Biology subjects

Verma, D. K.

Publications and source records attributed to Verma, D. K..

3 recordsLinked to original sources

bioRxiv2023

Assessment of structural behaviour of new L-asparaginase and SAXS data-based evidence for catalytic activity in its monomeric form

The present study reports the structural and functional characterization of a new glutaminase-free recombinant L-asparaginase (PrASNase) from Pseudomonas resinovorans IGS-131. PrASNase showed substrate specificity to L-asparagine, and its kinetic parameters, Km, Vmax, and kcat were 9.49x10-3 M, 25.13 IUmL-1min-1, and 3.01x103 s-1, respectively. The CD spectra showed that PrASNase consists of 30.9% -helix and 69.1% other structures in its native form. FTIR was used for the functional characterization, and molecular docking predicted that the substrate interacts with serine, alanine, and glutamine in the binding pocket of PrASNase. Different from known asparaginases, structural characterization by small-angle X-ray scattering (SAXS) and analytical ultracentrifugation (AUC) unambiguously revealed PrASNase to exist as a monomer in solution at low temperatures and oligomerized to a higher state with temperature rise. Through SAXS studies and enzyme assay, PrASNase was found to be mostly monomer and catalytically active at 37{degrees}C. Furthermore, this glutaminase-free PrASNase showed killing effects against WIL2-S and TF-1.28 cells with IC50 of 7.4 {micro}g.mL-1 and 5.6 {micro}g.mL-1, respectively. This is probably the first report with significant findings of fully active L-asparaginase in monomeric form using SAXS and AUC and demonstrates the potential of PrASNase in inhibiting cancerous cells, making it a potential therapeutic candidate. HIGHLIGHTSO_LIA new L-asparaginase (PrASNase) was structurally and functionally characterized. C_LIO_LISAXS revealed PrASNase is functionally active in monomeric form and oligomerizes with temperature rise. C_LIO_LIMonomeric PrASNase showed an IC50 value of 7.4 and 5.6 {micro}g mL-1 against WIL2-S and TF-1.28 cells. C_LIO_LICytotoxicity of PrASNase against leukemic cell lines showed its potential as a biotherapeutic. C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=133 SRC="FIGDIR/small/522448v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@ece494org.highwire.dtl.DTLVardef@92cdbeorg.highwire.dtl.DTLVardef@19a2borg.highwire.dtl.DTLVardef@1308c70_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2021

Zinc2+ ion inhibits SARS-CoV-2 main protease and viral replication in vitro.

Zinc deficiency is linked to poor prognosis in COVID-19 patients while clinical trials with Zinc demonstrate better clinical outcome. The molecular target and mechanistic details of anti-coronaviral activity of Zinc remain obscure. We show that ionic Zinc not only inhibits SARS-CoV-2 main protease (Mpro) with nanomolar affinity, but also viral replication. We present the first crystal structure of Mpro-Zn2+ complex at 1.9 [A] and provide the structural basis of viral replication inhibition. We show that Zn2+ coordinates with the catalytic dyad at the enzyme active site along with two previously unknown water molecules in a tetrahedral geometry to form a stable inhibited Mpro-Zn2+ complex. Further, natural ionophore quercetin increases the anti-viral potency of Zn2+. As the catalytic dyad is highly conserved across SARS-CoV, MERS-CoV and all variants of SARS-CoV-2, Zn2+ mediated inhibition of Mpro may have wider implications.

biochemistry↗

bioRxiv2017

The immunomodulatory CEA cell adhesion molecule 6 (CEACAM6/CD66c) is a candidate receptor for the influenza A virus

To establish a productive infection in host cells, viruses often use one or multiple host membrane glycoprotein as their receptors. For Influenza A virus (IAV) such a glycoprotein receptor has not been described, to date. Here we show that IAV is using the host membrane glycoprotein CD66c as a receptor for entry into human epithelial lung cells. Neuraminidase (NA), a viral spike protein binds to CD66c on the cell surface during IAV entry into the host cells. Lung cells overexpressing CD66c showed an increase in virus binding and subsequent entry into the cell. Upon comparison, CD66c demonstrated higher binding capacity than other membrane glycoproteins (EGFR and DC-SIGN) reported earlier to facilitate IAV entry into host cells. siRNA mediated knockdown of CD66c from lung cells inhibited virus binding on cell surface and entry into cells. Blocking CD66c by antibody on the cell surface resulted in decreased virus entry. We found CD66c is a specific glycoprotein receptor for influenza A virus that did not affect entry of non-IAV RNA virus (Hepatitis C virus). Finally, IAV pre-incubated with recombinant CD66c protein when administered intranasally in mice showed decreased cytopathic effects in mice lungs. This publication is the first to report CD66c (CEACAM6) as a glycoprotein receptor for Influenza A virus.\n\nSignificance StatementCells are enclosed by a semipermeable membrane that allows selective exchange of biomolecules between cells and their surroundings. A set of specialized proteins in this semipermeable membrane, work like gatekeepers to the cell and regulate entry of these biomolecules. One class of such surface proteins is termed as receptors. Viruses bind to one or more of these receptors and manipulate gatekeepers for their own successful entry into host-cells. A membrane protein that influenza A virus (Flu virus) uses for entry into the cells was not discovered till date. This study reports for the first time, a receptor for influenza A virus, that was sought after by researchers for decades. The viral receptor is a promising target that can be used to inhibit virus entry into host cells.

microbiology↗