scholarly journals Mitoxantrone dihydrochloride, an FDA approved drug, binds with SARS-CoV-2 NSP1 C-terminal

2021 ◽  
Author(s):  
Prateek Kumar ◽  
Taniya Bhardwaj ◽  
Rajanish Giri
Keyword(s):  
Structural Dynamics ◽  
Immune Evasion ◽  
Drug Target ◽  
Mrna Translation ◽  
Small Subunit ◽  
Therapeutic Strategies ◽  
Spectroscopic Technique ◽  
Terminal Region ◽  
Binding Assays ◽  
Computational Simulations

One of the major virulence factors of SARS-CoV-2, NSP1, is a vital drug target due to its role in host immune evasion through multiple pathways. NSP1 protein is associated with inhibiting host mRNA translation by binding to the small subunit of ribosome through its C-terminal region. Previously, we have shown the structural dynamics of NSP1 C-terminal region (NSP1-CTR) in different physiological environments. So, it would be very interesting to investigate the druggable compounds that could bind with NSP1-CTR. Here, in this article, we have performed the different spectroscopic technique-based binding assays of an anticancer drug Mitoxantrone dihydrochloride (MTX) against the NSP1-CTR. We have also performed molecular docking followed by computational simulations with two different forcefields up to one microsecond. Overall, our results have suggested good binding between NSP1-CTR and MTX and may have implications in developing therapeutic strategies targeting NSP1 protein of SARS-CoV-2.

scholarly journals Structural dynamics of the SARS-CoV-2 frameshift-stimulatory pseudoknot reveal topologically distinct conformers

2020 ◽  
Author(s):  
Krishna Neupane ◽  
Meng Zhao ◽  
Aaron Lyons ◽  
Sneha Munshi ◽  
Sandaru M Ileperuma ◽  
...  
Keyword(s):  
Drug Discovery ◽  
Optical Tweezers ◽  
Structural Dynamics ◽  
Drug Target ◽  
Conformational Heterogeneity ◽  
Computational Simulations ◽  
Mechanical Tension ◽  
Stem Loop ◽  
Ribosomal Frameshifting ◽  
Rna Pseudoknot

The RNA pseudoknot that stimulates −1 programmed ribosomal frameshifting in SARS coronavirus-2 (SARS-CoV-2) is a possible drug target. To understand how this 3-stemmed pseudoknot responds to the mechanical tension applied by ribosomes during translation, which is thought to play a key role during frameshifting, we probed its structural dynamics under tension using optical tweezers. Unfolding curves revealed that the frameshift signal formed multiple different structures: at least two distinct pseudoknotted conformers with different unfolding forces and energy barriers, as well as alternative stem-loop structures. Refolding curves showed that stem 1 formed first in the pseudoknotted conformers, followed by stem 3 and then stem 2. By extending the handle holding the RNA to occlude the 5′ end of stem 1, the proportion of the different pseudoknot conformers could be altered systematically, consistent with structures observed in cryo-EM images and computational simulations that had distinct topologies: the 5′ end of the RNA threaded through the 3-helix junction to form a ring-knot, or unthreaded as in more standard H-type pseudoknots. These results resolve the folding mechanism of the frameshift signal in SARS-CoV-2 and highlight the dynamic conformational heterogeneity of this RNA, with important implications for structure-based drug-discovery efforts.

scholarly journals Recent Advancements in the Mechanisms Underlying Resistance to PD-1/PD-L1 Blockade Immunotherapy

Cancers ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 663
Author(s):  
Yu Yuan ◽  
Abdalla Adam ◽  
Chen Zhao ◽  
Honglei Chen
Keyword(s):  
Immune Evasion ◽  
Immune Checkpoint ◽  
Poor Prognosis ◽  
Target Therapy ◽  
Acquired Resistance ◽  
Immune Checkpoint Blockade ◽  
Therapeutic Strategies ◽  
Present Evidence ◽  
Immunosuppressive Factor

Release of immunoreactive negative regulatory factors such as immune checkpoint limits antitumor responses. PD-L1 as a significant immunosuppressive factor has been involved in resistance to therapies such as chemotherapy and target therapy in various cancers. Via interacting with PD-1, PD-L1 can regulate other factors or lead to immune evasion of cancer cells. Besides, immune checkpoint blockade targeting PD-1/PD-L1 has promising therapeutic efficacy in the different tumors, but a significant percentage of patients cannot benefit from this therapy due to primary and acquired resistance during treatment. In this review, we described the utility of PD-L1 expression levels for predicting poor prognosis in some tumors and present evidence for a role of PD-L1 in resistance to therapies through PD-1/PD-L1 pathway and other correlating signaling pathways. Afterwards, we elaborate the key mechanisms underlying resistance to PD-1/PD-L1 blockade in cancer immunotherapy. Furthermore, promising combination of therapeutic strategies for patients resistant to PD-1/PD-L1 blockade therapy or other therapies associated with PD-L1 expression was also summarized.

scholarly journals A UV-Induced Mutation in Neurospora That Affects Translational Regulation in Response to Arginine

Genetics ◽  
1996 ◽  
Vol 142 (1) ◽  
pp. 117-127 ◽  
Author(s):  
Michael Freitag ◽  
Nelima Dighde ◽  
Matthew S Sachs
Keyword(s):  
Translational Control ◽  
Translational Regulation ◽  
Fusion Gene ◽  
Mrna Translation ◽  
Small Subunit ◽  
Upstream Open Reading Frame ◽  
Control Mechanisms ◽  
Carbamoyl Phosphate ◽  
Altered Expression ◽  
Reading Frame

The Neurospora crmsu arg-2 gene encodes the small subunit of arginine-specific carbamoyl phosphate synthetase. The levels of arg-2 mRNA and mRNA translation are negatively regulated by arginine. An upstream open reading frame (uORF) in the transcript’s 5′ region has been implicated in arginine-specific control. An arg-2-hph fusion gene encoding hygromycin phosphotransferase conferred arginine-regulated resistance to hygromycin when introduced into N. crassa. We used an arg-2-hph strain to select for UV-induced mutants that grew in the presence of hygromycin and arginine, and we isolated 46 mutants that had either of two phenotypes. One phenotype indicated altered expression of both arg-2-hph and urg-2 genes; the other, altered expression of urg-2-hph but not arg-2. One of the latter mutations, which was genetically closely linked to arg-2-hph, was recovered from the 5′ region of the arg-2-hph gene using PCR. Sequence analyses and transformation experiments revealed a mutation at uORF codon 12 (Asp to Asn) that abrogated negative regulation. Examination of the distribution of ribosomes on arg-2-hph transcripts showed that loss of regulation had a translational component, indicating the uORF sequence was important for Arg-specific translational control. Comparisons with other uORFS suggest common elements in translational control mechanisms.

scholarly journals Enzyme Specificity of 2-Nitrotoluene 2,3-Dioxygenase from Pseudomonas sp. Strain JS42 Is Determined by the C-Terminal Region of the α Subunit of the Oxygenase Component

1998 ◽  
Vol 180 (5) ◽  
pp. 1194-1199 ◽  
Author(s):  
Juanito V. Parales ◽  
Rebecca E. Parales ◽  
Sol M. Resnick ◽  
David T. Gibson
Keyword(s):  
Amino Acid Sequence Identity ◽  
Large Subunit ◽  
Small Subunit ◽  
Terminal Region ◽  
Oxidation Products ◽  
Enzyme Specificity ◽  
Α Subunit ◽  
Sequence Identity ◽  
Wide Range ◽  
Genes Encoding

ABSTRACT Biotransformations with recombinant Escherichia coliexpressing the genes encoding 2-nitrotoluene 2,3-dioxygenase (2NTDO) from Pseudomonas sp. strain JS42 demonstrated that 2NTDO catalyzes the dihydroxylation and/or monohydroxylation of a wide range of aromatic compounds. Extremely high nucleotide and deduced amino acid sequence identity exists between the components from 2NTDO and the corresponding components from 2,4-dinitrotoluene dioxygenase (2,4-DNTDO) from Burkholderia sp. strain DNT (formerlyPseudomonas sp. strain DNT). However, comparisons of the substrates oxidized by these dioxygenases show that they differ in substrate specificity, regiospecificity, and the enantiomeric composition of their oxidation products. Hybrid dioxygenases were constructed with the genes encoding 2NTDO and 2,4-DNTDO. Biotransformation experiments with these hybrid dioxygenases showed that the C-terminal region of the large subunit of the oxygenase component (ISPα) was responsible for the enzyme specificity differences observed between 2NTDO and 2,4-DNTDO. The small subunit of the terminal oxygenase component (ISPβ) was shown to play no role in determining the specificities of these dioxygenases.

scholarly journals Fibrillin Assembly Requires Fibronectin

2009 ◽  
Vol 20 (3) ◽  
pp. 846-858 ◽  
Author(s):  
Laetitia Sabatier ◽  
Daliang Chen ◽  
Christine Fagotto-Kaufmann ◽  
Dirk Hubmacher ◽  
Marc D. McKee ◽  
...  
Keyword(s):  
Dermal Fibroblasts ◽  
Terminal Region ◽  
Electron Microscopic Level ◽  
Binding Assays ◽  
Electron Microscopic ◽  
Binding Interaction ◽  
Fibrillin 1 ◽  
And Function ◽  
Sirna Knockdown ◽  
Fibronectin Fibrils

Fibrillins constitute the major backbone of multifunctional microfibrils in elastic and nonelastic extracellular matrices. Proper assembly mechanisms are central to the formation and function of these microfibrils, and their properties are often compromised in pathological circumstances such as in Marfan syndrome and in other fibrillinopathies. Here, we have used human dermal fibroblasts to analyze the assembly of fibrillin-1 in dependence of other matrix-forming proteins. siRNA knockdown experiments demonstrated that the assembly of fibrillin-1 is strictly dependent on the presence of extracellular fibronectin fibrils. Immunolabeling performed at the light and electron microscopic level showed colocalization of fibrillin-1 with fibronectin fibrils at the early stages of the assembly process. Protein-binding assays demonstrated interactions of fibronectin with a C-terminal region of fibrillin-1, -2, and -3 and with an N-terminal region of fibrillin-1. The C-terminal half of fibrillin-2 and -3 had propensities to multimerize, as has been previously shown for fibrillin-1. The C-terminal of all three fibrillins interacted strongly with fibronectin as multimers, but not as monomers. Mapping studies revealed that the major binding interaction between fibrillins and fibronectin involves the collagen/gelatin-binding region between domains FNI6 and FNI9.

Biosensor Analysis of Drug–Target Interactions: Direct and Competitive Binding Assays for Investigation of Interactions between Thrombin and Thrombin Inhibitors

2001 ◽  
Vol 291 (2) ◽  
pp. 306 ◽  
Author(s):  
Robert Karlsson ◽  
Mari Kullman-Magnusson ◽  
Markku D. Hämäläinen ◽  
Annika Remaeus ◽  
Karl Andersson ◽  
...  
Keyword(s):  
Drug Target ◽  
Competitive Binding ◽  
Thrombin Inhibitors ◽  
Binding Assays ◽  
Biosensor Analysis ◽  
Competitive Binding Assays

scholarly journals Identification of β-Glucosidase Aggregating Factor (BGAF) and Mapping of BGAF Binding Regions on Maize β-Glucosidase

2000 ◽  
Vol 276 (15) ◽  
pp. 11895-11901 ◽  
Author(s):  
David J. Blanchard ◽  
Muzaffer Cicek ◽  
Jialun Chen ◽  
Asim Esen
Keyword(s):  
Null Allele ◽  
Terminal Region ◽  
Binding Assays ◽  
Terminal Sequence ◽  
Chain Reaction ◽  
Chimeric Enzymes ◽  
Repeat Units ◽  
Maize Genotypes ◽  
Polymerase Chain ◽  
A Site

In certain maize genotypes (nulls), β-glucosidase does not enter the gel and therefore cannot be detected on zymograms. Such genotypes were initially thought to be homozygous for a null allele at theglu1gene. We have shown that a β-glucosidase aggregating factor (BGAF) is responsible for the null phenotype, and it specifically interacts with maize β-glucosidases and forms large insoluble aggregates. To understand the mechanism of the β-glucosidase-BGAF interaction, we constructed chimeric enzymes by domain swapping between the maize β-glucosidase isozymes Glu1 and Gu2, to which BGAF binds, and the sorghum β-glucosidase (dhurrinase) isozyme Dhr1, to which BGAF does not bind. The results of binding assays with 12 different chimeric enzymes showed that an N-terminal region (Glu50-Val145) and an extreme C-terminal region (Phe466-Ala512) together form the BGAF binding site on the enzyme surface. In addition, we purified BGAF, determined its N-terminal sequence, amplified the BGAF cDNA by reverse transcriptase-polymerase chain reaction, expressed it inEscherichia coli, and showed that it encodes a protein whose binding and immunological properties are identical to the native BGAF isolated from maize tissues. A data base search revealed that BGAF is a member of the jasmonite-induced protein family. Interestingly, the deduced BGAF sequence contained an octapeptide sequence (G(P/R)WGGSGG) repeated twice. Each of these repeat units is postulated to be involved in forming a site for binding to maize β-glucosidases and thus provides a plausible explanation for the divalent function of BGAF predicted from binding assays.

scholarly journals Translational regulation in response to changes in amino acid availability in Neurospora crassa.

1995 ◽  
Vol 15 (10) ◽  
pp. 5235-5245 ◽  
Author(s):  
Z Luo ◽  
M Freitag ◽  
M S Sachs
Keyword(s):  
Amino Acid ◽  
Neurospora Crassa ◽  
Translational Regulation ◽  
Mrna Translation ◽  
Small Subunit ◽  
Negative Regulation ◽  
Start Codon ◽  
Mrna Levels ◽  
Carbamoyl Phosphate ◽  
Amino Acid Availability

We examined the regulation of Neurospora crassa arg-2 and cpc-1 in response to amino acid availability.arg-2 encodes the small subunit of arginine-specific carbamoyl phosphate synthetase; it is subject to unique negative regulation by Arg and is positively regulated in response to limitation for many different amino acids through a mechanism known as cross-pathway control. cpc-1 specifies a transcriptional activator important for crosspathway control. Expression of these genes was compared with that of the cytochrome oxidase subunit V gene, cox-5. Analyses of mRNA levels, polypeptide pulse-labeling results, and the distribution of mRNA in polysomes indicated that Arg-specific negative regulation of arg-2 affected the levels of both arg-2 mRNA and arg-2 mRNA translation. Negative translational effects on arg-2 and positive translational effects on cpc-1 were apparent soon after cells were provided with exogenous Arg. In cells limited for His, increased expression of arg-2 and cpc-1, and decreased expression of cox-5, also had translational and transcriptional components. The arg-2 and cpc-1 transcripts contain upstream open reading frames (uORFs), as do their Saccharomyces cerevisiae homologs CPA1 and GCN4. We examined the regulation of arg-2-lacZ reporter genes containing or lacking the uORF start codon; the capacity for arg-2 uORF translation appeared critical for controlling gene expression.

scholarly journals Immune evasion mechanisms and therapeutic strategies in gastric cancer

2022 ◽  
Vol 14 (1) ◽  
pp. 216-229
Author(s):  
En-Si Ma ◽  
Zheng-Xin Wang ◽  
Meng-Qi Zhu ◽  
Jing Zhao
Keyword(s):  
Gastric Cancer ◽  
Immune Evasion ◽  
Therapeutic Strategies
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