2016-2019 BioXFEL Research

2019 BioXFEL Lab


Virginia Commonwealth University

Nadia (in Jane Tao’s lab) aimed to study the protein structure of human astrovirus (HAstV), a pathogen associated with gastroenteritis. Specifically, her plan was to express and purify the VP90 dimer, the precursor, immature capsid protein of HAstV. With the purified VP90, she conducted crystal screenings under various conditions. The crystals that were produced will be used in X-ray crystallography in order to capture images of the protein that reveal its specific structure. Having this knowledge is vital in understanding the life cycle of the virus and possibly creating vaccines against it.


University of Vermont

Trace (in Yousif Shamoo's lab) studied different variants of penicillin-binding proteins (PBPs) from the hospital pathogen Enterococcus faecium in hopes of determining the structural basis that confers antibiotic resistance. This was done by purifying these PBPs with the ultimate goal of growing protein crystals from them. Protein crystals are necessary for X-ray crystallography which allows us to determine the molecular structure of said proteins.


University of Puerto Rico
Industrial Microbiology

Gabriela (in Joff Silberg’s lab) worked to purify cyanophage ferredoxin (Fd), an iron-sulfur (Fe-S) cluster-containing protein that functions as an electron carrier in biological cells. These low potential oxidoreductases serve as electron hubs, participating in electron transfer reactions with a variety of protein partners. Specifically, Gabriela’s research focused on the analysis of phage Fd stability, which is believed to be more robust than host Fds. To determine their stability through in vitro assays, she first overexpressed the phage Fd in the Escherichia coli BL21(DE3) strain and then purified the protein from cell lysates using chromatography techniques. She confirmed the homogeneity of purified samples through SDS-PAGE. Finally, she evaluated Fd stability by using circular dichroism to quantify the loss of folded protein in incrementally denaturated samples.


University of North Carolina

Jonathan (in George Phillip’s lab) worked on structure determination of the New Delhi metallo-beta-lactamase 1 (NDM-1) enzyme in complex with novel inhibitors developed by collaborators from the Bugni Lab at the University of Wisconsin. NDM-1 is a class B lactamase which confers antibiotic resistance to Klebsiella pneumoniae by catalyzing the hydrolysis of beta-lactam antibiotics in a zinc dependent manner. Jonathan worked on protein expression in E. coli and subsequent protein purification via a combination of affinity and size exclusion chromatography. Successful structure determination of the protein-ligand complex would provide insight for the mechanism of hydrolysis of NDM-1 and could inform the design of novel antibiotics.


University of Texas at Austin

Esther (in George Phillip’s lab) focused on the expression, purification, characterization, and crystallization of Adenylate Kinase (AK) from the hyperthermophile, Thermus thermophilus. AK is a model enzyme for the study of protein dynamics, as it is a small enzyme that undergoes large conformational rearrangements. It catalyzes the interconversion of the nucleotides ATP, ADP, and AMP. Protein production included molecular cloning of a DNA construct and protein expression in Escherichia coli cells, while protein purification involved affinity and size exclusion chromatography. The purified AK was used for temperature dependent activity assays and crystallization experiments. The overall goal of her research was to use X-ray crystallography to investigate the structure and dynamics of AK, with the ultimate goal of producing a protein movie through time-resolved crystallography.


Howard University
Computer Science

Jurnee (in Jane Tao’s lab) worked to solve the atomic level structures related to the human astrovirus (HAstV), which is an emerging virus that is a leading cause of gastroenteritis. More specifically, her role in this project was to test new purification methods to isolate fully assembled viral particles. Viral protein 90 (VP90) is the capsid polyprotein “foundation” of HAstV. Within the the cell, it is assembled into viral particles and is further matured to yield viral protein 70 (VP70), which correlates with the cell release of the virus. Because of this relationship, solving and understanding the structure of VP90 has potential applications in vaccine development and the creation of small-molecule drugs that would target astrovirus assembly and maturation. The goal of this project was to isolate the construct of VP90 in oligomer form in order to visualize the particle using cryo-electron microscopy and later create 3D images.


University of Arkansas at Pine Bluff

Justin (in Yousif Shamoo's lab) worked on purifying YycF, which is a response regulator (RR) of the YycFG two-component signal transduction system in Enterococcus faecium. Clinical isolates of E. faecium have shown high resistance to antibiotics. E. faecium is a human pathogen that causes nosocomial bacteremia, surgical wound infection, and other conditions. His goal was to produce high yields of purified YycF protein, to grow crystals and then solve the X-ray crystal structure to determine the molecular basis for antibiotic resistance.


Rice University
Mathematics and Computer Science

Robert (in George Phillips’ lab) worked on recovering the three-dimensional structure of proteins from diffraction data. This involved working with the Hybrid Input Output Algorithm and focusing on the applications of specific code, while attempting to maximize its efficiency and accuracy.


Rice University
Computer Science and Statistics

Yong (in George Phillips’ lab) researched areas involving the field of computational biology, particularly in the realms of x-ray crystallography. His project involved working on overcoming the infamous phase problem regarding devising protein structures. The Hybrid Input Output (HIO) algorithm has had its successes in overcoming the problem but the scope of structures for which it can overcome the problem is limited. He combined specific code with the HIO code to result in an algorithm that has minimal constraints on what structures it can figure out.

2018 BioXFEL Lab


St. Mary's University

Alyssa (in Joff Silberg’s lab) worked on expressing and purifying a Clostridium acetobutylicum flavodoxin (CacFld6) in Escherichia coli cells for structural analysis. Using plasmids containing the gene for CacFld6, obtained from a graduate lab member, Alyssa transformed Rosetta DE3 cells with the expression vector and identified the optimal conditions of expression by varying incubation temperatures and IPTG induction concentrations. She measured expression by analysis on SDS-PAGE gels. From this, she overexpressed the protein in E. coli, purified CacFld6 by ion exchange, and will be screening for conditions where CacFld6 crystallizes for crystallography studies. The overall goal of this project is to determine the structure of the protein for applications in bioelectronics.


Houston Community College

German (in George Phillips’ lab) worked on determining CalU17’s molecular structure through X-ray crystallography. A protein from the Calicheamicin biosynthetic gene cluster of Micromonospora echinospora, CalU17 is believed to be involved in the synthesis of Calicheamicin thorough post-translational sulfatase modification. German’s work involves the cloning of the gene calU17 into a DNA plasmid as well as the expression of CalU17 in bacterial cells to produce a soluble protein. He hopes to crystalize the protein in order to obtain diffraction data and generate a model through which the specific functions of CalU17 can be determined. Calicheamicin is currently being studied as a viable anticancer drug treatment due to its cytotoxic properties and potential for selectivity of human cancer cells. Determining the role CalU17 plays in the production of Calicheamicin can further the production of pharmaceuticals to treat the disease.


Rice University
Biochemistry and Cell Biology

Sarah (in Bonnie Bartel’s lab) worked to express and purify three peroxisomal proteins from Arabidopsis thaliana: PEX5, and PEX4 in complex with PEX22. After designing multiple construct variations, she has expressed these proteins in E. coli to be purified and later crystallized. She will then investigate the structural changes that accompany ligand binding and interactions with binding partners.


University of Texas at Austin

Esther (in Natasha Kirienko’s lab) focused on the expression and purification of wild-type JMJC-1, human homolog RIOX1 and JMJC-19ala. The proteins were expressed in Escherichia coli cells, were lysed using lysozyme and sonication, and purified with a Ni-NTA resin. She aims to determine whether mutations altered the protein folding once the protein purification is optimized. The purified proteins will then be used in the determination of its role in the novel ESRE (Ethanol and Stress Response Element) mitochondrial surveillance pathway.


Texas A&M University
Biomedical Science

Haania (in Yizhi (Jane) Tao’s lab) worked on optimizing conditions for expressing and purifying the DNA polymerase of nackedna-viruses, a family of fish viruses that bear striking similarities to the Hepatitis B Virus (HBV). After transforming competent E. coli cells with transgenic plasmids containing the gene for her protein, she expressed and purified it with affinity chromatography and size-exclusion. Ultimately, Haania hopes to obtain DNA polymerase solutions to grow crystals for x-ray crystallographic analysis, providing new insight into the structure of the enzyme. This will in turn inform new anti-viral therapies for HBV.


UT MDACC School of Health Professions
Molecular Genetics Technology

Tasmia (in Yousif Shamoo’s lab) studied the function of mutated genes FolP from Nocardia Strain that were resistance to TMP-SMX antibiotics. Her work involved studying the structure and function of mutated FolP gene. She is currently trying to make constructs of FolP Asp172Glu mutated gene with a solubility tag to test the protein solubility. Based on the protein solubility she hopes to purify and foster the protein for crystallization to understand the mechanism behind the structure and enzymatic assays of the mutated protein.


University of North Carolina at Charlotte
Biology and Psychology

Chiagoziem (in George Phillips’ lab) worked to express and purify 2-hydroxyacyl-CoA lyase (HACL). The discovery that HACL catalyzes a novel C-C bond forming reaction with C1 substrates shows great potential for use in industry. We plan to understand its protein structure and function with its application in the context of our synthetic alpha-reduction cycle. As of now, we are focused on biochemical characterization where we are expressing and purifying proteins, ultimately, for X-ray crystallization. The lab is striving to demonstrate and improve the performance of a simple biological conversion platform that integrates C1 molecules into multi-carbon products, to determine the crystal structure of HACL and its tertiary homologues, and to improve operation of the alpha-reduction cycle.


Prairie View A&M University
Chemical Engineering

Courage (in George Phillips’ lab) worked on the purification and crystallization of the protein Adenylate Kinase (ADK). I used the plasmid containing the DNA sequence of ADK and grew them in an E. coli for expression and added IPTG to induce the protein expression. To purify the ADK I used a column with Affi-gel cibacron blue for ion exchange chromatography. After that I did size exclusion chromatography. At the end I was able to obtain purified ADK for crystallization. My final goal is to use my crystals for X-ray diffraction for a better understanding of the structure and dynamics of ADK.


Austin College

Carlos (in Angel Marti's lab) focused on synthesizing Ru[phen]3/Cy5 doped Silicon Nano-particles for intracellular thermometry to aid in Radio Frequency Hyperthermia (RFH) treatments. The release and absorption of heat goes hand-to-hand with many of the cellular functions a cell performs, making the cell a platform for fluctuating temperatures. In addition to RFH treatments, this thermometer would be able to study these variations in temperature to give us a better understanding of the cell's functions. Specifically, we would be able to learn more about cellular pathogenesis; cancer. Cellular pathogenesis is characterized as the cells producing a tremendous amount of heat, so our molecular thermometer would be able to quantize this measurement to be able to learn more about such events.


University of Texas at Austin

Megan (in Zach Ball's lab) studied a copper catalyzed reaction generated by the Ball Lab similar to an already known reaction called Chan-Lam coupling. The ability to engineer reactivity within complex protein structures allows for applications such as elucidating cellular pathways, constructing biomaterials, and synthesizing therapeutic agents. Her lab tested this reaction on silicon surfaces and gold nanoparticles to discover how robust the reaction was and to see if it holds potential in other fields. Off-target modification of the protein was also found after the reaction was completed. An additional project she completed was synthesizing peptides and screening where this modification occurred using tandem mass spectrometry.

2017 BioXFEL Lab


Middlebury College
Natural Science

Oriyomi (in George Phillips Lab) worked to express and purify wild-type Myoglobin from the Dwarf Sperm whale using the PV80K vector. The overarching goal is to create human blood substitutes as a temporary substitute for those waiting on a suitable blood match for transfusion. Hemoglobin carries oxygen from lungs to various tissues in the body. Her lab looked for a stable hemoglobin-based oxygen carrier (HbOC) that would not disintegrate outside of the red blood cells and thus be used in the blood substitute. They used Dwarf Whale Myoglobin, one of the most stable myoglobin’s, as a model. She expressed the myoglobin gene in E. coli and purifying ultimately for X-ray crystallization. Once done, the cause of the myoglobin’s stability can be discovered and applied to making stable HbOC.


Houston Community College

Christopher (in John Olson’s Lab) focused on the expression and purification of the human sickle hemoglobin mutant protein(HbS) and one of its providence mutation Hbβ(E6V/K82D). He worked towards performing crystallization trials to better understand and characterize the effects of HbS Providence and its resistance to oxidative degradation through X-Ray diffraction. Understanding the molecular conformation of HbS providence clinically opens up doors to new therapies to treat Sickle Cell Disease patients.


Norfolk State University

Alexys (in Yousif Shamoo’s Lab) conducted site directed mutagenesis on the transmembrane protein LiaS of the LiaFSR antibiotic resistance pathway in Enterococcus faecalis. She aimed to lock the protein into its ‘phosphomimetic’ and ‘inactive’ states and crystallize it. The crystallization of LiaS in both states helped to determine the structure of the protein and provide insight into making resistant bacteria susceptible to antibiotics.


University of Texas at Austin

Laura (in Yizhi Jane Tao’s Lab) worked on expressing and purifying the Human Astrovirus Serotype-8 (HAstV-8) Capsid Protein VP90 for Functional and Structural Analysis. She tried to isolate the viral capsid precursor VP90(28-782) in order to study how it matures via proteolytic processes that the virus needs to become infectious. Structural studies of VP90 provide insight into how the capsid protein undergoes maturation and targets for antiviral compounds.


Harvard University

Caleb (in George Phillips’s Lab) worked on the purification and crystallization of the protein Adenylate Kinase (AK). Using plasmids containing the sequence for his protein, Caleb successfully transformed them into E. coli cells for expression. Through both affinity chromatography and size exclusion, Caleb was able to extract and purify his protein. With the utilization of various optimal crystallization conditions in several trays, Caleb successfully grew Adenylate Kinase crystals. Ultimately, Caleb hopes to diffract x-rays through the protein crystals to better understand AK’s structure and dynamics.


St. Mary's University

Jesus (in George Phillip's Lab) is on the forefront of the worldwide battle against antibiotic resistance. He investigated the New Delhi Metallo B-Lactamase (NDM-1) since it has given bacteria the ability of resistance to a wide range of antibiotics. Through the use of cloning and transformation methods NDM-1 has successfully been expressed. The protein was then purified via size exclusion and affinity chromatography and prepped for crystallization. Jesus not only used the protein crystals to determine the structure of NDM-1 but to also create a molecular movie of NDM-1 in action.


San Jacinto Community College

Marvin (in Yizhi Jane Tao’s Lab) overexpressed and purified the SO0839 protein in Escherichia coli, which originally comes from the Shewanella oneidensis bacteria. His work involved testing the binding conditions of the protein, along with crystal screening and structural investigation. The protein is believed to be a transcription factor in metabolic functions for bacteria. Finding a way to interrupt this protein’s function could potentially inhibit or stop biofilms from forming.


Rice University

Jessica (in Yousif Shamoo's Lab) focused on evolving resistance of Nocardia bacteria species to Trimethoprim-Sulfamethoxazole (TMP-SMX), an antibiotic of last resort used against bacterial infections. She looked at mutation comparisons among different Nocardia strains and species (N. nova and N. cyriacigeorgica). Jessica researched determining mechanisms of resistance of Nocardia to TMP-SMX.

2016 BioXFEL Lab


University of Houston

Eduardo (in John Olson’s lab) worked on the expression and purification of a recombinant hemoglobin mutant protein. His work involved crystallization trials of the isolated protein for data collection in an X-ray diffraction experiment. The structural investigation of hemoglobin mutants will help facilitate the development of Hemoglobin-based blood substitutes.


Rice University
Biochemistry & Cell Biology

Hector (in George Phillip's lab) is attempting to fight the spread of beta-lactamase inflicted bacterial pathogens by the means of X-ray crystallography. Through the use of cloning methods a plasmid containing NDM-1 has been synthesized and successfully transformed into BL21 E. coli cells. The purification of this protein was carried out through both size-exclusion and affinity chromatography using FPLC. Hector will ultimately determine optimal crystallization conditions for this protein and then go on to diffract the protein using X-rays and solve the structure.


Prairie View A&M University
Natural Sciences

Deja (In John Olson's lab) worked on expressing the protein, myoglobin, in E. coli in an attempt to try to find what makes the protein so stable. Her work involves purification of the recombinant protein for crystallization trials via the sitting drop diffusion method. Optimized crystals will be used for data collection in an X-ray diffraction experiment. Elucidating the structure of myoglobin will facilitate the investigation of key residues involved in protein stability.


Houston Community College

Nia (in George Phillips’ lab) worked on the beta-lactamase protein, KPC-2, a gram-negative bacterial enzyme from Klebsiella pneumoniae. Her work involves molecular cloning of the DNA encoding the KPC-2 sequence into a circular DNA plasmid, recombinant protein expression in E. coli, and protein purification. She is currently trying to generate a structural model of the enzyme. By crystallizing the specific enzyme, she hopes to obtain diffraction patterns for crystallographic data analysis by using a bright X-ray light source, such as a third-generation synchrotron.


Rice University
Natural Sciences

Mukund (in George Phillips' lab) is using X-ray crystallography to solve the structure of an enzyme, TEM-60. Through molecular cloning and recombinant protein expression methods, large amounts of TEM-60 can be produced in E. coli. This protein is then purified and concentrated to carry out crystallization experiments for the optimization of protein crystals. Mukund will ultimately collect data in the form of X-ray diffraction patterns to generate a structural model of TEM-60.


Trinity University

Maia (in Yousif Shamoo's lab) is focusing on molecular evolution to resistance in bacteria. She is looking at the mutations of Bacillus subtilis after adaptation to a chemically synthesized pyrrolidinone antibiotic and Pseudomonas aeruginosa after adaptation to a clinical last resort drug for gram negative bacteria. She has assisted with setting up growth curves, biochemical tests and minimum inhibitory concentration tests.


University of Texas at Austin

Kelly (in Yousif Shamoo's lab) worked on isolating LiaS, a transmembrane protein that is a part of the LiaFSR antibiotic resistance pathway of Enterococcus faecium and Enterococcus faecalis. Her project involved cloning and expressing the correct LiaS construct that was able to retain its kinase activity. Purification of the LiaS protein will allow for in vitro phosphorylation experiments of LiaR and lead to attempts to crystallize the histidine-kinase region of LiaS. Finding the crystal structure of LiaS would open new doors to finding drugs to combat common antibiotic resistant diseases.