SARS-CoV-2 variants infectivity prediction and therapeutic peptide design using computational approaches.

The outbreak of severe acute respiratory coronavirus 2 (SARS-CoV-2) has created a public health emergency globally. SARS-CoV-2 enters the human cell through the binding of the spike protein to human angiotensin converting enzyme 2 (ACE2) receptor. Significant changes have been reported in the mutational landscape of SARS-CoV-2 in the receptor binding domain (RBD) of S protein, subsequent to evolution of the pandemic. The present study examines the correlation between the binding affinity of mutated S-proteins and the rate of viral infectivity. For this, the binding affinity of SARS-CoV and variants of SARS-CoV-2 towards ACE2 was computationally determined. Subsequently, the RBD mutations were classified on the basis of the number of strains identified with respect to each mutation and the resulting variation in the binding affinity was computationally examined. The molecular docking studies indicated a significant correlation between the Z-Rank score of mutated S proteins and the rate of infectivity, suitable for predicting SARS-CoV-2 infectivity. Accordingly, a 30-mer peptide was designed and the inhibitory properties were computationally analyzed. Single amino acid-wise mutation was performed subsequently to identify the peptide with the highest binding affinity. Molecular dynamics and free energy calculations were then performed to examine the stability of the peptide-protein complexes. Additionally, selected peptides were synthesized and screened using a colorimetric assay. Together, this study developed a model to predict the rate of infectivity of SARS-CoV-2 variants and propose a potential peptide that can be used as an inhibitor for the viral entry to human.Communicated by Ramaswamy H. Sarma.

Angiogenic Potential, Circulating Angiogenic Factors and Insulin Resistance in Subjects with Obesity.

Altered vascular function and pathological angiogenesis are important factors common to the development of obesity and obesity-associated diseases. Most human studies relating obesity and angiogenesis have compared levels of angiogenic factors in obesity without looking at the serum angiogenic capacity which reflects the balance between the effects of angiogenic and angiostatic factors. Therefore, in this cross-sectional study, the serum angiogenic potential and levels of angiogenic factors in serum of obese (BMI > 25 kg/m2) and lean subjects (BMI < 23 kg/m2), with no history of obesity associated co-morbidities, were assessed. Serum angiogenic potential was significantly higher (p < 0.0001) in both male (n = 67) and female (n = 35) obese subjects and showed a positive correlation (r = 0.4, p < 0.0001) with BMI. Serum levels of the angiogenic factors, vascular endothelial growth factor (VEGF) and angiopoietin were significantly higher in obese subjects. Levels of angiostatic factors such as angiostatin, endostatin were not altered in obese male subjects but were elevated in female obese subjects. Angiogenic potential and levels of VEGF did not vary in obese subjects with high HOMA-IR compared to obese subjects with low HOMA-IR. These results suggest that the angiogenic potential of serum was elevated in obesity and that insulin resistance may not contribute to the increased angiogenic potential in obesity.

C-Reactive Protein (CRP) and Leptin Receptor in Obesity: Binding of Monomeric CRP to Leptin Receptor.

While leptin deficiency or dysfunction leads to morbid obesity, obese subjects are characterized paradoxically by hyperleptinemia indicating lack of response to leptin. C-reactive protein (CRP) has been suggested to be a key plasma protein that could bind to leptin. To examine whether CRP interferes with leptin action, mediated through its cell surface receptor, docking studies of CRP with the extracellular domain of the leptin receptor were done employing bioinformatics tools. Monomeric CRP docked with better Z-rank score and more non-bond interactions than pentameric CRP at the CRH2-FNIII domain proximal to the cell membrane, distinct from the leptin-docking site. Interaction of CRP with leptin receptor was validated by solid phase binding assay and co-immunoprecipitation of CRP and soluble leptin receptor (sOb R) from human plasma. Analysis of the serum levels of leptin, CRP, and sOb R by ELISA showed that CRP levels were significantly elevated (p < 0.0001) in non-morbid obese subjects (n = 42) compared to lean subjects (n = 32) and correlated positively with body mass index (BMI) (r = 0.74, p < 0.0001) and leptin (r = 0.8, p < 0.0001); levels of sOb R were significantly low in obese subjects (p < 0.001) and showed a negative correlation with BMI (r = -0.26, p < 0.05) and leptin (r = -0.23, p < 0.05) indicating a minimal role for sOb R in sequestering leptin.

Changes in expression of VE-cadherin and MMPs in endothelial cells: Implications for angiogenesis.

The mechanism of cell-cell contact dependent regulation of pericellular proteolysis in angiogenesis was examined by studying the expression of MMPs using isolated HUVECs in culture. Zymography, Immunoblot and RT-PCR analysis showed that the production and secretion of matrixmetalloproteinase-2 and matrixmetalloproteinase-9 by HUVECs in culture were high when they remain as individual cells and significantly decreased during later stages of culture when cells developed cell-cell contact and tubular network-like structure. As MMPs decreased there was significant upregulation of VE-cadherin in cells undergoing angiogenic transition. Investigations to understand the signaling pathways downstream of VE-cadherin showed a relatively high level of ß-catenin in the nucleus of endothelial cells in culture during initial stages and decrease in its levels in the nucleus, associated with an increase in the cytosol during later stages of culture. The distribution of ß-catenin was found to be regulated by Tyr/Ser phosphorylation status of this protein. Cell-cell contact dependent downregulation of MMPs during angiogenesis was also observed in experiments using proangiogenic substances which caused a rapid rate of downregulation of MMP-2 and MMP-9 and absence of downregulation of MMPs when treated with anti-angiogenic agents.