Investigating the Structure of the Papain-Inhibitor Complex using SPR and NMR

Cysteine proteases (CPs) are enzymes with a nucleophilic thiol in their active sites. Inhibitors of cysteine proteases (ICPs) occur naturally in bacterial pathogens and some protozoa. In parasites, ICPs are often virulence factors, contributing to the formation and survival of amastigotes within host cells. These amastigotes have higher CP activity, therefore making both ICPs and CPs potential drug targets. Despite great genetic variability, ICPs contain highly conserved structural features, including a series of defined loops that play a significant role in binding CPs. Papain, a CP from Carica papaya, complexes with ICP from Leishmania mexicana. Although the individual 3-D structures of ICP and papain have been determined, as of this work, the structure of the papain-ICP complex has only been predicted, not solved. This research details the development of a technique for determining quaternary structure of the papain-ICP complex using paramagnetic relaxation enhancement NMR (PRE-NMR). A paramagnetic tag (MTSL) was added to various cysteine-mutants of ICP to measure distances to reductively 13C-methylated papain. The modification of ICP with MTSL was quantified using EPR, and the effects of labeling on the binding kinetics of papain and ICP were determined using SPR. 13C-methyl peak perturbations due to PRE were observed when papain was bound to spin-labeled E102C-ICP and K27C-ICP. Intermolecular distances were predicted using modeling software and a working model of the complex was created. Data from additional mutants will help to further determine complex structure and perfect the model.The penultimate chapter of this dissertation includes work towards the development of a method for studying protein-protein interactions using atomic force microscopy. Papain-ICP was used as a model system, with the intention to apply this method to the study of another system: filamentous actin (f-actin) and the actin-binding domain of abelson tyrosine-protein kinase (ABL2-FABD). The creation of nanopores on an AFM sensor chip surface was successful. ICP monomers bound selectively into the pores. Attempts to form the papain-ICP complex on the chip surface were unsuccessful, and future work is needed to perfect this method. The final chapter of this dissertation is a literature review outlining previous work in this area

Impact of mask wearing during high-intensity exercise on post-exercise hemodynamics

Background: Few studies examining face mask wearing during high-intensity interval exercise (HIE) have measured blood pressure (BP) and cardiac output (Q) during exercise and none have examined these variables post-exercise. Methods: Participants were randomly assigned to complete four exercise and two control conditions while wearing different face masks. Participants followed a 4x4 protocol on a cycle ergometer. Participants exercised at 85% of VO2max for 4-min, followed by a 3-min rest, repeated four times. Measurements of Q, systemic vascular resistance (SVR), and BP were measured pre-exercise for 20-min, during exercise, and postexercise for 60-min. Linear mixed models were used to detect differences between conditions. Results: Ten young (20.3 ± 1.4 yr.) male (n = 5) and female (n = 5) participants with an average BMI of 28.1 ± 7.3 kg/m2 and VO2max of 37.0 ± 7.1 ml.kg-1.min-1 completed this. There were no group differences during exercise on outcomes of Q, SVR, HR, SBP, DBP, MAP, or RPE (all p > .05). During exercise, EXS-N95 had a lower SV than CON-E (p = .014) and EXS-CL (p = .006). All mask conditions had a higher post-exercise HR than CON-E (all p > .05). Only EXS-SUR differed in post-exercise brachial SBP compared to CON-E (3.1 ± 1.6 mmHg, p < .043). Of the exercise conditions, only EXS-N95 differed from CON-E with an increase of 2.0 ± .88 mmHg for brachial DBP (p = .022) and 2.1 ± .92 mmHg for central DBP (p = .022), SV (-11.8 ± 3.5 mL.min-1, p < .001), Q (-.52 ± .26 L.min-1, p = .045), and SVR (73.7 ± 29.8 Dyn.s/cm5, p = .014). Conclusion: The current study shows that in healthy populations, wearing a face covering of any type during HIE does not impactfully change the hemodynamic response during exercise or recovery period

How does maximal aerobic capacity predict the performance of ROTC cadets on the ACFT

The Army Combat Fitness Test (ACFT) is a newly developed test that assesses the combat readiness of U.S. Army soldiers. Maximal aerobic capacity (VO2max) characteristics vary between soldiers and have the ability to affect performance outcomes. The purpose of this cross-sectional study is to determine if VO2max can predict performance outcomes of the ACFT in ROTC cadets. Methods: ROTC cadets (44 males, 15 females; aged 21.06 ± 3.6 years) completed the 6-event ACFT (3-repetition maximum trap-bar deadlift [MDL], standing power toss [SPT], hand-release pushups [HRPU], sprint-drag-carry shuttle run [SDC], plank [PLK], and 2-mile run [2MR]). The cadets were invited into the laboratory to conduct a maximal treadmill running test following the Bruce protocol. The ability for VO2max (mL·kg-1·min-1) to predict ACFT performance was determined with a linear regression model. Significance was set at p \u3c 0.05. Results: VO2max was significantly and positively correlated to MDL (r = .301, p = .018), HRP (r = .525, p \u3c .001), SDC (r = .573, p \u3c .001), PLK (r = .668, p \u3c .001) 2MR (r = .642, p \u3c .001) and overall ACFT score (r = .666, p \u3c .001) except MDL (r = -.021, p = .920), SPT (r = -.173, p = .408), HRP (r = .280, p = .175). VO2max significantly explained 43% (p = .001) of the variance on the total ACFT scores with a beta coefficient of 4.911. Conclusion: There is a gap in the understanding of how VO2max impacts performance in the newly implemented ACFT. VO2max is a predictor of the ACFT total and significantly correlates with the MDL, HRP, SDC, PLK, and 2MR. VO2max did not correlate with the individual event SPT. For every .715 mL·kg-1·min-1 increase in VO2max, ACFT total scores increased by 4.911 points. These findings create the need for further research due to the majority of U.S. Army personnel failing the PLK and 2MR, which can be associated with an insignificant aerobic capacity

Fat-Fit Index on Predicting Cardiovascular Outcomes in College Aged Students

Globally, 70% of all deaths annually occur from chronic diseases due to lifestyle risk factors such as obesity and low physical activity. These modifiable factors contribute to the result of chronic cardiovascular disease, respiratory disease, and diabetes. Body Mass Index (BMI) is a common measurement used to predict risk obesity. Furthermore, maximum oxygen consumption (VO2) is used to measure an individual\u27s cardiorespiratory fitness. Fitness Fat Index (FFI) has been suggested to measure chronic disease risks. FFI represents an individual’s cardiorespiratory fitness (CRF) divided by their waist to height ratio (WHR). Higher FFI is associated with cardiovascular disease and the reduction of all-cause mortality. FFI has been suggested as superior to BMI and VO2 in predicting health outcomes. The ability of FFI to predict measures of cardiovascular outcomes in college aged students is unknown. PURPOSE: The purpose of this study was to examine if FFI is superior to BMI or VO2 in predicting cardiovascular outcomes in college aged students. METHODS: 217 total college-aged subjects (20.9 ± 3.5) with an average BMI of 25.2 ±4.9 completed the study. BMI, FFI measurements were performed and VO2 max test was conducted. Pulse Wave Velocity (PWV), augmentation pressure (AP), augmentation index (AIX), brachial systolic blood pressure (BSBP), brachial diastolic blood Pressure (BDBP), central systolic blood pressure (CSBP), and central diastolic blood pressure (DBP) were measured. Hierarchical regression analysis was used with age entered in the first block and the predictor variables of BMI, VO2, and FFI were individually entered in the second block. RESULTS: After adjusting for age, BMI explained more of the variance on outcomes of AP (R2 change 5.9%,

Body Adiposity Index Superior to Body Mass Index in Predicting Adiposity in Army ROTC Cadets

The Army’s standards to assess body composition were established in the 1960s. The current body composition assessment is due for change as there are newer alternate methods to assess body composition using adiposity. Body Mass Index (BMI) is moderately correlated with % body fat and often used to assess obesity in the general population. The Body Adiposity Index (BAI) is an alternative anthropometric measurement suggested to be superior to BMI at predicting adiposity, but has not been well assessed within military populations. PURPOSE: The purpose of this correlational study was to determine the accuracy of the BMI overweight category as a measure of fatness in ROTC cadets, and to determine if BAI is a superior predictor of % body fat to BMI. METHODS: ROTC cadets were invited into the laboratory for measurements of height, weight, and body fat via BOD POD. Bodyfat above 33% was considered overfat and BMI \u3e25 kg/m2 was considered overweight. BAI was calculated as (Hip Circumference/Height1.5)–18. Pearson correlation determined the strength of relationships. RESULTS: 83 collegiate-aged (20.91 ± 3.37 yr.) ROTC cadets with an average BMI of 24.85 ± 4.87 kg/m2 and % body fat of 23.69 ± 5.95 participated in the study. BMI and BAI significantly predicted % body fat (r = .508, p \u3c 0.001, r = .482, p \u3c 0.001, respectively). When using BMI to determine overfat, 4% (n = 2) of ROTC cadets had a false positive (overweight, normal fat) and 10% (n = 5) had a false negative (normal weight, overfat). BMI predicts 25.8% of body fat and BAI predicts 23.2% of body fat. Because BMI and BAI are statistically similar, both are good predictors of % body fat. Conclusion: Due to the army’s outdated recent measures of body composition, alternative methods could improve quality and consistency of assessments in the future with the use of BMI and BAI

How does body composition predict the performance of ROTC cadets on the ACFT

The Army Combat Fitness Test (ACFT) is a newly developed test assessing U.S. Army soldiers\u27 combat readiness. Body composition characteristics vary between soldiers and could affect performance outcomes. The purpose of this cross-sectional study is to determine whether fat-mass and fat-free-mass can predict performance outcomes of the ACFT in ROTC cadets. Methods: ROTC cadets (31 males, 21 females; aged 20.5 ± 2.2 years) completed the 6-event ACFT (3-repetition maximum trap-bar deadlift [MDL], standing power toss [SPT], hand-release pushups [HRPU], sprint-drag-carry shuttle run [SDC], plank [PLK], and 2-mile run [2MR]). The cadets were invited into the laboratory for measurements of anthropometrics (height, weight, and body mass index [BMI]) and body composition analysis via the air displacement plethysmograph (body fat mass [BFM], fat-free mass [FFM]). The ability of body composition to predict ACFT performance was determined with a linear regression model. Significance was set at p \u3c 0.05. Results: BFM was significantly and negatively correlated to SDC (r = -.383, p = .005), PLK (r = -.567, p \u3c .001), 2MR (r = -.577, p \u3c .001), HRP (r = -.501, p \u3c .001) and overall ACFT score (r = -.574, p = .001) except 3DL (r = .199, p = .154) and SPT (r = -.193, p = .166). FFM was significantly correlated to SDC (r = .411, p = .001), PLK (r = .249, p = .047), 3DL (r = .266, p = .034), SPT (r = .458, p = .001), and overall ACFT score (r = .364, p = .003) except 2MR (r = .137, p \u3c .279) HRP (r = .126, p = .322). BFM significantly explained 33% (p = .001) of the variance on the total ACFT scores with a beta coefficient -4.632. Conclusion: Body composition measurements of BFM and FFM are predictors of the ACFT total score. These data show that both BFM and FFM are important metrics for assessing a soldier’s combat readiness. For every 1% increase in body fat, ACFT scores decreased by 4 points. For every 1kg increase in FFM, ACFT score increased by 1.5 points

Towards the identification of the soil microbiome community associated with Longleaf Pine

This project is part of a larger study looking at the restoration of the Long-leaf Pine ecosystem in certain Wildlife Management Areas (WMA) in Northwest Georgia. As part of this larger study another group of researchers has been looking at changes in the plant community in the same plots used for this soil microbiome research. Our long-term aim is to look for potential associations between the above and below-ground community structures. Although research has shown that the health and composition of the microbiome surrounding the roots of plants has a significant impact on the ability of plants to fight and survive various stressors (Wei et al. 2019), the full extent of the complex system of feedback mechanisms between the rhizosphere and soil microbiome, and the above ground plant communities is not yet understood. We collected soil samples from six plots located in the Sheffield WMA located in Paulding County, GA. Except for two plots at the savanna site, with slopes south or west facing; soil samples were collected from either north or south facing hillsides. Within each plot, samples were collected in sterile plastic tubes at the center of the plot and 10 m above and below from the center. Five random sub samples were taken from each tube for DNA extraction using a commercially available kit. Given that plant communities appear to differ between north and south facing slopes, we hypothesize that the soil microbiome will also differ significantly in north versus south facing areas. We also hypothesize that the microbiome in the savanna plots, which are actively being restored for Longleaf pine, will be distinct from all others

How does diet quality predict the performance of ROTC cadets on the ACFT

The Army Combat Fitness Test (ACFT) is a newly developed test assessing U.S. Army soldiers\u27 combat readiness. The Healthy Eating Index 2015 (HEI-2015) is a questionnaire established by the USDA that quantifies overall diet quality. The HEI-2015 uses a scale of 0 to 100 with higher scores aligning more with the recommendations from the Dietary Guidelines for Americans. The HEI-2015 is comprised of 13 subcategories: Total Fruits [TF], Whole Fruits [WF], Total Vegetables [TV], Greens and Beans [GB], Whole Grains [WG], Dairy [D], Total Protein foods [TP], Seafood and Plant Proteins [SPP], Fatty Acids [FA], Refined Grains [RG], Sodium [S], Added Sugars [AS], and Saturated Fats [SF]. Purpose: The purpose of this cross-sectional study is to determine if diet quality can predict performance outcomes of the ACFT in ROTC cadets. Methods: ROTC cadets (31 males, 21 females; aged 20.5 ± 2.2 years) completed the 6-event ACFT (3-repetition maximum trap-bar deadlift [MDL], standing power toss [SPT], hand-release pushups [HRPU], sprint-drag-carry [SDC], plank [PLK], and 2-mile run [2MR]). The cadets were invited into the laboratory to complete the HEI-2015. The ability of the HEI-2015 to predict ACFT performance was determined with a linear regression model. Significance was set at p \u3c 0.05. Results: HEI-2015 total score was significantly and positively correlated to ACFT total score, showing (r = .319, p = .014). Subcategories of the HEI-2015 were positively and significantly correlated with ACFT total scores, GB (r = .326, p = .012), SPP (r = .349, p = .007), and FA (r = .263, p = .044). HEI-2015 total score significantly explained 10% (p = .014) of the variance on the ACFT total scores with a beta coefficient 3.121. Conclusion: HEI-2015 total score, GB, SPP, and FA are all predictors of the ACFT total score. This data shows that overall diet quality is an important factor in determining a soldier’s combat readiness. For every 1.2 point increase in HEI-2015 total score, it is predicted that ACFT total score will increase by 3.1 points

The Role of Body Fat Percentage and VO2MAX in Predicting Arterial Stiffness in Female College Students

Within the college population, high blood pressure (BP), as well as obesity and sedentary lifestyles, are seen. Bodyfat influences these outcomes. Much of the literature ignores the possible effects that fitness may have on these outcomes, specifically in females. Purpose: The intent of this study was to determine if VOMAX can predict arterial stiffness after adjusting for body fat% in college females. Methods: Healthy young females were recruited from Grand Canyon University. Subjects came to the lab for a single visit for the following assessments: height, weight, waist circumference, hip circumference, aortic BP, augmentation pressure (AP), augmentation index adjusted at a heart rate of 75 (Aix@75), carotid-femoral pulse wave velocity (cfPWV) and a VO2MAX. Hierarchical regression was used with body fat% entered into the first block and VO2MAX entered into the second block. Results: 92 female college students completed the study. The mean BMI was 25.4 +/-5.1 kg/m2, and the mean age was 20.5 +/- 2.2 years. Body fat% and VO2MAX did not statistically predict brachial systolic BP (P=0.137) or cfPWV (P=0.439). Body fat % significantly predicted brachial diastolic BP (R2=0.047, P=0.040), and VO2MAX significantly explains 4.4% more of the variance (P=0.043). Body fat % significantly predicted central systolic BP (R2=0.126, P=2MAX did not significantly add to the model (P=0.320). For central diastolic BP, body fat % was not an independent predictor (P=0.075); however, VO2MAX was found to be an independent predictor (R2=0.100, P=0.015). Body fat % was not an independent predictor (P=0.075) of AP; however, VO2MAX was found to be an independent predictor of AP (R2=0.113, P=0.008). Body fat % significantly predicted Aix@75 (R2=0.067, P=0.014), and VO2MAX was an independent predictor (R2=0.193, P=Conclusion: After adjusting for body fat%, VO2MAX significantly predicted measures of arterial stiffness in college aged females. Additionally, VO2MAX seemed to be a better predictor of arterial stiffness than body fat%. It is crucial to also consider physical fitness when considering body fat% and health