Digital Commons Network^™

Metformin Alleviates Hepatic Iron Overload And Ferroptosis Through Ampk-Ferroportin Pathway In Hfd-Induced Nafld, Fangzhi Yue, Ying Shi, Shanyu Wu, Lin Xing, Dan He, Lin Wei, Anqi Qiu, Ryan D. Russell, Dongmei Zhang

Health & Human Performance Faculty Publications and Presentations

Highlights

• Metformin alleviates HIO and ferroptosis in HFD-induced NAFLD

• FPN is involved in the molecular mechanism of metformin on HIO in HFD-induced NAFLD

• Metformin upregulates FPN expression by reducing lysosomal ubiquitination degradation

Summary

Metformin prevents progression of non-alcoholic fatty liver disease (NAFLD). However, the potential mechanism is not entirely understood. Ferroptosis, a recently recognized nonapoptotic form of regulated cell death, has been reported to be involved in the pathogenesis of NAFLD. Here, we investigated the effects of metformin on ferroptosis and its potential mechanism in NAFLD. We found that metformin prevented the progression of NAFLD, and alleviated hepatic iron …

Metformin Alleviates Hepatic Iron Overload And Ferroptosis Through Ampk-Ferroportin Pathway In Hfd-Induced Nafld, Fangzhi Yue, Ying Shi, Shanyu Wu, Lin Xing, Dan He, Lin Wei, Anqi Qiu, Ryan D. Russell, Dongmei Zhang

Health & Human Performance Faculty Publications and Presentations

Highlights

• Metformin alleviates HIO and ferroptosis in HFD-induced NAFLD

• FPN is involved in the molecular mechanism of metformin on HIO in HFD-induced NAFLD

• Metformin upregulates FPN expression by reducing lysosomal ubiquitination degradation

Summary

Metformin prevents progression of non-alcoholic fatty liver disease (NAFLD). However, the potential mechanism is not entirely understood. Ferroptosis, a recently recognized nonapoptotic form of regulated cell death, has been reported to be involved in the pathogenesis of NAFLD. Here, we investigated the effects of metformin on ferroptosis and its potential mechanism in NAFLD. We found that metformin prevented the progression of NAFLD, and alleviated hepatic iron …

Hydrogen Peroxide-Dependent Oxidation Of Erk2 Within Its D-Recruitment Site Alters Its Substrate Selection, Anthony E. Postiglione, Laquaundra L. Adams, Ese S. Ekhator, Anuoluwapo E. Odelade, Supriya Patwardhan, Meenal Chaudhari, Avery S. Pardue, Anjali Kumari, William A. Lefever, Olivia P. Tornow, Tamer S. Kaoud, Johnathan Neiswinger, Jun Seop Jeong, Derek Parsonage, Kimberly J. Nelson, Dukka Kc, Cristina M. Furdui, Heng Zhu, Andrew J. Wommack, Kevin N. Dalby, Ming Dong, Leslie B. Poole, Jeremiah D. Keyes, Robert H. Newman

Michigan Tech Publications, Part 2

Extracellular signal-regulated kinases 1 and 2 (ERK1/2) are dysregulated in many pervasive diseases. Recently, we discovered that ERK1/2 is oxidized by signal-generated hydrogen peroxide in various cell types. Since the putative sites of oxidation lie within or near ERK1/2’s ligand-binding surfaces, we investigated how oxidation of ERK2 regulates interactions with the model substrates Sub-D and Sub-F. These studies revealed that ERK2 undergoes sulfenylation at C159 on its D-recruitment site surface and that this modification modulates ERK2 activity differentially between substrates. Integrated biochemical, computational, and mutational analyses suggest a plausible mechanism for peroxide-dependent changes in ERK2-substrate interactions. Interestingly, oxidation decreased ERK2’s …

Increased Sirt3 Combined With Parp Inhibition Rescues Motor Function Of Sbma Mice., David R Garcia Castro, Joseph R Mazuk, Erin M Heine, Daniel Simpson, R Seth Pinches, Caroline Lozzi, Kathryn Hoffman, Phillip Morrin, Dylan Mathis, Maria V Lebedev, Elyse Nissley, Kang Hoo Han, Tyler Farmer, Diane E Merry, Qiang Tong, Maria Pennuto, Heather L. Montie

PCOM Scholarly Papers

Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular disease with substantial mitochondrial and metabolic dysfunctions. SBMA is caused by polyglutamine (polyQ) expansion in the androgen receptor (AR). Activating or increasing the NAD^+-dependent deacetylase, SIRT3, reduced oxidative stress and death of cells modeling SBMA. However, increasing diminished SIRT3 in AR100Q mice failed to reduce acetylation of the SIRT3 target/antioxidant, SOD2, and had no effect on increased total acetylated peptides in quadriceps. Yet, overexpressing SIRT3 resulted in a trend of motor recovery, and corrected TCA cycle activity by decreasing acetylation of SIRT3 target proteins. We sought to boost blunted …

Social Transmission Of Predator Information Through Referential Alarm Calls Within And Across Species, Carly Trebac

Honors Theses

Animals can encode information about a predator’s size, identity, or threat through alarm calls. This can provide referential information that can be used by individuals within and across species receiving the information. Referential alarm calls can encode information through changes in structure or rate of the call, and these variances in their call can elicit different responses, such as fleeing or mobbing, in conspecifics and heterospecifics. We aimed to investigate whether white-breasted nuthatch alarm calls can encode referential information and elicit different responses from conspecifics. Previous studies showed that a related species of nuthatch varies their call rate in the …

Frankenbacteriosis Targeting Interactions Between Pathogen And Symbiont To Control Infection In The Tick Vector, Lorena Mazuecos, Pilar Alberdi, Angélica Hernández-Jarguín, Marinela Contreras, Margarita Villar, Alejandro Cabezas-Cruz, Ladislav Simo, Almudena González-García, Sandra Díaz-Sánchez, Girish Neelakanta, Sarah I. Bonnet, Erol Fikrig, José De La Fuente

Biological Sciences Faculty Publications

(Summary) Tick microbiota can be targeted for the control of tick-borne diseases such as human granulocytic anaplasmosis (HGA) caused by model pathogen, Anaplasma phagocytophilum. Frankenbacteriosis is inspired by Frankenstein and defined here as paratransgenesis of tick symbiotic/commensal bacteria to mimic and compete with tick-borne pathogens. Interactions between A. phagocytophilum and symbiotic Sphingomonas identified by metaproteomics analysis in Ixodes scapularis midgut showed competition between both bacteria. Consequently, Sphingomonas was selected for frankenbacteriosis for the control of A. phagocytophilum infection and transmission. The results showed that Franken Sphingomonas producing A. phagocytophilum major surface protein 4 (MSP4) mimic pathogen and reduce infection …

Pollutants Corrupt Resilience Pathways Of Aging In The Nematode C. Elegans, Andrea Scharf, Annette Limke, Karl Heinz Guehrs, Anna Von Mikecz

Biological Sciences Faculty Research & Creative Works

Delaying aging while prolonging health and lifespan is a major goal in aging research. One promising strategy is to focus on reducing negative interventions such as pollution and their accelerating effect on age-related degeneration and disease. Here, we used the short-lived model organism C. elegans to analyze whether two candidate pollutants corrupt general aging pathways. We show that the emergent pollutant silica nanoparticles (NPs) and the classic xenobiotic inorganic mercury reduce lifespan and cause a premature protein aggregation phenotype. Comparative mass spectrometry revealed that increased insolubility of proteins with important functions in proteostasis is a shared phenotype of intrinsic- and …

Investigating The Effect Of Bacterial Ev-Packaged Rna On Human Gene Expression, Emily Marino

Student Research and Creative Activity Expo

Extracellular vesicles (EVs) produced by S. aureus at 2 physiologically relevant temperatures affect human gene expression.

Genetic Diversity And Differentiation Across Geographically Separated Populations Of North American Small Terrestrial Mammals., Molly Haugen

Student Research and Creative Activity Expo

We assess the extent of genetic connectivity between geographically separated populations of three small terrestrial mammals.

Pparα And Pparγ Activation Is Associated With Pleural Mesothelioma Invasion But Therapeutic Inhibition Is Ineffective, M. Lizeth Orozco Morales, Catherine A. Rinaldi, Emma De Jong, Sally M. Lansley, Joel P. A. Gummer, Bence Olasz, Shabarinath Nambiar, Danika E. Hope, Thomas H. Casey, Y. C. Gary Lee, Connull Leslie, Gareth Nealon, David M. Shackleford, Andrew K. Powell, Marina Grimaldi, Patrick Balaguer, Rachael M. Zemek, Anthony Bosco, Matthew J. Piggott, Alice Vrielink, Richard A. Lake, W. Joost Lesterhuis

Research outputs 2022 to 2026

Mesothelioma is a cancer that typically originates in the pleura of the lungs. It rapidly invades the surrounding tissues, causing pain and shortness of breath. We compared cell lines injected either subcutaneously or intrapleurally and found that only the latter resulted in invasive and rapid growth. Pleural tumors displayed a transcriptional signature consistent with increased activity of nuclear receptors PPARα and PPARγ and with an increased abundance of endogenous PPAR-activating ligands. We found that chemical probe GW6471 is a potent, dual PPARα/γ antagonist with anti-invasive and anti-proliferative activity in vitro. However, administration of GW6471 at doses that provided sustained plasma …

The Mrna-Lnp Platform's Lipid Nanoparticle Component Used In Preclinical Vaccine Studies Is Highly Inflammatory, Sonia Ndeupen, Zhen Qin, Sonya Jacobsen, Aurélie Bouteau, Henri Estanbouli, Botond Z. Igyártó

Department of Microbiology and Immunology Faculty Papers

Vaccines based on mRNA-containing lipid nanoparticles (LNPs) are a promising new platform used by two leading vaccines against COVID-19. Clinical trials and ongoing vaccinations present with varying degrees of protection levels and side effects. However, the drivers of the reported side effects remain poorly defined. Here we present evidence that Acuitas' LNPs used in preclinical nucleoside-modified mRNA vaccine studies are highly inflammatory in mice. Intradermal and intramuscular injection of these LNPs led to rapid and robust inflammatory responses, characterized by massive neutrophil infiltration, activation of diverse inflammatory pathways, and production of various inflammatory cytokines and chemokines. The same dose of …

Stabilin Receptors Clear Lps And Control Systemic Inflammation, Fatima Cabral, Mustafa Al-Rahem, John Skaggs, Thushara A. Thomas, Naresh Kumar, Qian Wu, Paolo Fadda, Lianbo Yu, John M. Robinson, Jonghan Kim, Ekta Pandey, Xinghui Sun, Wael N. Jarjour, Murugesan V.S. Rajaram, Edward N. Harris, Latha P. Ganesan

Department of Biochemistry: Faculty Publications

Lipopolysaccharides (LPSs) cause lethal endotoxemia if not rapidly cleared from blood circulation. Liver sinusoidal endothelial cells (LSEC) systemically clear LPS by unknown mechanisms. We discovered that LPS clearance through LSEC involves endocytosis and lysosomal inactivation via Stabilin-1 and 2 (Stab1 and Stab2) but does not involve TLR4. Cytokine production was inversely related to clearance/endocytosis of LPS by LSEC. When exposed to LPS, Stabilin double knockout mice (Stab DK) and Stab1 KO, but not Stab2 KO, showed significantly enhanced systemic inflammatory cytokine production and early death compared with WT mice. Stab1 KO is not significantly different from Stab DK in circulatory …

Articulating The Importance Of Broader Relevance In Cures: From Definition To Actualization, Nikki T. Donegan

Open Access Theses & Dissertations

Course-based undergraduate research experiences (CUREs) have emerged as a means to foster student' scientific reasoning, promote positive attitudes and motivations toward research, and increase persistence in the science, technology, engineering, and mathematics (STEM) fields. In contrast to prescriptive laboratories, students enrolled in CUREs employ techniques and skills to collaboratively and iteratively address novel questions of broad relevance to audiences beyond the scope of the course itself. While many of the core components of CUREs are perceived to be relatively straightforward, definitionally-speaking, this latter dimension of broader relevance has, in contrast, remained largely ill-defined. In the series of studies presented herein, …

Myonuclear Transcriptional Dynamics In Response To Exercise Following Satellite Cell Depletion, Yuan Wen, Davis A. Englund, Bailey D. Peck, Kevin A. Murach, John J. Mccarthy, Charlotte A. Peterson

Physical Therapy Faculty Publications

Skeletal muscle is composed of post-mitotic myofibers that form a syncytium containing hundreds of myonuclei. Using a progressive exercise training model in the mouse and single nucleus RNA-sequencing (snRNA-seq) for high-resolution characterization of myonuclear transcription, we show myonuclear functional specialization in muscle. After 4 weeks of exercise training, snRNA-seq reveals that resident muscle stem cells, or satellite cells, are activated with acute exercise but demonstrate limited lineage progression while contributing to muscle adaptation. In the absence of satellite cells, a portion of nuclei demonstrates divergent transcriptional dynamics associated with mixed-fate identities compared with satellite cell replete muscles. These data provide …

Understanding Climate Change: Communicating Topics On Climate Change To Non-Experts, Michaela K. Plagwitz

Honors Capstones

Climate change is an increasingly prevalent phenomenon in recent years as it affects every corner of the world. However, there is a gap of understanding between scientists and non-experts. Some non-experts are unsure if climate change affects local areas, and whether they can do anything to help mitigate it. Of course, climate change indeed affects every environment, even urban areas, and small actions are helpful in the overall mitigation of climate change. Therefore, this project was created in order to teach this information to non-experts. In order to do so, a target demographic was chosen, and an initial survey was …

Vitamin D3 Induces Mesenchymal-To-Endothelial Transition And Promotes A Proangiogenic Niche Through Igf-1 Signaling, Lei Chen, Anweshan Samanta, Lin Zhao, Nathaniel R. Dudley, Tanner Buehler, Robert J. Vincent, Jeryl Hauptman, Magdy Girgis, Buddhadeb Dawn

School of Medicine Faculty Publications

Biological Sciences; Physiology; Molecular Biology; Cell Biology

Enhanced Gamma Band Mutual Information Is Associated With Impaired Consciousness During Temporal Lobe Seizures, Puneet Dheer, Sandipan Pati, Kalyan Kumar Chowdhury, Kaushik Kumar Majumdar

Journal Articles

Background: Epileptic seizures are characterized by aberrant synchronization. We hypothesized that higher synchronization across the seizure onset zone (SOZ) channels during a temporal lobe seizure contributes to impaired consciousness. New method: All symmetric bivariate synchronization measures were extended to multivariate measure by a principal component analysis (PCA) based technique. A novel nonparametric method has been proposed to test the statistical significance between increased synchronization across the seizure onset zone (SOZ) channels and reduced consciousness. Results: Increased synchronization in the gamma band towards seizure termination significantly contributes to impaired consciousness (p < 0.1). Synchronization reaches its peak in the extratemporal region (frontal lobe) ahead of the temporal region (p < 0.05). Synchronization is prominent in beta and gamma bands by most methods and it is more in the second half of seizure duration than in the first (p < 0.05). Conclusions: Mutual information is the only synchronization measure out of the six that we studied, whose increase can be associated with the loss of consciousness in a statistically significant way.

Chronic Consumption Of A Western Diet Induces Robust Glial Activation In Aging Mice And In A Mouse Model Of Alzheimer’S Disease, Leah C. Graham, Jeffrey M. Harder, Ileana Soto Reyes, Wilhelmine N. De Vries, Simon W. M. John, Gareth R. Howell

Ileana Soto Reyes

Studies have assessed individual components of a western diet, but no study has assessed the long-term, cumulative effects of a western diet on aging and Alzheimer’s disease (AD). Therefore, we have formulated the first western-style diet that mimics the fat, carbohydrate, protein, vitamin and mineral levels of western diets. This diet was fed to aging C57BL/6J (B6) mice to identify phenotypes that may increase susceptibility to AD, and to APP/PS1 mice, a mouse model of AD, to determine the effects of the diet in AD. Astrocytosis and microglia/monocyte activation were dramatically increased in response to diet and was further increased …

Fungal Physiological Response To Temperature And Nitrogen Availability, Shana Whitney

Master's Theses and Capstones

Microbial carbon use efficiency (CUE; the proportion of assimilated C allocated towards biomass production) is a powerful regulator of global soil C stocks, and there is a need to integrate accurate and dynamic CUE parameters into C-cycling models. However, there is uncertainty in the CUE-response to warming and increased N deposition and how this response may vary among groups of microbes. Here, we conducted a laboratory incubation of ten photogenically diverse fungal isolates grown in liquid culture to evaluate potential environmental and physiological controls over CUE. Isolates were grown at 15ºC or 25ºC and under low or high N concentrations. …

Investigating Limitations To Nitrogen Fixation By Leguminous Cover Crops In South Texas, Stephanie L. Kasper

Theses and Dissertations

Many farms use leguminous cover crops as a nutrient management strategy to reduce their need for nitrogen fertilizer. Biological nitrogen fixation (BNF) depends on a symbiotic relationship between legumes and nitrogen-fixing rhizobia. Under certain conditions despite inoculation with appropriate rhizobial strains, this symbiosis fails to form. Such failure was observed in a 14-acre winter cover crop trial in the Rio Grande Valley of Texas when three legume species (forage pea, crimson clover, and hairy vetch) produced no signs of nodulation and nitrogen fixation. This study examined nitrogen, phosphorus, moisture, micronutrients and biotic interference as potential causes for the failure and …

Effects Of Elevated Temperature On Gonadal Functions, Cellular Apoptosis, And Oxidative Stress In Atlantic Sea Urchin (Arbacia Punctulata), Jackson B. Johnstone

Theses and Dissertations

Increasing surface sea water temperatures effect on growth, reproduction and development in marine organisms. Sea urchins are excellent indicator species and ideal model organisms to focus on. In this study, I tested the effect of higher temperatures on reproductive functions, heat shock protein (HSP, a chaperone protein produced by cells in response to heat stress) and nitrotyrosine protein (NTP, an indicator of reactive nitrogen species, RNS) expressions, cellular apoptosis, and ceolomic fluid (CF, a body fluid which regulates important physiological processes) conditions in Atlantic sea urchin at three different temperatures. Ten sea urchins were placed in each of six aquariums …

Social Influences On Vocal Development In A Wild Parrot, Caleb Michael Mand Arellano

Theses and Dissertations

The Social Intelligence Hypothesis (SIH) proposes that navigating complex societies requires increased neural processing to monitor shifting relationships. SIH evolutionary focus of brain size variation in adults originally ignored the developmental frameworks leading to increased sociality. I studied the ontogeny of early sibling interactions in wild Green‐rumped Parrotlets (Forpus passerinus) in a Venezuelan population that exhibited large variation in brood size and engaged in elaborate social interactions and complex social learning during nestling development. To test if nests of varying brood sizes exhibit differences in social complexity, I quantified the number and strength of sibling play partnerships, as a proxy …

Object Enhancement, Noise Reduction, Conversion And Collection Of Spatiotemporal Image Data, Michael Izaguirre Jr.

Theses and Dissertations

In this report, a variety of cellular dynamics are enhanced and analyzed utilizing various algorithms and filter for contrast enhancement. This report will also illustrate the underlying complexities of processing compressed data received from certain type of sensors, their default applications, various methods in converting compressed data to compatible universal uncompressed formats allowed in scientific applications, various methods of image and video capture, guidelines in ethical image manipulation, various methods of frame extraction, and analyzing/processing video images. These methods and processes purposely utilize freeware and public domain software to lower the cost of reproducibility for all.

A Snapshot Of The Age, Growth, And Reproductive Status Of Gray Triggerfish (Balistes Capriscus, Gmelin 1789) On Three Artificial Reefs In The Northwest Gulf Of Mexico, Adam M. Lee

Theses and Dissertations

Age, growth, and reproductive status of gray triggerfish (Balistes capriscus) were identified from 2015-2016 on artificial reefs in the northwest Gulf of Mexico. Individuals ranged from 232-432 mm fork length with and a mean fork length of 319 mm. Individuals from age 0.2 to 5.2 yrs were observed with a weight to length relationship of Wg = 1.1 x -104 x FL2.7 (r2 = 0.94, n = 112), where FL = fork length (mm) and Wg = weight (g). A von Bertalanffy growth equation of Lt = 326(1 - e - 0.9 (t + 1.71)) was calculated irrespective …

The Snail Communities Of The Lower Rio Grande Valley, Briante Shevon Lewis Najev

Theses and Dissertations

The lower Rio Grande Valley are three lowest latitudinal counties of south Texas. This unique, subtropical, semi-arid environment has the last remaining Tamaulipan Thornforest in the USA. The Tamaulipan Thornforest of Texas occupies less than 2% of their former range. Before this habitat possibly dwindles even more there has been a comprehensive survey of terrestrial snails in intact, altered, and urban areas of the valley.

We have found a 16397 of snails in intact sites with a high species richness at 17; followed by altered sites with an abundance of 12143 snails and a species richness of 15. There were …

High-Powered Led Assembly As Replacement For Conventional Hbo Lamps In High Resolution, High Magnification Microscopy System, Christian Y. Miranda Solis

Theses and Dissertations

Through decades, fluorescence microscopy has relied on mercury vapor lamps as its main light source due to their high power, broad spectrum, and bright bands generated. The use of mercury vapor lamps has remained fundamentally associated with the required lighting equipment of research laboratories. However, as time progresses, mercury vapor lamps have been slowly replaced with technologies that offered longer lifetimes, less hazardous waste, and output light at similar intensities. LED technology has been actively used in biological research, but was limited by price, lower intensities, and stability. The recent development of LED technology has allowed for further understanding and …

Effects Of Global Warming On Gonadal Functions, Cellular Apoptosis, And Oxidative Stress In The American Oyster, Sarah B. Nash

Theses and Dissertations

Global warming due to climate change is predicted to intensify the heat stress in marine and coastal organisms, affecting their development, growth and reproductive functions. In this study. I analyzed gonadal development, heat shock protein 70 (HSP70), nitrotyrosine protein (NTP), dinitrophenyl (DNP) expressions, cellular apoptosis, and coelomic fluid (CF) conditions in American oyster. Oysters were placed in six aquariums and exposed to control (24°C), medium (28°C), and high (32°C) temperatures for one week. Higher temperature significantly decreased the number of eggs and sperm. CF protein concentrations also declined compared to control. In contrast, CF pH and HSP70 expression in gonad …

Comparative, Lithology-Based Biogeography Of Phreatophytic And Non-Phreatophytic Matrices Along A North-South Gradient Of The South Texas Sand Sheet, Christopher Joaquin Munoz

Theses and Dissertations

A 70km-long, N-S stratigraphic cross-section spanning the entirety of the South Texas Sand Sheet (STSS) has been rendered along Texas Interstate 69C. The cross-section indicates localized zones of substantial STSS thickness (>4m) as well as zones where the underlying Goliad Formation is either exposed or just below the surface (<0.5m). Spatial patterns of both phreatophytic and non-phreatophytic vegetation assemblages were analyzed by means of remote sensing and in situ herbaceous species evaluations. Biogeographic patterns were comprised of three vegetation matrices: P. glandulosa-dominated, Q. virginiana-dominated and a matrix of herbaceous plants. The geographic distribution of vegetation assemblages corresponded to specific lithological parameters and features. Namely, Q. virginiana concentration exhibited a positive linear relationship with the lithological parameter of STSS thickness (P =2.122x10-11) (α = 0.05). P. glandulosa/V. farnesiana …

Investigating The Efficacy Of Jak2/Stat3 And Pi3k/Mtor Inhibitory Therapy On Glioblastoma And Basal Breast Cancer Cell Lines, Neftali Vazquez

Theses and Dissertations

The JAK2/STAT3 and PI3K/mTOR pathways are upregulated in cancer cells. STAT3 promotes a positive feedback loop for the stem program whereas PI3K inhibits FOXOs tumor suppressor role and mTOR promotes tumorigenic advancement. Particularly GBM and BBC are aggressive cancers that have a stem cell population that makes them difficult to treat and metastatic. We investigate the effects of a combination therapy that inhibits JAK2/STAT3 and PI3K/mTOR output aiming to lower stem cell expression and promote apoptosis. Our collected data indicates that the dual inhibition induced apoptosis, but it also increased the stem cell signature. The findings pave way for the …

Studies On Regulation Of Dihydrotestosterone Binding To And Releasing From The Androgen Receptor, Tahyra M. Resto Santos

Theses and Dissertations

The androgen receptor (AR) is a nuclear receptor that responds to testosterone and 5α-dihydrotestosterone (DHT) by modulating gene expression in a variety of cells, including prostate and muscle tissues. Studies of small molecule interaction with AR have shown diphenyl compounds inhibiting through a binding site labeled Binding Function 3 (BF3). The goal of this research is to analyze the interaction of the BF3 site with DDT and related compounds. Amino acids that are exposed on the surface of the BF3 that might interact with different compounds were mutated. Mutant proteins were tested experimentally for their impact on the inhibition the …