have no or weak binding for the receptors despite sharing ∼70% sequence identity with Cbln1. Here, we report crystal structures of the homotrimers of the C1q domain of Cbln1 and Cbln4 at 2.2 and 2.3 Å resolution, respectively. Comparison of the structures suggests that the difference between Cbln1 and Cbln4 in GluD2 binding might be because of their sequence and structural divergence in loop CD. Surprisingly, we show that Cbln4 binds to Nrxn1ß and forms a stable complex with the laminin, nectin, sex-hormone binding globulin (LNS) domain of Nrxn1ß. Furthermore, the negative-stain electron microscopy reconstruction of hexameric full-length Cbln1 at 13 Å resolution and that of the Cbln4/Nrxn1ß complex at 19 Å resolution suggest that Nrxn1ß binds to the N-terminal region of Cbln4, probably through strand ß10 of the S4 insert.