bluuebunny/crossref_metadata_2025_split huggingface dataset:
Synthesis and Characterization of Esterified Fmoc‐Amino Esters as Potential Butyrylcholinesterase Inhibitors
Authors: Pingul Bianca Ysabel , Ramirez Jennifer , Schwans Jason 
Published: June 2021

Neurodegenerative diseases such as Alzheimer's disease (AD) is the sixth leading cause of death in the United States and affects 5.7 million Americans. While cures for this disease have not yet been discovered, several pharmaceuticals are available to alleviate symptoms. These compounds typically target the enzymes acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Specifically, for individuals with AD it has been found that while AChE activity is slightly decreased or unaffected, BChE activity is increased. The increased BChE activity leads to a depletion of the neurotransmitter acetylcholine, and this is implicated in the progression of dementia. Thus, inhibitors of BChE are sought in AD treatment. We previously found Fluorenylmethyloxycarbonyl (Fmoc)‐amino acids selectively inhibit BChE, leading to a potential new class of cholinesterase inhibitors. While the role of the amino acid side chain was explored, the effects of modifying the carboxylate group were not investigated. Specifically, the enzyme binds the cationic substrate acetylcholine, but the Fmoc‐amino acids are anionic. We postulated Fmoc‐amino acid esters may be more potent inhibitors, as the ester ablates the negative charge. In addition, the ester allows incorporation of substituents that increase van der Waals interactions that may interact favorably with the enzyme. To test this model, a series of Fmoc‐amino acids were esterified using an alcohol and EEDQ (N‐Ethoxycarbonyl‐2‐ethoxy‐1,2‐dihydroquinoline), purified by chromatography, and characterized by NMR and HPLC. The solubility and inhibition properties for the Fmoc‐containing esters were evaluated using UV‐Vis spectroscopy. Kinetic studies suggest the Fmoc‐Leu‐esters are more potent inhibitors compared to the Fmoc‐Leu‐O− but are limited by solubility. We are now investigating additional Fmoc‐amino acid esters bearing cationic groups to enhance solubility such as lysine, and our initial results suggest the lysine‐based compounds are potent BChE inhibitors. Overall, the results may guide the design of new Fmoc‐containing compounds that specifically and effectively inhibit BChE.Support or Funding InformationThis project is supported by the National Institute of General Medical Sciences of the National Institutes of Health under Award Numbers R25GM071638, UL1GM118979, TL4GM118980, RL5GM118978, and T34 GM008074.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Targeting Dxr/IspC to develop drugs against malaria and tuberculosis
Authors: Girma Misgina Belachew , Ball Haley , Dowd Cynthia , Couch Robin 
Published: June 2021

The methylerythritol phosphate pathway (the MEP pathway) of isoprenoid biosynthesis is essential for the survival of many pathogenic organisms, including Plasmodium falciparum and Mycobacterium tuberculosis, causative agents of malaria and tuberculosis, respectively. The MEP pathway is absent in humans, making it an important drug target. There are seven enzymes in this pathway. 1‐Deoxy‐D‐xylose 5‐phosphate reductoisomerase (Dxr/IspC) catalyzes the first committed step in the pathway. It converts the substrate 1‐Deoxy‐D‐xylose 5‐phosphate (DXP) into 2‐C‐methyl‐D‐erythritol 4‐phosphate (MEP), which in turn is the substrate for the downstream enzymatic steps. We have expressed and purified IspC of Plasmodium falciparum (PfIspC) and Mycobacterium tuberculosis (MtIspC) using E. coli as an expression system. Finally, we carried out enzyme inhibition assays using novel IspC inhibitors, showing IC50 results ranging from 0.28 – 5.3 μM.Support or Funding InformationSelf‐sponsored.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Mechanism Studies of Thiosemicarbazone Inhibition on Human Topoisomerase IIα
Authors: Lyons Kyle Robert , Toma Maryo , Lisic Edward , Jiang Xiaohua 
Published: June 2021

Thiosemicarbazones (TSCs) has been shown as noncompetitive inhibitors of human Topoisomerase IIα (TopoIIα). Metal‐TSC complexes demonstrated higher activities than their ligands. Metal‐TSC was shown to bind on TopoIIα near the ATP binding pocket. Thus in the absence of ATP, metal‐TSC complexes can increase TopoIIα‐induced cleavage complexes, unlike other inhibitors of TopoIIα. The metal ions of TSC play a predominant role in the activity of TSCs. Cu‐TSC complexes are the most active ones compared to their Pd and Pt counterpart. In this study, we study the activity of metal‐TSC effects on TopoIIα in the presence and absence of ATP molecule and try to understand the mechanism how metal‐TSC interact with TopoIIα.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigations of the Mechanism of Action for Lung Cancer Cell Death by a 4′‐Trifluoromethoxy Substituted Chalcone Derivative
Authors: Stantliff Trevor Martin , Krzysiak Amanda J. 
Published: June 2021

Chalcones are a diphenyl compound that serves as a natural precursor to flavanones in plants.Chalcones have been shown to have anticancer and antimicrobial activities. Chemoprevention activity of chalcones are of high interest in medicinal chemistry because of the simple laboratory synthesis and modification via aldol condensation. Previously this lab created and screened a library of synthetic chalcones against A549 lung adenocarcinoma cell line for antiproliferation properties. We identified a strong drug candidate (4′‐Trifluoromethoxy substituted chalcone) for A549 growth inhibition. However, the cause of inhibition by the substituted chalcone remains to be identified. We began to explore the mechanism of action of this drug by looking at physical characteristics of cell death with microscopy. Then, we conducted viability and cytotoxic assays such as MTS and Trypan blue for cell death quantification. Cytotoxicity was determined with a Lactate Dehydrogenase Assay. Western Blots were ran to identify dose dependency on the specific apoptotic proteins PARP and Caspase‐3. Using flow cytometry, we investigated a more accurate quantification of cells undergoing apoptosis versus necrosis. Here we present the results of our investigation into the apoptotic and necrotic cell death of lung cancer A549 cells by a chalcone derivative.Support or Funding InformationProject Funded by Bellarmine University Department of Chemistry and Physics and Bellarmine University Student Government Association.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Yiqi‐Huoxue Granule reduces pro‐thrombotic factors induced by lipopolysaccharide in HUVECs via upregulating KLF2 and inhibiting NF‐κB
Authors: Wang Zhentao , Wang Xinzhou , Gao Shuibo , WU HONG 
Published: June 2021

Yiqi‐Huoxue Granule (YQHX) is a widely used traditional Chinese formula for the treatment of cardiovascular diseases in clinical practice in China. Our present study explored the effects of YQHX on ameliorating inflammatory events triggered by lipopolysaccharide (LPS) in HUVECs. YQHX significantly suppressed the generation of pro‐thrombotic factors such as plasminogen activator inhibitor‐1 (PAI‐1) and tissue factor (TF), whereas increased the protein expression of Kruppel‐like factor 2 (KLF2). The increase of PAI‐1 and TF were reduced by KLF2 knockdown with Lenti‐shKLF2 vector. Furthermore, YQHX remarkably lowered the phosphorylation levels of nuclear factor κB (NF‐κB) p65 and IκB induced by LPS. In addition, YQHX inhibited the production of PAI‐1 and TF in an NF‐κB‐dependent manner, which is manifested by using PDTC, a specific NF‐κB inhibitor. In short, the results demonstrated that YQHX notably exerts the anti‐thrombotic reaction induced by LPS via inhibiting the upregulation of KLF2 expression and the activity of NF‐κB signaling in HUVECs, suggesting that KLF2 and NF‐κB signaling may be promising therapeutic strategies for intervention of inflammation‐associated atherosclerosis.Support or Funding InformationThis work was financially supported by grants from the National Natural Science Foundation of China (No. 81473453, 81673800), the Science & Technology Innovation Talents in Universities of Henan Province (No. 14HASTIT028), and the Henan Science and Technology Project (No. 142102310040).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Optimization of Shell Grinding Techinques for Purification of Proteins from Zebra Mussel Shells
Authors: Freiermuth Margaret , Grant Jennifer E. 
Published: June 2021

Wisconsin and Minnesota lakes are under attack from invasive Zebra Mussels, Dreissena polymorpha, a freshwater mussel species native to Eurasia. The Zebra Mussels were introduced to the Great Lakes around 1985–1986 and have continued to aggressively invade the surrounding areas of Minnesota and Wisconsin. Outcompeting native species, including fish, for food sources, Zebra Mussels are thriving. Oddly, this confluence of attributes, ie fast growth but undesired presence, offers a unique opportunity to capitalize on the economic potential of this invasive species' presence in Wisconsin. Here we examine the protein content found in the Zebra Mussel's shell, and optimization of the protocol for extracting proteins from this matrix, with particular emphasis on techniques for grinding the shells. This will be the first step in determining a possible commercial use in a large pool representing an underutilized potential resource that is also a pest.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Towards Extraction of Proteins from the Shells of Dreissena polymorpha
Authors: Junaidi Nabil , Grant Jennifer E. 
Published: June 2021

The zebra mussel, Dreissena polymorpha, is one of the most invasive aquatic species of Midwest United States. Its biogeochemical impact on American waters is regarded as a quintessential example of a difficult and current ecological problem, since there does not exist a working management scheme to control its biofouling and parasitic effects. Yet, the untapped proteomic diversity of aquatic species, especially freshwater mollusks, leave the zebra mussel as an attractive resource for the large‐scale screening of peptides that may confer selective advantages to its life cycle, such as antibiotic resistance. The first step along this pathway is to optimize procedures for extracting functional peptides from zebra mussel shells. Here we discuss basic research aimed at identifying successful extraction of proteins using detergent‐based strategies.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
New tools for autoimmune kidney disease: Design of antigen binding proteins against the thrombospondin type‐1 containing domain 7A
Authors: Stoddard Serena Daniella , Stoddard Shana Victoria 
Published: June 2021

Idiopathic membranous nephritis, (IMN) is an autoimmune disease (AD) affecting the kidney. Conventional treatment for AD uses immunosuppressive medicines that suppress the entire immune system, rather than target specific autoimmune disorder antibodies. This causes patients to become immunocompromised, and at risk of not being able to fight off common infections. A more specific therapy to treat AD is needed to prevent the suppression of the entire immune system. Thus the goal of this work was to develop antigen binding proteins (ABP) which could prevent autoantibody binding to the antigen of interest in an AD. ABP targeting the thrombospondin type‐1 containing domain 7A (THSD7A) antigen of IMN were developed. Domain 2 of THSD7A was used as the target site for binding. These designs were carried out through in silico mutagenesis of template monobody candidates. Homology modeling of designed proteins was performed using the Phyre2 server for all ABP. The Rosetta docking2 protocol was used to determine strength of protein‐protein interactions between the ABP and domain 2 of THSD7A. Molecular interactions were evaluated to determine which mutations strengthened binding of the ABP to the THSD7A. Several mutations were shown to improve overall binding compared to the template monobody (3UYO) which was found to have an interface score of −2.511, measured in Rosetta energy units (REU). The most promising binding efficiency scores and placement of an ABP was shown to be the C‐side location of the 3UYO ABP template. Introduction of an electrostatic interaction through the single point mutation S88D, produced an interface score of −5.557 REU. The four point mutation G43W, T72R, S88D, S91D introducing three electrostatic interactions, one hydrophobic interaction, resulted in an interface score of −4.793 REU. Currently protein expression and experimental testing is underway to evaluate binding in vitro. This work will assist in the design of antigen specific therapies for AD, leading to a more targeted therapeutic suppression of the immune system.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Development of antigen‐specific blocking monobodies against the β2‐glycoprotein I receptor for the inhibition of systemic lupus erythematosus
Authors: Rasasack Itthipoaln , Stoddard Shana Victoria 
Published: June 2021

Autoimmune diseases occur when the body's immune system attacks healthy tissues and organs through the use of antibodies. Current treatments suppress the patient's entire immune system through immunosuppressive drug therapies. These drugs decrease the body's effective immune response to genuine pathogens, increasing the patient's susceptibility to having major complications from basic infections. Therefore, treatments that can eliminate disease progression, as well as preserve the healthy function of the patient's immune system, should be considered. Systemic lupus erythematosus (SLE) is a systemic autoimmune disorder affecting a predicted five million individuals worldwide. One autoantigen targeted in SLE is the β2‐glycoprotein I receptor (β2GPI). The work here focuses on the development of protein monobodies capable of inhibiting interactions between the autoantibody and autoantigen that elicit the immune response. Binding between the monobody and autoantigen may prevent antibody binding, decrease autoimmune complexes, and ultimately halt disease progression. Using in silico programs Phyre2 and Chimera, mutations to the monobody templates 3K2M, 3T04, and 5A43 were introduced to improve binding between the monobody and proposed antibody‐binding β2GPI domains 1, 2, and 5. In silico program ROSIE was used to predict protein‐protein binding scores between the monobody candidates and proposed β2GPI domains. Initial binding scores (in Rosetta Energy Units, or REU) between monobody templates 3K2M, 3T04, and 5A43 with β2GPI domain 5 were −3.105 REU, −2.795 REU, and −3.038 REU, respectively. A double mutation to 5A43 (V46R‐E48H) improved 5A43's binding score with β2GPI domain 5 to −4.122 REU. A slight improvement to binding was made to 3K2M with a single mutation (N47H), with a score of −3.446 REU to β2GPI domain 5. More mutations to each monobody template are being evaluated. Enzyme‐linked immunosorbent assays (ELISAs) will be conducted to evaluate antibody inhibition by monobody candidates in vitro. The goal of the research presented here is to develop safer and effective treatments of autoimmune disease.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Computational Approaches for the Analysis of Tandem Affinity and Proximity‐Based Purifications in R
Authors: Velasquez Erick Francisco , Garcia Yenni , Gao Lucy , Whitelegge Julian , Torres Jorge 
Published: June 2021

During the past decade, the incorporation of mass spectrometry data to identify protein‐protein interactions and associations has become extremely popular, and presents an exciting opportunity to exploit the protein interactome. However, the experimentation often produces large amounts of proteomic data that can be hard to handle, delaying scientific discovery. It has become clear that when handling the data, cleaning the data of false positives is as important as verifying associations. To address this issue, scientists have developed databases like the Contaminant Repository for Affinity Purification (CRAPome) that provide spectral count data for a collection of controls. However, the data can be hard to interpret without a more rigorous statistical analysis. This work describes an open source proteomic pipeline that incorporates tools to identify false positives and visualize interactions/associations in a user friendly manner. Taking advantage of R, a popular statistical programming language, and Shinny apps we developed a program that aims to standardize analysis of protein interaction/association experiments. The pipeline allows scientists to submit proteomic data and easily apply suggested significance tests to clean their experiments against their own control and the CRAPome if they wish to do so. Often these types of experiments are done through a purification step and so we have extended the platform to include predictive machine learning algorithms that highlight false positives based on intrinsic data that describes a protein that might stick to the column. After analysis, is carried out we couple the platform to protein interaction network visualization tools such as Cytoscape, to create a comprehensive analysis pipeline that is user friendly. In the future we plan on extending this platform by applying neuronal networks to further predict protein‐protein interactions, adding to a fully automated pipeline, for the analysis of protein‐interaction/association experiments.Support or Funding InformationThe work was possible through USPHS National Research Service Award 5T32GM008496 and the Whitcome Pre‐doctoral Training Program and the UCLA MolecularBiology InstituteThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
An Integrated Framework for Genome Analysis Reveals Numerous Previously Unrecognizable Structural Variants in Leukemia Patient Samples
Authors: Broach James , Xu Jie , Schleicher Emily , Song Fan , Pool Christopher , Hennessy Max , Sheldon Kathryn , Annageldiyev Charyguly , Sharma Arati , Chang Yuanyuan , Hastie Alex , Miller Barbara , Goldenberg David , Claxton David , Moldovan Geroge‐Lucian , Yue Feng 
Published: June 2021

We have applied a combination of whole genome sequencing and optical genomic mapping to a number of adult and pediatric leukemia samples, which revealed in each of these samples a large number of structural variants not recognizable by current standard genomic analyses. We developed computational methods to determine which of those variants likely arose as somatic mutations. The method identified 97% of the translocations previously reported by karyotype analysis of these samples and revealed an additional fivefold more such somatic rearrangements. The method identified on average tens of previously unrecognizable inversions and duplications and hundreds of previously unrecognizable insertions and deletions. These structural variants affected a number of leukemia associated genes as well as cancer driver genes not previously associated with leukemia and genes not previously associated with cancer. A number of variants only affected intergenic regions but caused cis‐acting alterations in expression of neighboring genes. Analysis of TCGA data indicates that several of the recurrently mutated genes significantly affect survival of AML patients. Our results suggest that current genomic analysis methods fail to identify a majority of structural variants in leukemia samples and this lacunae may hamper diagnostic and prognostic efforts.Support or Funding InformationNIH grants R35GM124820, R01HG009906, and U01CA200060; Four Diamonds Fund; George Lafferty FoundationThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
GLP1 targets lncRNA Gas5 in diabetic adipocytes
Authors: Lui Ashley , Kraja Gearta , Rupani Rea , Bader Deena , Patel Niketa 
Published: June 2021

Diabetes type 2 affects 10% of the US population whereas 80% of patients affected are insulin resistant. We previously showed that lncRNA Gas5 was downregulated in the serum of T2DM patients. Here we sought to investigate the underlying molecular mechanisms of Gas5. Our data shows that Gas5 is down regulated in type II diabetic adipocytes and their secretome. Depletion of Gas5 in non‐diabetic adipocytes resulted in decreased glucose uptake. Using an unbiased diabetes transcriptomics screen, we demonstrated that this reduction of Gas5 resulted in a decrease of both insulin receptors A and B. Glucagon‐Like Peptide‐1 mimetics are new therapeutics available for diabetic patients and have recently been seen as the leading treatment for patients who are no longer responsive to metformin. Our data has shown treatment with GLP‐1 in adipocytes results in an increase in Gas5 resulting in an increase in glucose uptake. Gas5 has been recognized as a biomarker for T2DM, but this is the first time it has been studied mechanistically in adipocytes and researched as a therapeutic target.Support or Funding InformationVAMRThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.