Bacterial anti-inflammatory lipid mediatorsProject Number: 1K99AI166116-01

Bacterial anti-inflammatory lipid mediatorsProject Number: 1K99AI166116-01

Postdoctoral Mentor: Charles O. Rock, Ph.D.• Operated Whitley M45 workstation to grow microbiome anaerobic bacteria Alistipes finegoldii and Bifidobacterium breve for metabolic radiolabeling experiments • Measured murine macrophage response to microbiome lipids under hundreds of conditions using high throughput luciferase reporter cell-based assay • Measured human T cell response to microbiome lipids presented by 4 unique CD1 antigen presenting molecules using flow cytometry • Operated analytical gas chromatography instrument to measure fatty acid composition of 5 unique bacterial genera • Utilized HPLC and UV-Vis for 10-point protein assay measuring acyl-coenzyme A oxidation • Measured lipid and nucleotide cofactor binding to 4 proteins using differential scanning fluorimetry and surface plasmon resonance • Implemented preparative Protein A-based purification of polyclonal antibody from rabbit serum and assessed antibody quality by immunoblot of wildtype and knockout cell lysates • Utilized small angle X-ray scattering and analytical ultracentrifugation to analyze stoichiometry and oligomerization of 10 unique protein-protein complexes • Developed novel method for co-crystallizing insoluble fatty acid ligands with fatty acid binding proteins by in-vitro protein-protein interactions and validation by X-ray diffraction • Developed and executed laboratory work plans and schedules for self and other members of a multidisciplinary team of 3 experienced researchers using performance standards and metrics

Thesis mentors: Lawrence J. DeLucas, O.D., Ph.D.; Stephen G. Aller, Ph.D. • Operated ÄKTA Avant Chromatography System to purify 10 unique proteins for graduate training and NASA project • Achieved 5x improvement in solubility of recombinant cysteine-rich protein by engineering constructs with point mutations based on computational homology modeling • Crystallized 2 membrane proteins by lipidic cubic phase-based crystallization • Purified 4-component ABC transporter by engineering nickel affinity tag on cytoplasmic ATPase • Developed novel method for purification of substrate-free metal atom-binding proteins by in-vivo protein-protein interactions and validation by X-ray fluorescence • Determined ligand-free and ligand-bound protein structures utilizing 2 synchrotron radiation sources (APS and CMCF) • Collected X-ray diffraction data from 40 protein crystals grown on Earth and in microgravity for NASA project • Processed X-ray diffraction data collection statistics from NASA project utilizing commonly used structural biology software packages • Responsible for maintaining a clear and precise laboratory notebook, and planning and coordinating literature searches • Managed the training and supervision of 2 undergraduates and 3 graduate students in structural biology projects through democratic leadership and consensus decision making to expand breadth and depth of their technical expertise

•Organized and performed multivariate statistical analysis on patient data collected over a multi-month period•Compiled results and assisted in writing report that was published and circulated in Orlando Health administration on behalf of Transition Services Department•Assisted in administrative protocol development for new program growth•Awarded 2011 ‘I Care’ volunteer award for extensive service to the Transition Services Spiritual Care Department

• Maintained and operated industrial 100-liter fermenter and hollow fiber harvest column for growing bacteria Azotobacter vinelandii and Klebsiella pneumoniae under anaerobic conditions for protein production • Purified 2 unique proteins sensitive to oxidative stress using ÄKTA Purifier Chromatography System • Used gas manifold for gas exchange to establish anaerobic conditions and crimp seal system for long-term storage of purified proteins sensitive to oxidative stress • Designed colorimetric ligand binding assay to measure enzymatic activity in an anaerobic glovebox in the dark • Managed the training of 5 undergraduates on laboratory instrumentation, troubleshooting, and problem-solving skills for independent execution of standard laboratory procedures and methods