Exhaustive mutagenesis of six secondary active-site residues in Escherichia coli chorismate mutase shows the importance of hydrophobic side chains and a helix N-capping position for stability and catalysis

Secondary active-site residues in enzymes, including hydrophobic amino acids, may contribute to catalysis through critical interactions that position the reacting molecule, organize hydrogen-bonding residues, and define the electrostatic environment of the active site. To ascertain the tolerance of an important model enzyme to mutation of active-site residues that do not directly hydrogen bond with the reacting molecule, all 19 possible amino acid substitutions were investigated in six positions of the engineered chorismate mutase domain of the Escherichia coli chorismate mutase-prephenate dehydratase. The six secondary active-site residues were selected to clarify results of a previous test of computational enzyme design procedures. Five of the positions encode hydrophobic side chains in the wild-type enzyme, and one forms a helix N-capping interaction as well as a salt bridge with a catalytically essential residue. Each mutant was evaluated for its ability to complement an auxotrophic chorismate mutase deletion strain. Kinetic parameters and thermal stabilities were measured for variants with in vivo activity. Altogether, we find that the enzyme tolerated 34% of the 114 possible substitutions, with a few mutations leading to increases in the catalytic efficiency of the enzyme. The results show the importance of secondary amino acid residues in determining enzymatic activity, and they point to strengths and weaknesses in current computational enzyme design procedures

Computationally designed variants of Escherichia coli chorismate mutase show altered catalytic activity

Computational protein design methods were used to predict five variants of monofunctional Escherichia coli chorismate mutase expected to maintain catalytic activity. The variants were tested experimentally and three active site mutants exhibited catalytic activity similar to or greater than the wild-type enzyme. One mutant, Ala32Ser, showed increased catalytic efficiency

Impacts of Barrier-Island Breaching on Mainland Flooding During Storm Events applied to Moriches, NY

Barrier islands can protect the mainland from flooding during storms through reduction of storm surge and dissipation of storm generated wave energy. However, the protective capability is reduced when barrier islands breach and a direct hydrodynamic connection between the water bodies on both sides of the barrier island is established. Breaching of barrier islands during large storm events is complicated, involving nonlinear processes that connect water, sediment transport, dune height, and island width among other factors. In order to assess how barrier island breaching impacts flooding on the mainland, we used a statistical approach to analyze the sensitivity of mainland storm-surge to barrier island breaching by randomizing the location, time, and extent of a breach event. We created a framework that allows breaching to develop during the course of a simulation and imposes a breach in an approximation of a Gaussian bell-curve that deepens over time. We show that simulating a storm event and varying the size, location, and number of breaches in the barrier island that mainland storm surge and horizontal inundation is affected by breaching; total inundation has a logarithmic relationship with total breach area which tapers off after the entire island is removed. Breach location is also an important predictor of inundation and bay surge. The insights we've gleaned from this study can help prepare shoreline communities for the differing ways that breaching affects the mainland coastline. Understanding which mainland locations are vulnerable to breaching, planners and coastal engineers can design interventions to reduce the likelihood of a breach occurring in areas adjacent to high flood risk

Evidence for Prosody in Silent Reading

English speakers and expressive readers emphasize new content in an ongoing discourse. Do silent readers emphasize new content in their inner voice? Because the inner voice cannot be directly observed, we borrowed the cap-emphasis technique (e.g., “toMAYto”) from the pronunciation guides of dictionaries to elicit prosodic emphasis. Extrapolating from linguistic theories of focus prosody in spoken English, we predicted and found that silent readers in experiment 1 preferred cap-emphasized, newsworthy content (“James stole the BRACELET ) when the just-read story left them wondering what was stolen (compared with control trials). Readers preferred “JAMES stole the bracelet” when left wondering who the thief was. Experiment 2 generalized our findings to newsworthy function words and to a new behavioral measure, reaction time. As predicted, “He CAN” was judged more quickly and accurately following “Can he swim,” whereas “HE can” was judged more quickly and accurately following “Who can swim?” Our results suggest that readers engage focus prosody when they read silently

The Sport Concussion Assessment Tool-5 (SCAT5): Baseline Assessments in NCAA Division I Collegiate Student-Athletes

International Journal of Exercise Science 13(3): 1143-1155, 2020. The purpose of this study was to report baseline values for the SCAT5 in NCAA Division I collegiate student-athletes, while also evaluating if sex, health diagnoses, or sport type influenced baseline performance. A sample of 462 collegiate student-athletes (212 females, 250 males, (19.21±1.32 years)) completed the SCAT5 prior to the 2017-18, 2018-19 or 2019-20 athletic seasons. Descriptive statistics were reported for symptom total (22 possible), symptom severity (132 possible), orientation (5 possible), immediate memory (30 possible), concentration (5 possible), delayed recall (10 possible), total SAC score (50 possible), 3 mBESS stances (10 possible), and mBESS score (30 possible). Separate Mann-Whitney U tests were conducted to identify sex, health diagnoses (concussion history, ADD/ADHD, depression/anxiety), and sport type (contact, non-contact) differences for all SCAT5 components. Alpha level was set a priori \u3c.05. Student-athletes reported 1.96± 3.37 symptoms with a severity of 3.43±7.63, and an overall SAC score of 35.14±5.23 (orientation 4.96±0.20, immediate memory 20.18±3.40, concentration 3.60±1.14, delayed recall 6.41±1.94). Student-athletes participating in contact sports, had ADD/ADHD, or depression/anxiety reported more symptoms and at greater severity (p=\u3c.001-.01). Those with ADD/ADHD performed worse on mBESS (p=.01-.03). No sex differences were found for any SCAT5 components (p=.08-.90). This study presents reference values for the SCAT5 by sex, health diagnoses, and sport type. Healthcare professionals may utilize these normative values when individual baseline references are unavailable

Temporal and Spatial Impact of Human Cadaver Decomposition on Soil Bacterial and Arthropod Community Structure and Function

As vertebrate carrion decomposes, there is a release of nutrient-rich fluids into theunderlying soil, which can impact associated biological community structure andfunction. How these changes alter soil biogeochemical cycles is relatively unknown and may prove useful in the identification of carrion decomposition islands that have long lasting, focal ecological effects. This study investigated the spatial (0, 1, and 5 m) and temporal (3–732 days) dynamics of human cadaver decomposition on soil bacterial and arthropod community structure and microbial function. We observed strong evidence of a predictable response to cadaver decomposition that varies over space for soil bacterial and arthropod community structure, carbon (C) mineralization and microbial substrate utilization patterns. In the presence of a cadaver (i.e., 0 m samples), the relative abundance of Bacteroidetes and Firmicutes was greater, while the relative abundance of Acidobacteria, Chloroflexi, Gemmatimonadetes, and Verrucomicrobia was lower when compared to samples at 1 and 5 m. Micro-arthropods were more abundant (15 to 17-fold) in soils collected at 0 m compared to either 1 or 5 m, but overall, micro-arthropod community composition was unrelated to either bacterial community composition or function. Bacterial community structure and microbial function also exhibited temporal relationships, whereas arthropod community structure did not. Cumulative precipitation was more effective in predicting temporal variations in bacterial abundance and microbial activity than accumulated degree days. In the presence of the cadaver (i.e., 0 m samples), the relative abundance of Actinobacteria increased significantly with cumulative precipitation. Furthermore, soil bacterial communities and C mineralization were sensitive to the introduction of human cadavers as they diverged from baseline levels and did not recover completely in approximately 2 years. These data are valuable for understanding ecosystem function surrounding carrion decomposition islands and can be applicable to environmental bio-monitoring and forensic sciences

Standardizing Umbilical Cord Mesenchymal Stromal Cells for Translation to Clinical Use: Selection of GMP-Compliant Medium and a Simplified Isolation Method

Citation: Smith, J. R., Pfeifer, K., Petry, F., Powell, N., Delzeit, J., & Weiss, M. L. (2016). Standardizing Umbilical Cord Mesenchymal Stromal Cells for Translation to Clinical Use: Selection of GMP-Compliant Medium and a Simplified Isolation Method. Stem Cells International, 14. doi:10.1155/2016/6810980Umbilical cord derived mesenchymal stromal cells (UC-MSCs) are a focus for clinical translation but standardized methods for isolation and expansion are lacking. Previously we published isolation and expansion methods for UC-MSCs which presented challenges when considering good manufacturing practices (GMP) for clinical translation. Here, a new and more standardized method for isolation and expansion of UC-MSCs is described. The new method eliminates dissection of blood vessels and uses a closed-vessel dissociation following enzymatic digestion which reduces contamination risk and manipulation time. The new method produced >10 times more cells per cm of UC than our previous method. When biographical variables were compared, more UC-MSCs per gram were isolated after vaginal birth compared to Caesarian-section births, an unexpected result. UC-MSCs were expanded in medium enriched with 2%, 5%, or 10% pooled human platelet lysate (HPL) eliminating the xenogeneic serum components. When the HPL concentrations were compared, media supplemented with 10% HPL had the highest growth rate, smallest cells, and the most viable cells at passage. UC-MSCs grown in 10% HPL had surface marker expression typical of MSCs, high colony forming efficiency, and could undergo trilineage differentiation. The new protocol standardizes manufacturing of UC-MSCs and enables clinical translation

UNLV College of Education Multicultural & Diversity Newsletter

A newsletter discussing a variety of topics dealing with multicultural topics and diversity in education

Always on my mind: Cross-brain associations of mental health symptoms during simultaneous parent-child scanning.

How parents manifest symptoms of anxiety or depression may affect how children learn to modulate their own distress, thereby influencing the children's risk for developing an anxiety or mood disorder. Conversely, children's mental health symptoms may impact parents' experiences of negative emotions. Therefore, mental health symptoms can have bidirectional effects in parent-child relationships, particularly during moments of distress or frustration (e.g., when a parent or child makes a costly mistake). The present study used simultaneous functional magnetic resonance imaging (fMRI) of parent-adolescent dyads to examine how brain activity when responding to each other's costly errors (i.e., dyadic error processing) may be associated with symptoms of anxiety and depression. While undergoing simultaneous fMRI scans, healthy dyads completed a task involving feigned errors that indicated their family member made a costly mistake. Inter-brain, random-effects multivariate modeling revealed that parents who exhibited decreased medial prefrontal cortex and posterior cingulate cortex activation when viewing their child's costly error response had children with more symptoms of depression and anxiety. Adolescents with increased anterior insula activation when viewing a costly error made by their parent had more anxious parents. These results reveal cross-brain associations between mental health symptomatology and brain activity during parent-child dyadic error processing