A5013114112- RNA and protein synthesis mechanisms
- Protein Structure and Dynamics
- Monoclonal and Polyclonal Antibodies Research
- RNA modifications and cancer
- Enzyme Structure and Function
- Chemical Synthesis and Analysis
- Bacterial Genetics and Biotechnology
- Genomics and Phylogenetic Studies
- T-cell and B-cell Immunology
- CRISPR and Genetic Engineering
- Paraoxonase enzyme and polymorphisms
- Glycosylation and Glycoproteins Research
- Advanced Proteomics Techniques and Applications
- Microbial Metabolic Engineering and Bioproduction
- Ubiquitin and proteasome pathways
- Cell Image Analysis Techniques
- Biotin and Related Studies
- Biochemical Acid Research Studies
- Advanced Fluorescence Microscopy Techniques
- Bioinformatics and Genomic Networks
- Cellular transport and secretion
- Data Visualization and Analytics
- Fungal and yeast genetics research
- Evolution and Genetic Dynamics
- Mass Spectrometry Techniques and Applications
The Ohio State University
2013-2024
The Ohio State University Comprehensive Cancer Center – Arthur G. James Cancer Hospital and Richard J. Solove Research Institute
2024
Ohio University
2017
Area Cooperative Education Services
2006-2012
Yale University
2004-2006
University of California, Berkeley
1997-2002
Scripps Research Institute
2000
Howard Hughes Medical Institute
1997
Kenyon College
1997
Identification of protein binding partners is one the key challenges proteomics. We recently introduced a screen for detecting protein−protein interactions based on reassembly dissected fragments green fluorescent fused to interacting peptides. Here, we present set comaintained Escherichia coli plasmids facile subcloning fusions fragments. Using library antiparallel leucine zippers, have shown that can detect very weak (KD ≈ 1 mM). In vitro kinetics show reaction essentially irreversible,...
The low stability of natural proteins often limits their use in therapeutic, industrial, and research applications. scale throughput methods such as circular dichroism, fluorescence spectroscopy, calorimetry severely limit the number variants that can be examined. Here we demonstrate a high-throughput thermal scanning (HTTS) method for determining approximate stabilities protein at high cost. is based on binding to hydrophobic dye akin ANS, which fluoresces upon molten globules denaturation...
In an effort to expand the scope of protein mutagenesis, we have completed first steps toward a general method allow site-specific incorporation unnatural amino acids into proteins in vivo. Our approach involves generation "orthogonal" suppressor tRNA that is uniquely acylated Escherichia coli by engineered aminoacyl-tRNA synthetase with desired acid. To this end, eight mutations were introduced tRNA2Gln based on analysis x-ray crystal structure glutaminyl-tRNA aminoacyl (GlnRS)-tRNA2Gln...
ADVERTISEMENT RETURN TO ISSUEPREVCommunicationNEXTA New Functional Suppressor tRNA/Aminoacyl−tRNA Synthetase Pair for the in Vivo Incorporation of Unnatural Amino Acids into ProteinsLei Wang, Thomas J. Magliery, David R. Liu, and Peter G. SchultzView Author Information Department Chemistry, The Scripps Research Institute 10550 North Torrey Pines Road, La Jolla, California 92037 Cite this: Am. Chem. Soc. 2000, 122, 20, 5010–5011Publication Date (Web):May 4, 2000Publication History Received18...
Biological activity in proteins requires them to share the energy landscape for folding and global conformational motions, 2 key determinants of function. Although most structural studies date have focused on fluctuations around a single basin, we directly observe coexistence symmetrically opposed conformations mutant Rop-homodimer (Repressor Primer) single-molecule fluorescence resonance transfer (smFRET) measurements. We find that mild denaturing conditions can affect sensitive balance...
Somatic hypermutation and clonal selection lead to B cells expressing high-affinity antibodies. Here we show that somatic mutations not only play a critical role in antigen binding, they also affect the thermodynamic stability of antibody molecule. directly involved recognition by 93F3, which binds relatively small hapten, reduce melting temperature compared with its germ-line precursor up 9 °C. The destabilizing effects these are compensated additional located on surface loops distal...
The recent explosion in the availability of complete genome sequences has led to cataloging tens thousands new proteins and putative proteins. Many these can be structurally or functionally categorized from sequence conservation alone. In contrast, little attention been given meaning poorly-conserved sites families proteins, which are typically assumed structural functional importance.Recently, using statistical free energy analysis tetratricopeptide repeat (TPR) domains, we observed that...
<h3>Background</h3> Notch intercellular communication instructs tissue-specific T-cell development and function. In this study, we explored the roles of dendritic cell (DC)-expressed ligands in regulation effector <h3>Methods</h3> We generated mice with CD11c lineage-specific deletion Delta-like ligand <i>(Dll)1</i> Jagged <i>(Jag)2.</i> Using these genetically-ablated engineered pharmacological constructs, various T-cell-mediated immunity were investigated. assessed tumor growth, mouse...
Human serum paraoxonase-1 (HuPON1) has the capacity to hydrolyze aryl esters, lactones, oxidized phospholipids, and organophosphorus (OP) compounds. HuPON1 bacterially expressed chimeric recombinant PON1s (G2E6 G3C9) differ by multiple amino acids, none of which are in putative enzyme active site. To address importance these acid differences, abilities HuPON1, G2E6, G3C9, several variants phenyl acetate, paraoxon, V-type OP nerve agents were examined. G2E6 have a 10-fold greater catalytic...
Abstract Mammalian paraoxonase-1 (PON1) is a ~ 39.45 kDa calcium-dependent hydrolytic enzyme with potential therapeutic applications in chemical defense and cardiovascular disease. The N-terminus of PON1 embedded the cellular membrane, imparting to hydrophobic character that leads increased aggregation propensity instability during purification. Although some advances have been made bacterial expression hosts by using solubility-enhancing fusion tags detergent solubilization strategies,...
Several steps have been completed toward the development of a method for site-specific incorporation unnatural amino acids into proteins in vivo. Our approach consists generation amber suppressor tRNA/aminoacyl-tRNA synthetase pairs that are orthogonal to all Escherichia coli endogenous tRNA/synthetase pairs, followed by directed evolution aminoacyl-tRNA synthetases alter their amino-acid specificities. A new pair E. has derived from Saccharomyces cerevisiae tRNAAsp and aspartyl-tRNA...
Abstract The X‐ray crystal structure of a bovine antibody (BLV1H12) revealed unique in its ultralong heavy chain complementarity determining region 3 (CDR3H) that folds into solvent‐exposed β‐strand “stalk” fused to disulfide crosslinked “knob” domain. We have substituted an antiparallel heterodimeric coiled‐coil motif for the stalk this antibody. resulting (Ab‐coil) expresses mammalian cells and has stability similar parent MS analysis H–D exchange supports peptides. Substitution...
Identification of protein–protein interactions is critical for understanding protein function and regulation. Split reassembly an in vivo probe that circumvents some the problems with yeast 2-hybrid (indirect interactions, false positives) co-immunoprecipitation (loss weak transient decompartmentalization). GFP reassembly, also called Bimolecular Fluorescence Complementation (BiFC), especially attractive because chromophore forms spontaneously on folding virtually every cell type tested....
The identification of founder mutations in cancer predisposing genes is important to improve risk assessment geographically defined populations, since it may provide specific targets resulting cost-effective genetic testing. Here, we report the characterization BRCA1 c.190T>C (p.Cys64Arg) mutation, mapped RING-finger domain coding region, that detected 43 hereditary breast/ovarian (HBOC) families, for large part originating from province Bergamo (Northern Italy). Haplotype analysis was...
The monoclonal antibody 48G7 differs from its germline precursor by 10 somatic mutations, a number of which appear to be functionally silent. We analyzed the effects individual mutations and combinations thereof on both binding affinity thermal stability. Individual that enhance hapten decrease stability antibody; combining these produced mutant with high for but exceptionally low Adding back each remaining restored These results, in conjunction recently published studies, suggest an...