Targeting spike glycans to inhibit SARS-CoV2 viral entry.

SARS-CoV-2 spike harbors glycans which function as ligands for lectins. Therefore, it should be possible to exploit lectins to target SARS-CoV-2 and inhibit cellular entry by binding glycans on the spike protein. Burkholderia oklahomensis agglutinin (BOA) is an antiviral lectin that interacts with viral glycoproteins via N-linked high mannose glycans. Here, we show that BOA binds to the spike protein and is a potent inhibitor of SARS-CoV-2 viral entry at nanomolar concentrations. Using a variety of biophysical approaches, we demonstrate that the interaction is avidity driven and that BOA cross-links the spike protein into soluble aggregates. Furthermore, using virus neutralization assays, we demonstrate that BOA effectively inhibits all tested variants of concern as well as SARS-CoV 2003, establishing that multivalent glycan-targeting molecules have the potential to act as pan-coronavirus inhibitors.

Ligand-Capped Cobalt(II) Multiplies the Value of the Double-Histidine Motif for PCS NMR Studies.

In structural studies by NMR, pseudocontact shifts (PCSs) provide both angular and distance information. For proteins, incorporation of a di-histidine (diHis) motif, coordinated to Co2+, has emerged as an important tool to measure PCS. Here, we show that using different Co(II)-chelating ligands, such as nitrilotriacetic acid (NTA) and iminodiacetic acid (IDA), resolves the isosurface ambiguity of Co2+-diHis and yields orthogonal PCS data sets with different Δχ-tensors for the same diHis-bearing protein. Importantly, such capping ligands effectively eliminate undesired intermolecular interactions, which can be detrimental to PCS studies. Devising and employing ligand-capping strategies afford versatile and powerful means to obtain multiple orthogonal PCS data sets, significantly extending the use of the diHis motif for structural studies by NMR.

Development and Validation of Fluorinated, Aromatic Amino Acid Parameters for Use with the AMBER ff15ipq Protein Force Field.

We developed force field parameters for fluorinated, aromatic amino acids enabling molecular dynamics (MD) simulations of fluorinated proteins. These parameters are tailored to the AMBER ff15ipq protein force field and enable the modeling of 4, 5, 6, and 7F-tryptophan, 3F- and 3,5F-tyrosine, and 4F- or 4-CF3-phenylalanine. The parameters include 181 unique atomic charges derived using the implicitly polarized charge (IPolQ) scheme in the presence of SPC/Eb explicit water molecules and 9 unique bond, angle, or torsion terms. Our simulations of benchmark peptides and proteins maintain expected conformational propensities on the µs time scale. In addition, we have developed an open-source Python program to calculate fluorine relaxation rates from MD simulations. The extracted relaxation rates from protein simulations are in good agreement with experimental values determined by 19F NMR. Collectively, our results illustrate the power and robustness of the IPolQ lineage of force fields for modeling the structure and dynamics of fluorine-containing proteins at the atomic level.

Targeting Spike Glycans to Inhibit SARS-CoV2 Viral Entry.

SARS-CoV-2 Spike harbors glycans which function as ligands for lectins. Therefore, it should be possible to exploit lectins to target SARS-CoV-2 and inhibit cellular entry by binding glycans on the Spike protein. Burkholderia oklahomensis agglutinin (BOA) is an antiviral lectin that interacts with viral glycoproteins via N-linked high mannose glycans. Here, we show that BOA binds to the Spike protein and is a potent inhibitor of SARS-CoV-2 viral entry at nanomolar concentrations. Using a variety of biophysical tools, we demonstrate that the interaction is avidity driven and that BOA crosslinks the Spike protein into soluble aggregates. Furthermore, using virus neutralization assays, we demonstrate that BOA effectively inhibits all tested variants of concern as well as SARS-CoV 2003, establishing that glycan-targeting molecules have the potential to be pan-coronavirus inhibitors.

A twist in the road less traveled: The AMBER ff15ipq-m force field for protein mimetics.

We present a new force field, AMBER ff15ipq-m, for simulations of protein mimetics in applications from therapeutics to biomaterials. This force field is an expansion of the AMBER ff15ipq force field that was developed for canonical proteins and enables the modeling of four classes of artificial backbone units that are commonly used alongside natural α residues in blended or "heterogeneous" backbones: chirality-reversed D-α-residues, the Cα-methylated α-residue Aib, homologated ß-residues (ß3) bearing proteinogenic side chains, and two cyclic ß residues (ßcyc; APC and ACPC). The ff15ipq-m force field includes 472 unique atomic charges and 148 unique torsion terms. Consistent with the AMBER IPolQ lineage of force fields, the charges were derived using the Implicitly Polarized Charge (IPolQ) scheme in the presence of explicit solvent. To our knowledge, no general force field reported to date models the combination of artificial building blocks examined here. In addition, we have derived Karplus coefficients for the calculation of backbone amide J-coupling constants for ß3Ala and ACPC ß residues. The AMBER ff15ipq-m force field reproduces experimentally observed J-coupling constants in simple tetrapeptides and maintains the expected conformational propensities in reported structures of proteins/peptides containing the artificial building blocks of interest-all on the µs timescale. These encouraging results demonstrate the power and robustness of the IPolQ lineage of force fields in modeling the structure and dynamics of natural proteins as well as mimetics with protein-inspired artificial backbones in atomic detail.