bluuebunny/crossref_metadata_2025_split huggingface dataset:
Crystal Structure and Bioinformatics Inferred Function of Protein A2617 from Methylibium petroleiphilum
Authors: Saleh Ali , Collazo Michael J. , Sawaya Michael , Cascio Duilio , Rasche Madeline 
Published: June 2021

The attempt to elucidate the biological function of enzymes involved in the biosynthesis of tetrahydromethanopterin for the use in one‐carbon metabolism by archaea and select bacteria has resulted in the crystal structure of the hypothetical protein A2617 from Methylibium petroleiphilum. The structure was initially solved at 2.0 Å resolution using a selenomethionine multi‐wavelength anomalous diffraction experiment, and further refined at 1.8 Å resolution using a native dataset. The Rfactor of the current model is 0.19 and the Rfree is 0.22. The crystal structure of A2617 is a homodimer with each monomer consisting of four α‐helices connected to a barrel fold of six anti‐parallel β‐sheets with an α‐helix by a pair of anti‐parallel β‐sheets. The monomers interface to form the homodimer at the region containing four α‐helices with the barrel folds flanking both ends of the structure. A deep cleft is present at the interface of the two monomers. The overall structure of A2617 resembles that of uncharacterized archaeal homologs; however, the structure is distinct due to the lack of bound FMN in the binding pocket that is found in the archaeal homolog crystal structures. Ligand docking studies using intermediates in the tetrahydromethanopterin biosynthetic pathway indicated that A2617 has the greatest binding affinity for 6‐hydroxymethyl‐7,8‐dihydropterin‐ribofuranosylaminobenzene‐5‐phosphate. This metabolite is the substrate for the third step of tetrahydromethanopterin side chain biosynthesis, which is an oxidoreductase reaction. The homology of A2617 with FMN‐containing archaeal proteins and the findings of the ligand binding studies provide support for the role of A2617 in catalyzing a reductive reaction in the biosynthesis of the tetrahydromethanopterin side chain.Support or Funding InformationThis research was supported by National Science Foundation grant number CHE‐1508801 and by a grant from the California State University Program for Education and Research in Biotechnology (CSUPERB).
A Fluorescent, HTS‐Adaptable Coupled‐Enzyme Assay for Measurement of Human Cytosolic Sulfotransferase (SULT) 2B1b Activity and Identification of Small‐Molecule Inhibitors
Authors: Zhou Qing , Mesecar Andrew 
Published: June 2021

Human cytosolic sulfotransferase (SULT) 2B1b, as a member in the SULT family, catalyzes the transfer of a sulfonate moiety (‐SO3) from the universal donor 3‐phopho‐adenosine 5′‐phophosulfate (PAPS) to the hydroxyl groups of various substrates, which leads to a change in water solubility and thus triggers various downstream effects. SULT2B1b highly prefers 3β‐hydroxysteroids, such as cholesterol, as its substrates. By switching the sulfonation state of its substrates, SULT2B1b is involved in the regulation of steroid synthesis, transportation and signaling. Furthermore, SULT2B1b has been suggested as a potential drug target for prostate cancer. Therefore, we aim to identify small‐molecule inhibitors from high‐throughput screening (HTS), and utilize them to study the physiological roles of SULT2B1b in both normal tissues and carcinogenesis as well as provide possible leads for drug discovery in the treatment of prostate cancer. Considering the lack of convenient and HTS‐adaptable activity assays for SULT2B1b, we have developed a fluorescent, coupled‐enzyme assay with real‐time fluorescence‐monitoring capabilities as opposed to the commonly used endpoint radioactivity assays. The new assay was adapted for HTS in 384‐well plates and after non‐trivial assay optimization, this activity assay resulted in a reliable performance (Z′‐factor=0.8 in preliminary study, Z‐factor=0.6~0.7 in the actual screening). Initial hits from HTS have been further validated in vitro and in cellulo. Our criteria for highly selective inhibitors of SULT2B1b include high potency in vitro, and success in mimicking the effects of SULT2B1b siRNA in cellular studies, including attenuating prostate cancer cell proliferation and decreasing cholesterol sulfate accumulation.Support or Funding InformationThis work is funded by a grant from the Department of Defense.
Expectations from Low‐Income Adults Regarding an Innovative Nutrition Education eLearning Program
Authors: Stotz Sarah , Lee Jung Sun , Hibbs Judy , White Alice 
Published: June 2021

As a step in a comprehensive multi‐year formative and process evaluation of a newly developed eLearning nutrition education program targeted to SNAP‐Ed eligible adults in Georgia (southeastern USA), this study intended to obtain qualitative understanding on expectations and desires of the program from the point‐of‐view of members of the target audience regarding both format and content. Low‐income adult Georgians were recruited to participate in a series of two focus groups. This study reports the findings from the first in this series based on the participants’ expectations of the eLearning program, which informed the iterative development process of the program, titled Food eTalk. Four focus groups were conducted in a convenience sample (n=25, age 41.5 ± 15.5 years, 96% female, 60% Black, 20% Hispanic). Participants from both rural and urban areas of Georgia described their Internet access as reliable and easily available both at home and through multitude of free WiFi sources in their communities. They discussed inexpensive online access though their smartphone using free WiFi or at a computer at the public library. When asked about the format of Food eTalk, participants prefer short lessons with interactive activities, videos, and the option to pause and resume a lesson where they left off. In response to what content they wanted in the Food eTalk lessons, participants discussed the need to expand beyond traditional basic nutrition education. Every focus group included a participant‐generated discussion about the specific need for diabetes diet information. Participants suggested that basic nutrition education, such as Choose My Plate, does not provide enough information to increase their knowledge nor does basic nutrition education address their actual barriers to eating healthfully. Barriers to eating healthfully include: limited food budget, high cost of fresh produce and low cost of ‘processed’ food, cultural taste preferences that have become habitual though generations, and resistance to change with regard to diet practices. To mitigate the last two barriers, participants discussed the need for education on how to cook healthier meals and tips on how to acculturate their children to prefer healthier foods. Finally, food safety was discussed at length and included concern about safety/hygiene of food when dining out, distrust in genetically modified food, and fears of antibiotic/steroid use in animals raised for food. The implications of this qualitative study are to assist in developing a user‐centered nutrition education eLearning program using an iterative process including members of the target audience in program development.Support or Funding InformationThis project is funded by USDA SNAP Ed
Recruitment Strategies Implemented Across a Four‐State Lifestyle Intervention: Get Fruved Study
Authors: Barr Makenzie L , Olfert Melissa D. , Zhou Wenjun , Riggsbee Kristin , Mathews Anne EW , Vilaro Melissa , Kattelmann Kendra K. , Andrea Hanson , White Adrienne , Horacek Tanya , Kidd Tandalayo , Brown Onikia , Colby Sarah E 
Published: June 2021

The objective of the ‘Get Fruved’ research project was to enroll college students from four universities into two courses. This community based participatory research (CBPR) multi‐university, peer‐led, social marketing and environmental intervention is aimed at improving healthful lifestyles (primarily focused on diet, stress management and physical activity) and changing campus environments to promote obesity prevention with older adolescent populations (students entering their first year of college). The four intervention universities (FL, SD, TN, and WV) attempted to recruit 30 social marketing and environmental intervention (SMEI) students, 50 peer mentors (PM), and student researchers per state (minimum of 320 students) over 5 months (August‐December 2015). Recruitment strategies included: emails, student digital announcements, talking with administrators, speaking in classes, informational booths, chalking sidewalks, flyers posted around campus in high traffic areas and word of mouth. Strategies for each state were logged weekly. The recruited students became 4‐H collegiate members and took a 3‐credit hour (SMEI) or a 1‐credit hour (PM) multi‐university course, and/or assisted with research. In these courses the students developed the intervention using a CBPR approach in collaboration with researchers and Extension Professionals. The baseline characteristics of the recruited SMEI, PM, and student researchers were assessed using demographic and behavior surveys and anthropometric data was collected. Weekly recruitment strategy logs were analyzed and states were found to have similar approaches. SMEI course (total of 74) signed up and 240 for the PM course. Approximately 60 undergraduate researchers, 40 graduate researchers, and 10 Doctoral students were recruited to work on the project as student researchers. Only 280 participants completed the survey process. Demographics between SMEI, PM, and student researchers across states were found to be statistically similar. Participants (N=280) were predominately Caucasian (84.4%) and female (87.89%), and 75.3% were 18–20 year olds. Most participants were sophomores (39.84%) followed by juniors (34.38%), freshmen (18.75%) and one graduate student (0.39%). Recruitment strategies resulted in a sample that exceeded minimum goals for PM but fell short for SMEI. Using a CBPR approach to develop partnerships with motivated students was essential to effectively plan and execute an intervention to promote healthful lifestyles on these college campuses that may help to prevent the development of obesity.Support or Funding InformationFunding support USDA/AFRI #2014‐67001‐21851
Metabolic Fate of Curcumin in the Gastrointestinal Tract in Mice
Authors: Li Zhengze , Sun Yue , Song Mingyue , Li Fang , Xiao Hang 
Published: June 2021

Curcumin (1,7‐bis (4‐hydroxy‐3‐methoxyphenyl)‐1,6‐heptadiene‐3,5‐dione) is a major yellow‐orange pigment found in turmeric, and has been used widely as a food ingredient. Curcumin has been associated with many health benefits, but its health‐promoting potential is limited by its poor bioavailability. This is due to the fact that curcumin is poorly absorbed, and absorbed curcumin is subject to extensive biotransformation. The detailed understanding of biotransformation of curcumin is needed to facilitate the elucidation of its biological significance. This study was aimed to establish the metabolic fate of curcumin in the gastrointestinal (GI) tract. The male CD‐1 mice were fed with curcumin in diet (0.05% w/w) for 5 weeks. The entire GI tract and its content were harvested for LC‐MS analysis. Our results demonstrated that curcumin underwent phase I metabolism in the small intestine to yield two major metabolites, namely hexahydro‐curcumin and octahydro‐curcumin. Curcumin, hexahydro‐curcumin and octahydro‐curcumin were all subject to phase II metabolism that converted them to their corresponding glucuronides and sulfates. The majority of curcumin, hexahydro‐curcumin and octahydro‐curcumin exist as phase II conjugates in the small intestine. However, due to the action of gut microbiota, curcumin, hexahydro‐curcumin and octahydro‐curcumin almost exclusively existed in the non‐conjugated forms in the cecum and colon. Furthermore, our results suggested that gut microbiota could break down curcumin, hexahydro‐curcumin and octahydro‐curcumin to produce different fission products. In summary, our results determined the metabolic fate of curcumin in the GI tract, which can help understand the biological effects of curcumin in the GI tract as well as other related organs.Support or Funding InformationThis study was partially supported by fund from USDA.
Creating a Riboswitch‐Based Whole‐Cell Biosensor for Bisphenol A
Authors: Zorawski Marek , Shaffer Jeremy , Velasquez Erick , Liu Jane M. 
Published: June 2021

Bisphenol A (BPA) is a known endocrine disruptor and potential carcinogen found in polycarbonate plastic products, food packaging, and drinking water supplies. A quick and affordable detector, such as a whole‐cell biosensor that measures the presence of BPA, would greatly benefit safety and health. We set out to develop such a method by using riboswitch‐based biosensors. Riboswitches are regions of mRNAs that control downstream gene expression and can be used as genetic sensors to create whole‐cell biosensors. Although there are no naturally occurring BPA‐riboswitches, we reasoned that we could use the power of genetic selections to generate one. Starting from a known riboswitch that regulates expression of the downstream selection marker tetA linked to the reporter gene gfp, we used PCR or Gibson Assembly to replace the aptamer domain of the switch with 40 random bases, generating plasmid libraries. We then transformed Escherichia coli with these plasmids such that each individual bacterium harbored a unique member of the library. Libraries made by Gibson Assembly resulted in 108 unique members, while those made by PCR were generally on the order of 106 members. With libraries in hand, the dualistic nature of TetA allowed us to apply dual genetic selection to identify riboswitches that express tetA‐gfp only in response to BPA. TetA is a transporter protein that pumps the antibiotic tetracycline out of the cell, rendering cells tetracycline resistant. TetA also allows toxic metals, such as Ni2+, to enter the cell, inhibiting growth. The E. coli library was grown in the presence of BPA and tetracycline. Surviving cells were switched to medium with Ni2+ and no BPA. Only those cells that express tetA‐gfp exclusively in the presence of BPA were expected to survive both positive and negative selection. We succeeded in isolating unique riboswitches that converged to single sequences from starting libraries by bringing each library through three rounds of selection with varying tetracycline and Ni2+ conditions. Ultimately, our hits proved largely unsuccessful in producing a significant increase in fluorescence when incubated with BPA for either 6 or 16 hours compared to no‐ligand controls. In order to improve the ON/OFF ratio of our riboswitches, we integrated fluorescence‐activated cell sorting into our riboswitch‐development platform, allowing us to both select for survival through dual genetic selection and screen for maximum or minimum fluorescence depending on presence or absence of the ligand of interest. Further studies will include optimizing the obtained riboswitches by mutagenesis and directed evolution.Support or Funding InformationResearch funding is provided by the NSF.
Variation of fasting plasma inflammatory markers in overweight and obese individuals
Authors: Beals Elizabeth G , Demmer Elieke , Rivera Nancy , Rogers Tara S , Gertz Erik , Van Loan Marta , German J Bruce , Smilowitz Jennifer T , Zivkovic Angela M 
Published: June 2021

MethodsThirty‐six obese individuals were asked to fast for 12 hours prior to a baseline blood draw, on four separate visits separated by one to two weeks. Inflammatory markers, including serum amyloid A (SAA) and C‐reactive protein (CRP), cytokines, and adhesion markers were measured using multiplex assays. Data was analyzed in Microsoft Excel and JMP Pro 12. Multivariate analysis was used to determine associations between markers.ResultsThe results showed both positive and negative associations in the coefficient of variation (CV) of certain biomarkers in each subject. Some biomarkers, such as soluble Intracellular Adhesion Molecule (sICAM) and soluble Vascular Adhesion Molecule (sVCAM) had relatively low average CVs for all subjects (8.9% and 7.58%, respectively), whereas other biomarkers had much higher average CVs (Interleukin‐4, 77.68%). Additionally, multivariate plots of CV values for several biomarkers indicated positive associations, suggesting that within‐subject variability for these biomarkers may track similarly.ConclusionThese results suggest that some inflammatory markers may be more appropriate as biomarkers to measure inflammation than others. The baseline blood draw was completed in the morning and in a fasting state, without an inflammatory stimulus. Therefore, biomarkers with high CV levels may fluctuate for reasons that are not well‐understood. The amount of fluctuation within each subject offers insight to better understand which biomarkers may be useful clinically. Changes in inflammatory markers are important for determining a patient's inflammatory status, which is useful for the development of personalized nutrition and medicine, and disease prevention.Support or Funding Information1This project was made possible by support from the National Dairy Council, Rosemont, IL. and the USDA, Agricultural Research Service, Western Human Nutrition Research Center 2This study is registered on ClinicalTrials.gov Identifier: NCT01811329
The anti‐inflammatory effects of Spirulina platensis extract are mediated, in part, through the induction of an endotoxin tolerance‐like mechanism
Authors: Pham Tho X , Lee Ji‐Young 
Published: June 2021

Endotoxin tolerance is a phenomenon where exposure of innate immune cells to lipopolysaccharide (LPS) induces a refractory state to subsequent endotoxin exposures, which is characterized by a lack of pro‐inflammatory cytokine and chemokine production. We previously showed that pretreatment of macrophages (MΦ) with Spirulina platensis organic extract (SPE) repressed LPS‐induced inflammatory gene expression and cytokine secretion through the inhibition of nuclear factor κB p65 nuclear translocation. We also demonstrated that splenocytes and resident peritoneal MΦ isolated from C57BL/6J mice fed a high fat/high sucrose (HF/HS) diet containing 0.25% (w/w) SPE for 16 weeks displayed significantly attenuated expression of pro‐inflammatory genes when they were stimulated ex vivo by LPS compared to those from mice on a HF/HS control diet. The objective of this study was to investigate if SPE ameliorates LPS‐induced inflammatory gene expression in MΦ via an endotoxin tolerance‐like mechanism. Compared to naïve MΦ exposed to LPS for the first time, endotoxin tolerant (ET) RAW 264.7 MΦ, bone marrow‐derived MΦ, and THP‐1 MΦ showed significantly less interleukin (IL)‐1β expression after stimulation with 100 ng/mL of LPS, which was also observed with SPE pretreatment. The expression of IL‐6, tumor necrosis factor α, and monocyte chemoattractant protein‐1 was also repressed in ET as well as SPE‐pretreated RAW 264.7 MΦ. Consistent with the repressed expression of pro‐inflammatory genes, the translocation of p65 into the nucleus was markedly reduced in both ET and SPE‐pretreated MΦ upon LPS stimulation. It has been demonstrated that nuclear p50 homodimer and B‐cell lymphoma 3‐encoded protein (BCL3) are increased in monocytes and MΦ with endotoxin tolerance, consequently attenuating p65 transcriptional activity. We found that both SPE‐pretreated and ET RAW 264.7 MΦ had increased nuclear protein levels of p50 and BCL3. Quantification of nuclear p50 and p65 proteins showed that in ET and SPE‐pretreated cells, p50/p65 ratios were increased by more than 3‐fold compared to naïve MΦ when they were stimulated with LPS. In addition, ET MΦ showed significantly increased expression of BCL3, p105, inhibitor of κBα and IL‐ receptor‐associated kinase M, all of which are shown to be increased in endotoxin tolerance. In contrast to the LPS‐induced endotoxin tolerance, SPE significantly increased only BCL3 and p105 expression. At the protein level, endotoxin tolerization increased nuclear p50 proteins, but not BCL3, while SPE increased the nuclear abundance of both. In conclusion, the anti‐inflammatory effect of SPE is at least partly attributable to the induction of an endotoxin tolerance‐like state in macrophages, which shares common characteristics of MΦ endotoxin tolerance, while features distinctive from endotoxin tolerance also exist with SPE treatment.Support or Funding InformationUSDA AFRI (2014‐01870)
Vasopressin Receptor Regulation in Maintaining Potassium Homeostasis in a Sus scrofa Model of Hemorrhagic Shock
Authors: Uyehara Catherine F T , Kajiura Lauren N , Murata Lee‐Ann M , Wong Sara A , Ichimura Wayne M , Hernandez Claudia A , Sarkar Joy , Rowland Michael R 
Published: June 2021

We previously showed that elevated potassium (K)levels seen with severe hemorrhage appear to stimulate enhanced vasopressin(VP) release, and that in turn, elevated VP levels help correct hyperkalemia by increasing renal tubular K secretion. We thus tested whether brain VP receptors controlling vasopressin release were regulated synergistically with kidney VP receptors whichmodulated renal K excretion. We hypothesized that brain VP receptor up‐regulation may enable positive feedback for K‐stimulated augmentation of VP responsiveness in hemorrhage, and that renal VP receptor expression would reflect VP receptor mediated action on renal K excretion. Thus, in anesthetized Yorkshire cross pigs (n=35) we compared plasma VP (pVP) and plasmaK (pK) before, at the end of hemorrhage (30 ml/kg over 20 minutes), and after resuscitation, against mRNA expression of V1aR, V1bR, and V2R receptors in brain and kidney tissues collected after resuscitation. Multiple regression analysis showed that increased pVP at the end of hemorrhage positively correlated (p<0.05) with expression of V1aR and V1bR in the hypothalamus, V1bR in the anterior pituitary, and V2R in the renal cortex. Plasma K levels at the end of resuscitation were negatively correlated (p<0.05) with renalcortex V2R expression. In a different group of control animals that did not undergo hemorrhage, pVP and pK remained unchanged throughout. Results suggest that an up‐regulation of V1a and V1b receptors in the brain contributed to an amplified VP response in hemorrhage. In addition, VP appears to stimulate renal V2R expression to lower pK levels and maintain K homeostasis via a renal V2R‐mediated negative feedback mechanism.Support or Funding InformationThis project was supported by U.S. Army Medical Resarch and Materiel Command Congressionally Directed Medical Research Program grant #DAMD17‐03‐1‐0072. The views expressed in this abstract are those of the authors and do not reflect the official policy or position of the Department of the Army, Department of Defense, or the U.S. Government.