Protein Sensors of Reactive Oxygen Species, Part A: Selenoproteins and Thioredoxin
- 1st Edition, Volume 347 - March 1, 2002
- Editors: Lester Packer, Helmut Sies
- Language: English
- Hardback ISBN:9 7 8 - 0 - 1 2 - 1 8 2 2 4 8 - 4
- eBook ISBN:9 7 8 - 0 - 0 8 - 0 4 9 6 9 5 - 5
This volume of Methods in Enzymology is concerned with the rapidly developing field of selenoprotein synthesis and its related molecular genetics. Progressive information on the to… Read more
This volume of Methods in Enzymology is concerned with the rapidly developing field of selenoprotein synthesis and its related molecular genetics. Progressive information on the topics of proteins as redox sensors, selenoproteins, and the thioredoxin system is studied using methods such as bioinformatics, DNA chip technology, cell biology, molecular genetics, and enzymology. The information on novel selenoproteins identified from genomic sequence data, as well as current knowledge on glutathione peroxidases, selenoprotein P, iodothyronine deiodinases, and thioredoxin reductases, is presented in a method-based approach.
Biochemists, pharmacologists, physiologists, cell biologists, molecular biologists, and biomedical researchers.
- Contributors to volume 347
- Preface
- Methods in enzymology
- Section I: Selenoproteins
- [1]: Selenoprotein Biosynthesis: Purification and Assay of Components Involved in Selenocysteine Biosynthesis and Insertion in Escherichia coli
- Introduction
- Purification of Selenocysteine-Inserting tRNASec
- Purification of Selenophosphate Synthetase
- Purification of Selenocysteine Synthase
- Preparation of Elongation Factor SelB
- Serylation of tRNASec
- Conversion of Seryl-tRNASec to Selenocysteyl-tRNASec
- Preparation of Selenocysteine Insertion Sequence Elements
- Analysis of Interaction of SelB with Its RNA Ligands
- [2]: Selenocysteine Insertion Sequence Element Characterization and Selenoprotein Expression
- Abstract
- Criteria for Identifying Functional Selenocysteine Insertion Sequence Elements in Databases
- Methods for Studying Selenocysteine Insertion Sequence Function and Selenoprotein Synthesis
- Selenocysteine Insertion Sequence Activity Assays
- Optimizing Expression in Transiently Transfected Cells
- Incorporation of 75Se into Selenoproteins in Transiently Transfected Cells
- Acknowledgment
- [3]: Transfer RNAs That Insert Selenocysteine
- Introduction
- Primary Structures
- Selenocysteine-Inserting tRNA Gene, Gene Expression, and Gene Product
- Selenocysteine-Inserting tRNA: Carrier Molecule for Biosynthesis of Its Amino Acid
- Selenocysteine-Inserting tRNA: Donor of Selenocysteine to Protein
- Functional Consequences of Over- and Underexpression of Mammalian Selenocysteine-Inserting tRNA and Expression of an i 6 A-Deficient Selenocysteine-Inserting tRNA
- Purification of Selenocysteine-Inserting tRNA
- Preparation of [75Se]Selenocysteyl-tRNA
- [4]: Purification and Analysis of Selenocysteine Insertion Sequence-Binding Protein 2
- Introduction
- Purification of Native Selenocysteine Insertion Sequence-Binding Protein 2
- Purification of Recombinant Selenocysteine Insertion Sequence-Binding Protein 2
- RNA-Binding Assays for Selenocysteine Insertion Sequence-Binding Protein 2
- In Vitro Translation of Phospholipid Hydroperoxide Glutathione Peroxidase
- Ribosome-Binding Assay
- Summary and Perspectives
- Acknowledgments
- [5]: Nonsense-Mediated Decay: Assaying for Effects on Selenoprotein mRNAs
- Introduction
- Rationale for Analyzing Transiently Expressed, in Vitro-Modified Glutathione Peroxidase 1 Alleles in Cultured Cells
- Methods Used to Determine the Mechanism by Which Selenium Concentration Affects Glutathione Peroxidase 1 Gene Expression
- Summary
- Acknowledgment
- [6]: Novel Selenoproteins Identified from Genomic Sequence Data
- Introduction
- Computational Detection of Selenocysteine Insertion Sequence Motifs
- Functional Screen of Selected Selenocysteine Insertion Sequence Hits
- Obtaining and Analyzing Selenoprotein cDNA Sequences
- Further Characterization of Selenoproteins
- Conclusion
- Acknowledgments
- [7]: Semisynthesis of Proteins Containing Selenocysteine
- Introduction
- Strategies for Synthesis of Proteins Containing Selenocysteine
- Preparation of Selenocysteine for Peptide Synthesis
- Solid-Phase Synthesis of Peptide Containing Selenocysteine
- Semisynthesis of a Protein Containing Selenocysteine
- Characterization of Proteins Containing Selenocysteine
- Use of Selenocysteine as Cleavage Reagent
- Summary
- Acknowledgments
- [8]: Mammalian Selenoprotein Gene Signature: Identification and Functional Analysis of Selenoprotein Genes Using Bioinformatics Methods
- Introduction
- Eukaryotic Selenoproteins
- Selenocysteine/Cysteine Pair in Homologous Sequences
- Mammalian Selenocysteine Insertion Sequence Element
- Identification of Selenocysteine Insertion Sequence Elements
- Mammalian Selenoprotein Gene Signature
- Applicability of Mammalian Selenoprotein Gene Signature Criteria
- Other Approaches for Identification of Selenoprotein Genes
- Strategy to Search for Selenoproteins in Nucleotide Sequence Databases
- Strategy to Test Whether a Newly Isolated Gene Encodes a Selenoprotein
- Functional Analyses of New Selenoproteins
- Concluding Remarks
- Acknowledgment
- [9]: Estimation of Individual Types of Glutathione Peroxidases
- Introduction
- Determination by Activity
- Determination by RNA Analysis
- Determination by Immunochemical Methods
- Immunochemical Detection
- Conclusions
- Acknowledgments
- [10]: High-Throughput 96-Well Microplate Assays for Determining Specific Activities of Glutathione Peroxidase and Thioredoxin Reductase
- Introduction
- Methods
- [11]: Selenoprotein P
- Introduction
- Structure
- Properties
- Proposed Functions
- Acknowledgments
- [12]: Iodothyronine Deiodinases
- Introduction
- Historical Reminiscence
- Type I 5′-Deiodinase
- Type II 5′-Deiodinase Activity
- Type III Iodothyronine Deiodinase
- Methods to Determine Deiodinase Activity
- Acknowledgments
- [13]: Expression and Regulation of Thioredoxin Reductases and Other Selenoproteins in Bone
- Bone Physiology
- Expression of Selenoproteins in Cells of Bone Microenvironment
- Expression and Regulation of Glutathione Peroxidases in Osteoblasts
- Expression and Regulation of Thioredoxin Reductases in Osteoblasts
- Expression of Glutathione Peroxidases and Thioredoxin Reductase α in Monocyte-Derived Cells
- Discussion
- Summary
- [14]: Selenoprotein W
- Abstract
- Procedures
- [15]: Genetic and Functional Analysis of Mammalian Sep15 Selenoprotein
- Introduction
- Materials and Methods
- Conclusions
- Acknowledgment
- [16]: Selenocysteine Lyase from Mouse Liver
- Introduction
- Cloning of cDNA for Mouse Selenocysteine Lyase
- Expression of Selenocysteine Lyase in Escherichia coli
- Assay Method
- Purification Procedure
- Properties
- Tissue Distribution and Intracellular Localization
- [17]: Selenocysteine Methyltransferase
- Introduction
- Enzyme Assays
- Semiquantitative Assay
- Quantitative Assay
- Crude Extract and Ammonium Sulfate Precipitation
- Column Chromatography
- Storage and Stability
- Properties
- Relation to Homocysteine Methyltransferases
- [18]: Phospholipid–Hydroperoxide Glutathione Peroxidase in Sperm
- Introduction
- Measurement of Phospholipid-Hydroperoxide Glutathione Peroxidase in Human Spermatozoa
- Discussion
- Acknowledgments
- [19]: In Vivo Antioxidant Role of Glutathione Peroxidase: Evidence from Knockout Mice
- Introduction
- Development and Characterization of GPX1(+) and GPX1(−/−) Mouse Models
- Protection by Glutathione Peroxidase against Acute, Lethal Oxidative Stress
- Protection by Glutathione Peroxidase against Moderate and Metabolic Oxidative Stress
- Unanswered Questions
- Acknowledgment
- [20]: Recombinant Expression of Mammalian Selenocysteine-Containing Thioredoxin Reductase and Other Selenoproteins in Escherichia coli
- Introduction
- tRNASec Defining the Selenoprotein World
- Species Barriers in Heterologous Selenoprotein Synthesis
- Introduction of Selenocysteine Insertion Sequence Element Compatible with Bacterial Selenoprotein Synthesis Machinery as Method for Recombinant Selenoprotein Production
- Critical Factors for Recombinant Selenoprotein Production in Escherichia coli
- Conclusions
- [21]: Mammalian Thioredoxln Reductases as Hydroperoxide Reductases
- Introduction
- Purification of Mammalian Thioredoxin Reductase from Wet Tissue
- Assay of Mammalian Thioredoxin Reductase Activity
- Assay of Hydroperoxide Reductase Activity
- Discussion
- Comments
- Acknowledgments
- [22]: Tryparedoxin and Tryparedoxin Peroxidase
- Introduction
- Tryparedoxins
- Tryparedoxin Peroxidases
- Sources of Purified Tryparedoxin and Tryparedoxin Peroxidase
- Conclusions
- Acknowledgments
- [23]: Trypanothione and Tryparedoxin in Ribonucleotide Reduction
- Introduction
- Methods
- Conclusions
- Acknowledgment
- [24]: Selenium- and Vitamin E-Dependent Gene Expression in Rats: Analysis of Differentially Expressed mRNAs
- Introduction
- Production of Selenium and Vitamin E Deficiency in Rats
- Analysis of Differentially Expressed mRNAs by Atlas cDNA Expression Arrays
- Typical Results
- [1]: Selenoprotein Biosynthesis: Purification and Assay of Components Involved in Selenocysteine Biosynthesis and Insertion in Escherichia coli
- Section II: Thioredoxin
- [25]: Overview
- Introduction
- Thioredoxin and Related Molecules
- Thioredoxin-Binding Proteins
- Redox Regulation of Transcriptional Factors by Thioredoxin
- Cytoprotective Action of Thioredoxin
- Environmental Stressors and Redox Signal
- Concluding Remarks and Perspectives
- Acknowledgments
- [26]: Thioredoxin and Glutaredoxin Isoforms
- Introduction
- Thioredoxins
- Escherichia coli Thioredoxins
- Saccharomyces cerevisiae Thioredoxins
- Human Thioredoxins
- Structural Similarities
- Assays for Thioredoxin Activity
- Glutaredoxins
- Catalytic Mechanism of Glutaredoxin/Thioredoxin
- Escherichia coli Glutaredoxins
- Saccharomyces cerevisiae Glutaredoxins
- Human Glutaredoxin
- Classification of Glutaredoxins
- Assays for Glutaredoxin Activity
- Acknowledgments
- [27]: Mammalian Thioredoxin Reductases
- Introduction
- Labeling Proteins with Selenium-75
- Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis Analysis and Autoradiography
- Identification of Selenocysteine Residues
- Precautions for Purifying Selenocysteine-Containing Proteins
- Separation of Thioredoxin Reductase 1 from Thioredoxin Reductase 2 in Rat Liver Homogenates
- Selective Alkylation of Selenocysteine Residue 498 of Thioredoxin Reductase 1
- Catalytic Role of Selenocysteine Residue
- [28]: Mitochondrial Thioredoxin Reductase and Thiol Status
- Introduction
- Determination of Total Mitochondrial Thiols
- Determination of Total Protein Thiols, Membrane Thiols, and Acid-Soluble Thiols
- Determination of Available Protein Thiols
- Mitochondrial 5,5′-Dithiobis(2-Nitrobenzoic Acid) Reductase Activity
- Purification of Mitochondrial Thioredoxin Reductase
- Preparation and Processing of Mitochondria
- DEAE-Sephacel Chromatography
- Assays
- Acknowledgment
- [29]: Protein Electrophoretic Mobility Shift Assay to Monitor Redox State of Thioredoxin in Cells
- Introduction
- Methods
- Results
- Acknowledgments
- [30]: Recycling of Vitamin C by Mammalian Thioredoxin Reductase
- Introduction
- Methods
- Conclusion
- [31]: Thioredoxin Cytokine Action
- Introduction
- Truncated Form of Thioredoxin
- Thioredoxin as Costimulatory Molecule of Cytokine Action
- Chemokine-Like Activity of Thioredoxin
- Circulating Thioredoxin Levels in Human Plasma
- Sandwich Enzyme-Linked Immunosorbent Assay for Human Thioredoxin
- Concluding Remarks
- Acknowledgment
- [32]: Identification of Thioredoxin-Linked Proteins by Fluorescence Labeling Combined with Isoelectric Focusing/Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis
- Introduction
- Sources of Materials and Chemicals
- Methods
- Applications
- [33]: Thioredoxin and Mechanism of Inflammatory Response
- Introduction
- NF-κB Immunostaining with Synovial Fibroblast Cultures
- Electrophoretic Mobility Shift Assay
- [34]: Redox State of Cytoplasmic Thioredoxin
- Introduction
- Determination of Redox State in Vivo
- Thioredoxin as Disulfide Bond Reductase
- Thioredoxin as Thiol Oxidant
- Acknowledgments
- [35]: Thioredoxin, Thioredoxin Reductase, and Thioredoxin Peroxidase of Malaria Parasite Plasmodium falciparum
- Introduction
- Expression and Purification of Recombinant Thioredoxin Reductase, Thioredoxin 1, and Thioredoxin Peroxidase 1
- Assays for Reactions and Compounds of Thioredoxin System
- Concluding Remarks
- Acknowledgments
- [36]: Human Placenta Thioredoxin Reductase: Preparation and Inhibitor Studies
- Introduction
- Assay Procedures
- [37]: Classification of Plant Thioredoxins by Sequence Similarity and Intron Position
- Arabidopsis Genes Encoding Thioredoxins or Proteins with Thioredoxin Domains
- Usability of Arabidopsis Data for Predicting Type of Plant Sequence
- [38]: Ferredoxin-Dependent Thioredoxin Reductase: A Unique Iron–Sulfur Protein
- Introduction
- Ferredoxin/Thioredoxin Reductase
- Spinach Thioredoxin f
- Acknowledgment
- [39]: Plant Thioredoxin Gene Expression: Control by Light, Circadian Clock, and Heavy Metals
- Introduction
- Experimental Procedures
- [40]: Thioredoxin Genes in Lens: Regulation by Oxidative Stress
- Introduction
- Experimental Procedures
- mRNAs Encoding Thioredoxin Genes in Lens
- Western Blot Analysis of Thioredoxin
- Thioredoxin Reductase Activity in Human Lens
- Concluding Remarks
- Acknowledgments
- [41]: Thioredoxin Overexpression in Transgenic Mice
- Introduction
- Thioredoxin Knockout Mice
- Characteristics of Thioredoxin-Transgenic Mice
- Concluding Remarks
- Acknowledgment
- [42]: Multiplex Reverse Transcription-Polymerase Chain Reaction for Determining Transcriptional Regulation of Thioredoxin and Glutaredoxin Pathways
- Introduction
- RNA
- cDNA
- Primers
- Multiplex Polymerase Chain Reaction
- Optimization of Multiplex Reverse Transcription-Polymerase Chain Reaction
- Application
- Final Remarks
- Acknowledgments
- [43]: Redox Regulation of Cell Signaling by Thioredoxin Reductases
- Introduction
- Isolation of Mammalian Thioredoxin Reductase Isozymes
- Analysis of Thioredoxin Reductase 1 Redox State, Using 5-Iodoacetamidofluorescein
- Analysis of Changes in Redox State of Isolated Mouse Liver Thioredoxin Reductase 1 in Vitro
- Analysis of Thioredoxin Reductase 1 Redox State in Cell Culture System
- Specific Alkylation of Selenocysteine in Thioredoxin Reductase 1 by 5-Iodoacetamidofluorescein
- Effect of Stimulation of A431 Cells with Epidermal Growth Factor or H2O2 on Expression of Thioredoxin Reductase 1
- Concluding Remarks
- [25]: Overview
- Author index
- Subject index
- Edition: 1
- Volume: 347
- Published: March 1, 2002
- Language: English
LP
Lester Packer
Lester Packer received a PhD in Microbiology and Biochemistry in 1956 from Yale University. In 1961, he joined the University of California at Berkeley serving as Professor of Cell and Molecular Biology until 2000, and then was appointed Adjunct Professor, Pharmacology and Pharmaceutical Sciences, School of Pharmacy at the University of Southern California.
Dr Packer received numerous distinctions including three honorary doctoral degrees, several distinguished Professor appointments. He was awarded Chevalier de l’Ordre National du Merite (Knight of the French National Order of Merit) and later promoted to the rank of Officier. He served as President of the Society for Free Radical Research International (SFRRI), founder and Honorary President of the Oxygen Club of California.
He has edited numerous books and published research; some of the most cited articles have become classics in the field of free radical biology:
Dr Packer is a member of many professional societies and editorial boards. His research elucidated - the Antioxidant Network concept. Exogenous lipoic acid was discovered to be one of the most potent natural antioxidants and placed as the ultimate reductant or in the pecking order of the “Antioxidant Network” regenerating vitamins C and E and stimulating glutathione synthesis, thereby improving the overall cellular antioxidant defense. The Antioxidant Network is a concept addressing the cell’s redox status. He established a world-wide network of research programs by supporting and co-organizing conferences on free radical research and redox biology in Asia, Europe, and America.
Affiliations and expertise
Department of Molecular Pharmacology and Toxicology, School of Pharmaceutical Sciences, University of Southern California, USAHS
Helmut Sies
Helmut Sies, MD, PhD (hon), studied medicine at the universities of Tübingen, Munich, and Paris. He was the professor and chair of the Institute for Biochemistry and Molecular Biology I at Heinrich-Heine-University Düsseldorf, Germany, where he is now professor emeritus. He is a member of the German National Academy of Sciences Leopoldina and was the president of the North Rhine-Westphalian Academy of Sciences and Arts. He was named ‘Redox Pioneer’; was the president of the Society for Free Radical Research International (SFRRI). Helmut Sies introduced the concept of “Oxidative Stress” in 1985, and was the first to reveal hydrogen peroxide as a normal constituent of aerobic cell metabolism. His research interests comprise redox biology, oxidants, antioxidants, micronutrients.
Affiliations and expertise
Heinrich-Heine-University Düsseldorf, GermanyRead Protein Sensors of Reactive Oxygen Species, Part A: Selenoproteins and Thioredoxin on ScienceDirect