bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effects of Hydrogen Bond Donors on Reactivity of Sulfite Oxidase
Authors: Bali Sofia 
Published: June 2021

Sulfite oxidase (SO) catalyzes the conversion of sulfite to sulfate as the last step in the degradation of cysteine and methionine [1]. Severe neonatal neurological problems are caused by SO deficiency in humans [2]. The overall mechanism of SO suggested that the reductive half reaction of SO is the rate limiting step for enzyme activity [3]. Using a computational approach, first I will study the elementary steps for the half reaction to determine the rate limiting step, then by introducing mutations in the active site of SO I will determine the effects of the hydrogen donors present in the active site.SO is part of the Molybdenum cofactor family of enzymes as it contains a mononuclear Molybdenum (Mo) that is penta‐coordinated [4]. At the Mo active site, the reductive half reaction of sulfite to sulfate takes place in two steps. The first step is a nucleophilic attack by the sulfite ion to the equatorial oxygen group. The second step is the release of sulfate facilitated by the spectator oxygen group [4]. By determining the energy profiles for both steps in this reaction I will determine which step is rate limiting and determine the route, associative or dissociative interchange, of the reaction.Model enzyme complexes have suggested that hydrogen donors in the active site play a key role in mediating this reaction. In previous studies performed we determined that the product release step is facilitated by pyridinium a hydrogen donor for the reaction. Pyridinium lowers the activation energy for the product release step (ΔH~ 30 kJ/mol) and promotes a dissociative interchange reaction route. In this study I will determine if hydrogen donors in the Mo active site behave as pyridinium does in the model enzyme complexes.Using Gaussian ONIOM to perform QM/MM calculations for the reaction of sulfite to sulfate catalyzed by SO, I will determine the energy profile of the reaction to identify the rate limiting step and the role of possible hydrogen donors in the active site of SO by comparing the reaction mechanism of two variants of SO with wild type SO. The variants will demonstrate the role as possible hydrogen bond donors of Tyrosine and water that are located in the active site of MO.The role of sulfite oxidase is crucial to the degradation of sulfite in the body and prevention of the accumulation of toxic sulfite in the body. Understanding how SO functions at a molecular level is crucial for the design and implementation of treatment for SO deficiency. The elementary steps of the enzyme activity can help determine specific roles of key residues in the enzyme. These steps cannot be seen in large scale enzyme kinetics studies, but computational studies allow the study of small movements and allow the study of these small‐scale reaction mechanics.Support or Funding InformationLouis Stokes Alliance for Minority Participation (LSAMP) HRD # 1305011This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Antioxidant and Ameliorative Potential of Aqueous Seed extracts of Delonix regia on High Fat Diet and Streptozotocin Induced Diabetes in Female Wistar Rats
Authors: ONYEGEME‐OKERENTA BLESSING MINAOPUNYE , OGBOYE PHILIP OSEIKHUENMHEN , MONAGO‐IGHORODJE COMFORT CHINAZO , MONAGO IFEOMA NWAMAKA 
Published: June 2021

Antioxidant and ameliorative potential of aqueous seed extracts of Delonix regia on high fat diet (HFD) and streptozotocin (STZ) induced diabetes in female wistar rats were investigated. Forty eight rats were grouped into eight, consisting of six per group. Diabetes was induced in all the groups except for group 1 which was used as normal control (NC) and was fed with normal diet. Diabetic state was achieved by feeding the rats with HFD which contained 20% sucrose, 20% lard and 60% grower mash for six weeks, followed by 40mg/kg body weight (BW) of a single dose intraperitoneal injection of STZ. Seven days after induction of diabetes, treatment of cooked Delonix regia seeds (A) and uncooked Delonix regia seeds (B) extracts commences and this lasted for six weeks. Rats in groups 3(MET) and 4(MET+VDG) received metformin 100 mg/kg and metformin/vildagliptin 50/25 mg/kg (BW) respectively as standard drugs. While the rats in groups, 5 (A1), 6(A2) and 7 (B1), 8(B2) were induced and treated with A and B extracts respectively by receiving 150 and 300 mg/kg (BW) orally and daily. Blood was obtained through cardiac puncture after the rats were anaesthetized and sacrificed. Proximate analysis showed that the seeds contained, moisture, ash, lipid, protein, carbohydrate and fiber contents of 3.83 ± 0.07, 3.50 ± 0.00, 7.82 ± 0.02, 17.50 ± 0.00, 21.79 ± 0.21 and 45.57 ± 0.26 respectively. The treatment with both extracts A and B significantly decreased (p<0.05) the fasting blood glucose concentration and pancreatic amylase activities in dose and time dependent manner, when compared to the DC. Similarly, treatments with A and B extracts significantly decreased (p<0.05) the pack cell volume (PCV), and platelets (PLT) count with, a significant decreased (p<0.05) in the granulocyte count in a dose dependent manner. The potassium and bicarbonate concentrations of the treated diabetic rats significantly decreased (p<0.05) when compared to the DC except for group 5(A1) extract treated group. Results obtained for superoxide dismutase (SOD) showed a significant increase (p<0.05) in activities of treated diabetic rats in a time dependent manner when compared to DC. Conclusively, treatments exhibited immune boosting and antioxidant properties, protective against disturbance of acid‐base balance, in treated diabetic rats. Therefore, the plant exhibited very impressive potency and promise in the management of diabetes and its complications and hence a potential source for the discovery of new orally active anti‐diabetic drugs and alternative source for food nutrientsSupport or Funding InformationSelf fundingThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
SuFEx based click chemistry for peptide stapling
Authors: Han Jennifer , Celaje Jeff , Thomas Joice , Fokin Valery 
Published: June 2021

Click chemistry is a chemical synthesis strategy which relies on covalent assembly of functional building blocks using high‐yielding reactions that are broad in scope, chemoselective, and compatible with water and oxygen. The sulfur (VI) fluoride exchange (SuFEx) is the latest addition to the click family due to the unique stability of the SO2F moiety, which confers inertness against acid‐base reactions, resistance to oxidation and reduction, and generally high yields of sulfonylated products. Ethenesulfonyl fluoride (ESF) is a powerful Michael acceptor for installing the SO2F group. We are exploring the use of SuFEx chemistry as a powerful tool in peptide stapling, a technique that locks peptide conformation to improve its pharmacological properties. Specifically, we take advantage of a complementary reactivity of two neighboring nucleophilic side chains of a peptide which can undergo a conjugate addition to ESF, followed by fluoride exchange, thus forming a chemical “staple” and locking a specific conformation of the resulting peptide. Such selective modification can inform the development of chemical probes to improve efficacy of a peptide therapeutic, may improve drug delivery, and facilitate the discovery process. Preliminary studies described herein demonstrate feasibility of this approach.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Proline‐Rich Tyrosine Kinase Phosphorylation's Effect On the Na+/H+ Exchanger Isoform 1
Authors: Bagnell Kyle P. , Provost Joseph J. , Wallert Mark 
Published: June 2021

The Na+‐H+ exchanger isoform 1 (NHE1) is a 12‐pass transmembrane protein that functions by exchanging one extracellular sodium ion for one intracellular proton. The transmembrane domain of NHE1 is involved in ion exchange, and the cytoplasmic tail is the regulatory domain which includes a series of phosphorylation sites as well as lipid and proteins binding sites that alter transport activity. NHE1 is a central driver of the hallmarks of cancer through its involvement in the regulation of cellular proliferation, migration, and invasion. While the exact number of phosphorylation sites on the cytoplasmic domain of NHE1 is still unclear, many studies have implicated 22 phosphorylation sites and 12 different kinases. One of the kinases believed to directly or indirectly lead to the phosphorylation of NHE1 at S602 and S605 is proline rich tyrosine kinase 2 (PyK2). To evaluate the role of PyK2 in NHE1 regulation we prepared three cell lines derived from Chinese hamster lung fibroblast lacking NHE1 expression (PS120 cells). These cell lines are PSN (PS120 cells expressing human NHE1), PSN S602A/S605A (PSN cells with serine to alanine mutations of the reported PyK2 phosphorylation sites) and PSN S602D/S605D (PSN cells with serine to aspartic acid mutations of the reported PyK2 phosphorylation sites to mimic phosphorylation). Some studies have found that PyK2 works inversely on NHE1 and causes inhibition upon phosphorylation, while other studies have shown PyK2 to enhance NHE1 activity. Here we evaluate the impact of PyK2 inhibition and mutation of the PyK2 phosphorylation sites on proliferation and migration. Western blot analysis is used to evaluate changes in Pyk2 expression and phosphorylation over time under low and normal serum conditions. Cell proliferation and cell migration are both evaluated. Our results suggest that serum stimulation increases Pyk2 phosphorylation and enhances both cell proliferation and cell migration. Inhibition of PyK2 by PF43I396 or the mutation of the Pyk2 phosphorylation sites on NHE1 decreases proliferation and migration rates in serum stimulated cells.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Mapping Domain Interaction Networks in Protein Kinases with Optical Tweezers
Authors: Maillard Rodrigo 
Published: June 2021

Signaling proteins are dynamic molecular switches that sample multiple conformational states. Such conformational plasticity allows these proteins to adapt and respond to different biological signals. Protein Kinase A (PKA) is a signaling protein complex composed of regulatory and catalytic subunits. The regulatory subunit harbors two cAMP binding domains (A and B) that oscillate between inactive and active conformations depending on cAMP concentration. In this study, we use optical tweezers to identify the networks of communication between cAMP binding domains that enable PKA to switch efficiently and cooperatively from inactive to active states. We show that in the inactive PKA conformation, domain B of the regulatory subunit serves as an energetic hub, controlling the magnitude of interaction between domain A and the catalytic subunit, and whose mechanical perturbation triggers the highly coordinated dissociation of the PKA complex. In the presence of cAMP, domains A and B establish an intricate network of communication that involves direct, interfacial domain contacts as well as long‐range interactions between non‐contiguous structural motifs. In contrast, without ligand the two domains behave as independent, non‐interacting structures. Lastly, we demonstrate the application of optical tweezers to dissect how loss‐of‐function mutations can disrupt very distinct communication networks in PKA. The experimental approach based on optical tweezers should be readily applicable to study disease mutations, the mechanism of action of small molecules, or other signaling proteins.Support or Funding InformationNSF MCB 1715572This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
MK5 and ERK3 play overlapping but distinct roles in regulating cardiac fibroblast function
Authors: Sahadevan Pramod , Nawaito Sherin A. , Calderone Angelo , Gaestel Matthias , Allen Bruce G. 
Published: June 2021

Background and objectivesImmunoreactivity for MAP kinase‐activated protein kinase‐5 (MK5), a protein serine/threonine kinase, is detected in cardiac ventricular fibroblasts but not myocytes. p38 and ERK3/4 MAP kinases are activators of MK5. However, the regulation of MK5 remains controversial and its physiological functions are poorly understood. Following hemodynamic overload, the increase in collagen mRNA was attenuated in ERK3 and MK5 haploinsufficient mice. In addition, following myocardial infarction, scar size and collagen content were reduced in MK5+/− mice, compared with MK5+/+ mice. In addition, MK5−/− fibroblasts show reduced motility and proliferation. Thus, MK5 may play a role in fibroblast function. The present study was to examine the role of ERK3, a putative activator of MK5, in cardiac fibroblast function.MethodsCardiac ventricular fibroblasts were isolated from MK5+/+ mice and male Sprague‐Dawley rats. Subconfluent cultures of fibroblasts from passages 2 and 3 were used. Ventricular myocytes were prepared from adult MK5+/+ mice. Protein expression was determined by immunoblotting. The presence of ERK3‐MK5 complexes was determined by proximity ligation assay (PLA) and co‐immunoprecipitation assays. Cell motility and myofibroblast contraction were studied by scratch‐wound and collagen gel contraction assays, respectively.ResultssiRNA‐mediated knockdown of MK5 (MK5‐kd) resulted in reduced contraction of collagen gels, reduced cell spreading, and formation of fewer dendritic extensions compared to fibroblasts transfected with scrambled RNA. ERK3 immunoreactivity was detected in fibroblasts but negligible in cardiomyocytes. MK5 immunoprecipitates from fibroblast lysates contained ERK3 immunoreactivity. Proximity ligation assays revealed ERK3‐MK5 complexes in the cytoplasm, which were less abundant following knockdown of MK5. ERK3‐MK5 complexes were not observed in the nucleus. Cell migration, in response to serum and/or angiotensin II, was reduced upon siRNA‐mediated knockdown of ERK3. However, whereas the secretion of type 1 collagen and fibronectin was increased in MK5−/− or MK5‐kd cells relative to MK5+/+ fibroblasts, knocking down ERK3 did not affect collagen secretion.ConclusionMK5 and ERK3 immunoreactivity was detected in fibroblasts but not myocytes. Co‐immunoprecipitation and PLA suggest that ERK3 and MK5 form cytoplasmic complexes in cardiac fibroblasts. Suppressing ERK3 or MK5 expression decreased cell motility; however, type 1 collagen secretion was enhanced by reduced MK5 expression but unaffected by knockdown of ERK3. Hence, MK5 and ERK3 play overlapping but distinct roles in regulating cardiac fibroblast function.Support or Funding InformationThis study was supported by a grant from the Heart and Stroke Foundation of Canada.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Teaching Biochemistry based Research Driven Course through Active Learning to Accelerate Student's Adaptation to College Study and Promote Freshman Research
Authors: Ray Supriyo , Fresquez Joseph , Esper Ronda , Clark Jeni , Corral Guadalupe , Xiao Chuan 
Published: June 2021

Freshman Year Research Intensive Sequence (FYRIS) program was implemented at The University of Texas at El Paso (UTEP) in 2015. The program promotes student enrollment in research driven course (RDCs) for two semesters to enhance student development and increase retention in STEM through early interventions by engaging them in research activities during the freshman and sophomore years. As the undergraduate population at UTEP is 85% Hispanic, programs such as FYRIS can play an important role in increasing the representation of minorities in STEM. UTEP has a near 100% acceptance for the undergraduate courses implies a class population represents students of diverse academic proficiencies. In our special freshman chemistry laboratory sections (CHEM 1105 and CHEM1106/4176), students conduct research on expression and purification of proteins. In the first semester, students gain understanding of basic chemistry and biochemistry concepts prior optimizing protein expression. In the second semester, they continue expressing proteins and also learn protein purification using Fast Protein Liquid Chromatography. To enhance student success in advanced research driven courses, it's important that all students have an accelerated understanding in the basic chemistry, biochemistry and molecular biology. Active learning methodologies such as personal response systems, case studies, problem based learning, flipped classroom, model building, game based quizzes, think pair share, peer led team learning were used to promote accelerated learning. Research‐Teaching Integration through peer led undergraduate students were implemented to boost student understanding and confidence. Overall, our evaluation showed that students' enjoyed the class sessions more and had a better clarity of complex research based concepts in our newly designed research driven coursesSupport or Funding InformationRDCs; S.R.; J.F. & R.E. National Institute of General Medical Sciences of the National Institutes of Health under linked Award Numbers RL5GM118969, TL4GM118971, and UL1GM118970.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Three web tools to aid genetics instruction developed by students in a unique undergraduate bioinformatics projects course
Authors: Rutherford Robert , Garcia Nic , Charczenko Richard , Levy Samuel , Bjelajac Jeremy 
Published: June 2021

At Seattle University, some of our undergraduate Cellular and Molecular biology (CMOL) majors enroll in 10 week bioinformatics course. In addition to learning to use a range of bioinformatics tools written by the research community, students learn to use Python programming language. Some of these students take a second course, where they work in teams to develop novel biological software to support the teaching and learning of threshold genetics concepts. Here we describe this method of teaching bioinformatics and programming to CMOL majors and also features three student projects that have extended beyond this class to become independent and ongoing research endeavors. These web ready, graphical software projects include: 1) EVO.py, a new pedagogical tool that simulates genetic drift and natural selection in snowshoe hare populations. Users control a multitude of variables, including climate, allele frequency, and population size, and run the simulation, observing how procedurally generated snowshoe hare populations evolve over time in response to those inputs. 2) Lactose Operon (LacOp), a new program that simulates the regulation and biochemistry of the E.coli Lac operon. LacOp allows students configure this pathway with a wide range of mutations and/or substrate concentrations and then observe the full set of molecular outputs through time in a graphical format. 3) An enhanced FaSE (FauxSequenceEvolver) tool, which subjects user supplied molecular sequence(s) to faux evolution, automating the generation of novel but biologically credible derivative sequences. These derivative sequences have been used, for example, as ‘unknown sequences' suitable to develop and test a classroom of students' abilities to use bioinformatics tools for characterization and annotation. Student software development on this project included integrating FaSE with BioPython to allow NCBI interactivity, to capture and import known characterization data, and to automate scanning for domains.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Determination of Apoptotic Stage of HCC827 Lung Adenocarcinoma Cell Line, Following Calcium Sulfide Nanoclusters Treatment
Authors: Rodriguez Irizarry Valerie J. , Torres Geraline Trossi , Forti Kevin Muñoz , Rivera Abigail Ruiz , Castro Miguel E. , Suarez Martínez Edu B. 
Published: June 2021

Lung cancer is the number one cause of death independently of gender. By the time of the initial diagnosis, more than 50% of all lung cancer patients have already developed secondary tumors due to metastases. Despite current treatments such as surgery, chemotherapy, and radiotherapy, the prognosis for these patients still poor. Subsequently, there exists the need for more efficient treatments in order to increase the survival rate of patients suffering this disease. Previously, in‐vitro experiments in our lab demonstrated that Calcium Sulfide (CaS) nanoclusters induced apoptosis and necrosis in the non‐small lung cell adenocarcinoma (NSCLC) cell line HCC827. Using the Flow Cytometry technology, in this project we aimed to determine the effect of CaS in these cells evaluating apoptotic markers including, Caspase‐3/7 activity, Bcl‐2 phosphorylation, mitochondria potential, DNA‐damage, and oxidative stress. We hypothesized that CaS will show significant differences in these assays when compared to the Dimethyl sulfoxide (DMSO) control. We added a single dose of CaS [3.8μM] (experimental treatment), DMSO (control vehicle) or Etoposide [10 nM] (positive control) at time 0 and then collected the data after 24, 48, and 72 hours. As our results, we did not observe any statistically significant effect on apoptotic markers, Caspase‐3/7 and Bcl‐2. Analysis of the Mitopotential assay showed an statistical increase (p≤0.01) of dead cells population at 48 hours and of depolarized/lives cells at 72 hours compared to DMSO, in HCC827 cells treated with CaS. However, at 48 hours cells treated with CaS decrease the numbers of non‐affected cells (p≤0.05), increase DNA double‐strand breaks (p≤0.001), decrease the activation of Ataxia‐telangiectasia mutated kinase (ATM) and the phosphorylation of H2A.X significantly (p≤0.01). These results suggested that the detrimental effect of CaS on malignant HCC827 cell line is influenced by the accumulation of oxidative stress damage also affecting the cell cycle as our lab previously reported. Additional studies are ongoing to validate these results. Within the next month we plan to study the lung non‐malignant cells MRC‐5 using the same assays.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.