There is nothing exempt from the peril of mutation - The Omicron spike.

The 2019 corona virus disease (COVID-19) has caused a global chaos, where a novel Omicron variant has challenged the healthcare system, followed by which it has been referred to as a variant of concern (VOC) by the World Health Organization (WHO), owing to its alarming transmission and infectivity rate. The large number of mutations in the receptor binding domain (RBD) of the spike protein is responsible for strengthening of the spike-angiotensin-converting enzyme 2 (ACE2) interaction, thereby explaining the elevated threat. This is supplemented by enhanced resistance of the variant towards pre-existing antibodies approved for the COVID-19 therapy. The manuscript brings into light failure of existing therapies to provide the desired effect, however simultaneously discussing the novel possibilities on the verge of establishing suitable treatment portfolio. The authors entail the risks associated with omicron resistance against antibodies and vaccine ineffectiveness on one side, and novel approaches and targets - kinase inhibitors, viral protease inhibitors, phytoconstituents, entry pathways - on the other. The manuscript aims to provide a holistic picture about the Omicron variant, by providing comprehensive discussions related to multiple aspects of the mutated spike variant, which might aid the global researchers and healthcare experts in finding an optimised solution to this pandemic.

Short linear motif candidates in the cell entry system used by SARS-CoV-2 and their potential therapeutic implications.

The first reported receptor for SARS-CoV-2 on host cells was the angiotensin-converting enzyme 2 (ACE2). However, the viral spike protein also has an RGD motif, suggesting that cell surface integrins may be co-receptors. We examined the sequences of ACE2 and integrins with the Eukaryotic Linear Motif (ELM) resource and identified candidate short linear motifs (SLiMs) in their short, unstructured, cytosolic tails with potential roles in endocytosis, membrane dynamics, autophagy, cytoskeleton, and cell signaling. These SLiM candidates are highly conserved in vertebrates and may interact with the µ2 subunit of the endocytosis-associated AP2 adaptor complex, as well as with various protein domains (namely, I-BAR, LC3, PDZ, PTB, and SH2) found in human signaling and regulatory proteins. Several motifs overlap in the tail sequences, suggesting that they may act as molecular switches, such as in response to tyrosine phosphorylation status. Candidate LC3-interacting region (LIR) motifs are present in the tails of integrin ß3 and ACE2, suggesting that these proteins could directly recruit autophagy components. Our findings identify several molecular links and testable hypotheses that could uncover mechanisms of SARS-CoV-2 attachment, entry, and replication against which it may be possible to develop host-directed therapies that dampen viral infection and disease progression. Several of these SLiMs have now been validated to mediate the predicted peptide interactions.

Design, Synthesis, Evaluation, and Structural Studies of C2-Symmetric Small Molecule Inhibitors of Programmed Cell Death-1/Programmed Death-Ligand 1 Protein-Protein Interaction.

A series of C2-symmetric inhibitors was designed and evaluated for inhibitory activity against the programmed cell death-1/programmed death-ligand 1(PD-1/PD-L1) protein-protein interaction (PPI) in a homogenous time-resolved fluorescence (HTRF) assay and PD-1 signaling in cell-based coculture assays. C2-symmetric inhibitors 2a (LH1306) and 2b (LH1307) exhibited IC50 values of 25 and 3.0 nM, respectively, in the HTRF assay. While 2a was ∼3.8-fold more potent than previously reported inhibitor 1a, 2b could not be differentiated from 1b due to their high potency and the limit of our HTRF assay conditions. In one cell-based coculture PD-1 signaling assay, 2a and 2b were 8.2- and 2.8-fold more potent in inhibiting PD-1 signaling than 1a and 1b, respectively. NMR and X-ray cocrystal structural studies provided more structural insights into the interaction between 2b and PD-L1; 2b binds to PD-L1 at the PD-1 binding site and induces the formation of a more symmetrically arranged PD-L1 homodimer than that previously reported for other inhibitors.

Deconstruction of an African folk medicine uncovers a novel molecular strategy for therapeutic potassium channel activation.

A third of the global population relies heavily upon traditional or folk medicines, such as the African shrub Mallotus oppositifolius. Here, we used pharmacological screening and electrophysiological analysis in combination with in silico docking and site-directed mutagenesis to elucidate the effects of M. oppositifolius constituents on KCNQ1, a ubiquitous and influential cardiac and epithelial voltage-gated potassium (Kv) channel. Two components of the M. oppositifolius leaf extract, mallotoxin (MTX) and 3-ethyl-2-hydroxy-2-cyclopenten-1-one (CPT1), augmented KCNQ1 current by negative shifting its voltage dependence of activation. MTX was also highly effective at augmenting currents generated by KCNQ1 in complexes with native partners KCNE1 or SMIT1; conversely, MTX inhibited KCNQ1-KCNE3 channels. MTX and CPT1 activated KCNQ1 by hydrogen bonding to the foot of the voltage sensor, a previously unidentified drug site which we also find to be essential for MTX activation of the related KCNQ2/3 channel. The findings elucidate the molecular mechanistic basis for modulation by a widely used folk medicine of an important human Kv channel and uncover novel molecular approaches for therapeutic modulation of potassium channel activity.

Human and mouse albumin bind their respective neonatal Fc receptors differently.

Albumin has a serum half-life of three weeks in humans and is utilized to extend the serum persistence of drugs that are genetically fused or conjugated directly to albumin or albumin-binding molecules. Responsible for the long half-life is FcRn that protects albumin from intracellular degradation. An in-depth understanding of how FcRn binds albumin across species is of importance for design and evaluation of albumin-based therapeutics. Albumin consists of three homologous domains where domain I and domain III of human albumin are crucial for binding to human FcRn. Here, we show that swapping of two loops in domain I or the whole domain with the corresponding sequence in mouse albumin results in reduced binding to human FcRn. In contrast, humanizing domain I of mouse albumin improves binding. We reveal that domain I of mouse albumin plays a minor role in the interaction with the mouse and human receptors, as domain III on its own binds with similar affinity as full-length mouse albumin. Further, we show that P573 in domain III of mouse albumin is required for strong receptor binding. Our study highlights distinct differences in structural requirements for the interactions between mouse and human albumin with their respective receptor, which should be taken into consideration in design of albumin-based drugs and evaluation in mouse models.

A critical evaluation of self-interaction chromatography as a predictive tool for the assessment of protein-protein interactions in protein formulation development: a case study of a therapeutic monoclonal antibody.

The aim of this study was to establish and evaluate a screening method for the physical characterization of protein-protein interactions of therapeutic proteins based on the determination of the osmotic second virial coefficient (B(22)). B(22) of an IgG1 was measured by self-interaction chromatography (SIC) and was compared to data obtained from static light scattering (SLS). As assessed by Fourier transform infrared spectroscopy (FTIR), the protein coupling to chromatography particles had no relevant influence on the three-dimensional native structure of the IgG1. B(22) variations could be measured for physiological relevant excipient concentrations. Significant positive B(22) values were observed for the following solution conditions of the investigated antibody: (i) acidic pH conditions, (ii) low buffer concentrations, (iii) low salt concentrations and (iv) high amino acid concentrations. B(22) was compared to IgG1 stability data derived from a study conducted for 12weeks at 40 degrees C. A concentration of 5mM histidine, which was the most promising buffer candidate according to B(22), showed a slightly better physical stability (as assessed by turbidity and size exclusion chromatography) compared to the other tested formulations. This is confirmed in a stress study investigating the colloidal stability. Thus, measuring protein-protein interactions with SIC appeared as a promising screening tool for physical characterization of protein formulations for cases in which the protein stability is governed by interparticle interactions.