Quantitative proteomics
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This document provides an introduction to quantitative proteomics using stable isotope labeling of amino acids in culture (SILAC). SILAC involves growing cells in media containing "light" or "heavy" isotopic forms of amino acids, which get incorporated into proteins and allow distinction and quantitation of proteins from different samples after mixing and mass spectrometry analysis. The document discusses applications of SILAC, how mass spectrometers identify isotopic species, and complementary labeling strategies like ICAT and iTRAQ.
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INTRODUCTION
STRUCTURAL PROTEOMICS
WHAT IS THE IMPORTANCE OF STUDY OF PROTEIN
METHODS FOR SOLVING PROTEIN STRUCTURE
1. X- RAY CRYSTALLOGRAPHY
INTRODUCTION
PROCEDURE
LIMITATIONS
2.NUCLEAR MAGNETIC RESONANCE
PROTEIN STRUCTURE DETERMINATION
3. MASS SPECTROMETER
MALDI
ESI
STRUCTURE MODELING
APPLICATIONS
CONCLUSION
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INTRODUCTION
STRUCTURAL PROTEOMICS
WHAT IS THE IMPORTANCE OF STUDY OF PROTEIN
METHODS FOR SOLVING PROTEIN STRUCTURE
1. X- RAY CRYSTALLOGRAPHY
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PROCEDURE
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2) Active site residues serine 17, arginine 54, and leucine 86 were selected for mutagenesis studies. Mutants showed altered substrate specificity and kinetic parameters compared to the wild-type.
3) Biophysical analysis found that AnAEst is most stable at pH 5.5 and temperatures of 25-45°C, with half
13128931.ppt
The document provides an introduction to metabolomics, including:
1. An overview of the techniques used to measure the metabolome, such as quenching metabolism, solvent extractions, HPLC, GC-MS, and NMR.
2. Descriptions of the complexity of the metabolome due to the wide range of molecular weights and concentrations of metabolites.
3. Explanations of applications of metabolomics including biomarker discovery, nutrigenomics, metabolic engineering, and plant functional genomics.
Pl ql1-mainchem0414
The document summarizes a scheme for distributing monthly liquid samples for external quality assessment. It includes mainline chemistry analytes distributed in 3.5ml and 1ml aliquots. The material is human serum manufactured to high specifications and sterilized to maintain integrity and commutability. Reference target values are assigned for key analytes to assess traceability. Performance reports identify components of imprecision and inaccuracy, and provide group reporting in graphical and tabular form to manage performance across sites. The scheme also includes samples for assessing linearity, validating new equipment and methods, and challenging laboratories.
1501 Refocus metabolomics
Metabolomics is often described as the study of “the complete set of low molecular weight intermediates, which are context dependent, varying according to the physiology, developmental or pathological state of the cell, tissue, organ or organism”. In fact, metabolomics is a new term for an old science in which classical biochemical concepts are investigated. New and unique to the current research that is being conducted is the combination with genomics information and full system biology. In this refocus we will discuss the challenges in today's metabolomics research and how to address them
Microbial taxonomy
This document discusses various methods used in microbial taxonomy, including chemotaxonomy and molecular methods. Chemotaxonomy involves analyzing chemical components of microbial cells like cell walls, lipids, fatty acids, isoprenoid quinones, and cytochromes. Molecular methods discussed include DNA homology, which involves measuring DNA reassociation between organisms to determine genetic relatedness, RNA sequencing to identify organisms, and analyzing G+C content. Overall, the document provides an overview of various chemical and molecular characterization techniques used to classify and identify microorganisms.
Host Cell Protein Analysis by Mass Spectrometry | KBI Biopharma
Host Cell Protein Analysis by Mass Spectrometry. Originally presented at the 2018 Sciex Users Meeting by Michael J Nold, Ph.D., Mass Spectrometry Core Facility at KBI Biopharma.
Data drivenapproach to medicinalchemistry
Learn how large-scale normalized data empowers the critical early phases of drug discovery.
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Characterizing the Complex Lipidic Composition of Small Extracellular Vesicle...
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lehninger(sixth edition) Ch 03: Amino acids, peptides and proteins
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The modern software delivery process (or the CI/CD process) includes many tools, distributed teams, open-source code, and cloud platforms. Constant focus on speed to release software to market, along with the traditional slow and manual security checks has caused gaps in continuous security as an important piece in the software supply chain. Today organizations feel more susceptible to external and internal cyber threats due to the vast attack surface in their applications supply chain and the lack of end-to-end governance and risk management.
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Bob Boule
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“Building and Scaling AI Applications with the Nx AI Manager,” a Presentation...
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Robin van Emden, Senior Director of Data Science at Network Optix, presents the “Building and Scaling AI Applications with the Nx AI Manager,” tutorial at the May 2024 Embedded Vision Summit.
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Monitoring and observability aren’t traditionally found in software curriculums and many of us cobble this knowledge together from whatever vendor or ecosystem we were first introduced to and whatever is a part of your current company’s observability stack.
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I, a former op, would like to extend an invitation to all application developers to join the observability party will share these foundational concepts to build on:
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Quantitative proteomics
- 2. Introduction of SILAC and its applications • Stable Isotope Labeling of Amino acids in Culture • Develop and promoted by Matthias Mann • Two papers: – Lipid Rafts (PNAS) – Focal Adhesion precursors (Cell under review)
- 3. What does a mass spectrometer do? Precise identification of mass Can trap a single ion species and fragment to get sub fragments (sequence)
- 4. Overview of Biological and Chemical Isotope Labeling Strategies Two complementary samples Labeling Analysis
- 5. SILAC vs. ICAT • Culture system only • De Novo Proteins (no serum contamination) • No optimization • Simplifies MS/MS • More complete peptide coverage • ALL protein samples • Labels selected moieties • Need to optimize labeling • Large linker group. • Reduces complexity
- 6. Stable isotopic amino acids (MW) Cambridge Isotope Catalog Sigma-Aldich Catalog L-Lysine 2HCl (223.13) (4,4,5,5-D4) DLM-2640 - L-Lysine HCl (186.67) (4,4,5,5-D4) - 616192 L-Lysine 2HCl (225.1) (13 C6) CLM-2247 - L-Lysine HCl (188.6) (13 C6) - 643459 L-Lysine 2HCl (227.1) (13 C6, 15 N2) CNLM-291 - L-Lysine HCl (190.59) (13 C6, 15 N2) - 608041 L-Arginine HCl (214.64) (15 N4) NLM-396 600113 L-Arginine HCl (216.62) (13 C6) CLM-2265 643440 L-Arginine HCl (220.59) (13 C6, 15 N4) CNLM-539 608033 L-Arginine HCl (221.6) (15 N4, D7) DNLM-7543 - L-Arginine HCl (227.6) (13 C6, 15 N4, D7) CDNLM-6801 SILAC Labels
- 7. “Light” “Heavy” “Light/heavy” mix How can you distinguish between isotoped species?
- 8. How do you actually get quantitation? 1) Once you’ve identified the complementary peptides, 2) Backtrack and integrate the Xtracted Ion Chromatogram
- 9. I-TRAQ (Isobaric Tag for Relative and Absolute Quantitation
- 10. Experimental Strategy Sample preparation Labeling Chromatographic separations Mass Spectrometry Analysis