Torrey Pines Institute for Molecular Studies

About: Torrey Pines Institute for Molecular Studies is a nonprofit organization based out in San Diego, California, United States. It is known for research contribution in the topics: Antigen & T cell. The organization has 2323 authors who have published 2217 publications receiving 112618 citations.

Proteomic tools for quantitation by mass spectrometry.

Abstract: I. Introduction 182 II. Relative Quantitation 184 A. 2-D Gel Electrophoresis 184 B. Metabolic Isotopic Labeling 185 1. 15N 185 2. 13C Enrichment and Depletion 185 3. Select Isotopic Amino Acid Incorporation 186 C. Chemical Labeling 186 1. Labeling During Proteolysis: 18O Incorporation During Enzymatic Cleavage 186 2. Isotopic Tags—Isotope-Codes Affinity Tag Reagents (ICAT) 187 3. Isotopic Tags—Acid-Labile Isotope-Coded Extractants (ALICE) 188 4. Lysine-Specific Labeling 189 5. Phosphoserine- and Phosphothreonine-Specific Labeling 189 6. N-Terminus Labeling 189 a. Nicotinyl-N-hyxroxysuccinimide 189 b. Acylation 189 c. Amidination-quantitation using enhanced signal tags (QUEST) 190 d. C-terminusu labeling 190 D. Differential Mass Mapping 190 III. Absolute Quantitation 191 IV. Choice of Instrumentation 191 V. Overview, Discussion, and Future Directions 191 References 192 Techniques for the quantitation of proteins and peptides by mass spectrometry (MS) are reviewed. A range of labeling processes is discussed, including metabolic, enzymatic, and chemical labeling, and techniques that can be employed for comparative and absolute quantitation are presented. Advantages and drawbacks of the techniques are discussed, and suggestions for the appropriate uses of the methodologies are explained. Overall, the metabolic incorporation of isotopic labels provides the most accurate labeling strategy, and is most useful when an internal standard for comparative quantitation is needed. However, that technique is limited to research that uses cultured cells. © 2003 Wiley Periodicals, Inc., Mass Spec Rev 22:182–194, 2003; Published online in Wiley Interscience (www.interscience.wiley.com). DOI 10.1002/mas.10048

Combining protein-based IMAC, peptide-based IMAC, and MudPIT for efficient phosphoproteomic analysis.

Abstract: Immobilized metal affinity chromatography (IMAC) is a common strategy used for the enrichment of phosphopeptides from digested protein mixtures. However, this strategy by itself is inefficient when analyzing complex protein mixtures. Here, we assess the effectiveness of using protein-based IMAC as a pre-enrichment step prior to peptide-based IMAC. Ultimately, we couple the two IMAC-based enrichments and MudPIT in a quantitative phosphoproteomic analysis of the epidermal growth factor pathway in mammalian cells identifying 4470 unique phosphopeptides containing 4729 phosphorylation sites.

Cartilage production by rabbit articular chondrocytes on polyglycolic acid scaffolds in a closed bioreactor system

Abstract: Rabbit articular chondrocytes were seeded onto three-dimensional polyglycolic acid (PGA) scaffolds and placed into a closed bioreactor system. After 4 weeks of growth, meshes were examined for cartilage formation. Gross examination revealed solid, glistening material that had the appearance of cartilaginous tissue. Histologic examination revealed cell growth and deposition of extracellular matrix throughout the mesh with a less dense central core. Alcian blue and Safranin 0 staining showed deposition of glycosaminoglycans (GAGs). Immunostaining showed positive reactivity for type II collagen and chondroitin sulfate and no reactivity for type I collagen. Biochemical analysis showed collagen and GAG values to be 15% and 25% dry weight, respectively. Our results indicate that this type of system compares well with those previously described and should be useful for producing cartilage for evaluation in a clinical setting. (c) 1995 John Wiley & Sons, Inc.

Discovery of diverse thyroid hormone receptor antagonists by high-throughput docking

Abstract: Treatment of hyperthyroidism, a common clinical condition that can have serious manifestations in the elderly, has remained essentially unchanged for >30 years. Directly antagonizing the effect of the thyroid hormone at the receptor level may be a significant improvement for the treatment of hyperthyroid patients. We built a computer model of the thyroid hormone receptor (TR) ligand-binding domain in its predicted antagonist-bound conformation and used a virtual screening algorithm to select 100 TR antagonist candidates out of a library of >250,000 compounds. We were able to obtain 75 of the compounds selected in silico and studied their ability to act as antagonists by using cultured cells that express TR. Fourteen of these compounds were found to antagonize the effect of T3 on TR with IC50s ranging from 1.5 to 30 μM. A small virtual library of compounds, derived from the highest affinity antagonist (1-850) that could be rapidly synthesized, was generated. A second round of virtual screening identified new compounds with predicted increased antagonist activity. These second generation compounds were synthesized, and their ability to act as TR antagonists was confirmed by transfection and receptor binding experiments. The extreme structural diversity of the antagonist compounds shows how receptor-based virtual screening can identify diverse chemistries that comply with the structural rules of TR antagonism.