Protein Synthesis: Methods and Protocols

ISBN-13: 9781617370250 / Angielski / Miękka / 2010 / 442 str.

Robin Martin

Opis

The synthesis of proteins from 20 or so constituent amino acids according to a strictly defined code with an accuracy of better than 1 in 10,000 at most loca tions is arguably the most complex task performed by cells. Protein Synthesis collects together methods and protocols covering a range of different approaches towards understanding how the cellular machinery accomplishes this task and how these ftinctions might be harnessed by the biotechnology industry to generate novel and useful proteins. The era in which the components of the translational machinery were being catalogued is over. This volume gathers together protocols that focus on preserving and describing the dynamic function as closely as possible. The need to understand exactly how ribosomes are positioned on messages or where tRNA molecules, translation factors, or control proteins are bound, has been appreciated by many of the authors. Several chapters that explore the fidelity and processivity of translation reflect this belief. Moreover, the fundamental importance of rRNA at the heart of the ribosome is a strong theme in a number of the protocols. These articles include in vitro and in vivo systems from bacterial, fungal, plant, and animal systems. Overall, Protein Synthesis might be characterized by the novelty of the approaches employed to illuminate the inner workings of the protein synthetic machinery as well as by the inventiveness of the attempts to harness these reactions for biotechnological applications."

Recenzje

". . . an excellent protocols book, a copy of which should be on the shelves of every molecular biology lab."-Microbiology Today

Spis treści

Analyses of Ribosome Distribution During In Vitro Translation, Sandra L. Wolin. Allele-Specific Priming in the Mapping of rRNA, Stephen Douthwaite. Permeabilized Mammalian Cells as a System for Protein Synthesis, Romualdas Stapulionis and Murray P. Deutscher. The Measurement of Processivity Errors in Protein Synthesis, Hengjiang Dong. A Biosynthetic Approach for the Incorporation of Unnatural Amino Acids into Proteins, Jon S. Thorson, Virginia W. Cornish, Jeannie E. Barrett, Sharon T. Cload, Takato Yano, and Peter G. Schultz. The Analysis of Translational Activity Using a Reporter Gene Constructed from Repeats of an Antibody-Binding Domain from Protein A, Asgeir Björnsson, Salim Mottagui-Tabar, and Leif A. Isaksson. In Vitro Engineering Using Acyl-Derivatized tRNAs, Obed W. Odom, Wieslaw Kudlicki, and Boyd Hardesty. In Vitro Engineering Using Synthetic tRNAs with Altered Anticodons Including Four-Nucleotide Antocodons, Gisela Kramer, Wieslaw Kudlicki, and Boyd Hardesty. Analysis of rRNA Function Using Specialized Ribosomes, Marcel F. Brink and Herman A. de Boer. Determination of the Peptide Elongation Rate In Vivo, Michael A. Sørensen and Steen Pedersen. The Measurement of Missense Errors During In Vivo Protein Synthesis, Bernard N. Violand and Gregg Bogosian. Heelprinting Analysis of In Vivo Ribosome Pause Sites, Margaret J. Hollingsworth, Jeong-Kook Kim, and Neil E. Stollar. Analysis of eIF-2a Kinases in Yeast, Thomas E. Dever. Continuous-Flow Cell-Free Translation, Transcription-Translation, and Replication-Translation Systems, Lyubov A. Ryabova, Igor Yu. Morozov, and Alexander S. Spirin. Analysis of Translational Activity of Extracts Derived from Oocytes and Eggs of Xenopus laevis, Virginia M. Pain, Tina D. Patrick, Richard Cox, and Simon J. Morley. A Fractionated Reticulocyte Lysate System for Studies on Protein Synthesis Initiation Factors, Michael Rau, Theophile Ohlmann,Virginia M. Pain, and Simon J. Morley. Measurement of Ribosomal Accuracy and Proofreading in E. coli Burst Systems, Måns Ehrenberg and Ne¸'e Bilgin. Measurement of Rate of Protein Synthesis In Vitro Preparation of Escherichia coli Burst Systems, Ne¸'e Bilgin and Måns Ehrenberg. In Vivo Mutational Analysis of Ribosomal RNA in Saccharomyces cerevisiae, Jaap Venema, Rudi J. Planta, and Hendrik A. Raué. Genetic Selection of rRNA Mutations, Steven T. Gregory, Carleen A. Brunelli, J. Stephen Lodmell, Michael O'Connor, and Albert E. Dahlberg. Toeprinting Assays: Mapping by Blocks to Reverse Transcriptase Primer Extension, Steven Ringquist and Larry Gold. Mapping of Pseudouridine Residues in RNA to Nucleotide Resolution, Andrey V. Bakin and James Ofengand. Chemical and Enzymatic Probing of Antibiotic-Ribosome Complexes, Cristina Rodriguez-Fonseca and Roger A. Garrett. Protein Engineering with Nonstandard Amino Acids. Lance E. Steward and A. Richard Chamberlin. Internal Ribosome Entry Sites Tests with Circular mRNAs, Chang-you Chen and Peter Sarnow. Gel Retardation and UV-Crosslinking Assays to Detect Specific RNA-Protein Interactions in the 5' or 3' UTRs of Translationally Regulated mRNAs, James Walker, Osvaldo de Melo Neto, and Nancy Standart. Translational Control by Repressor Proteins Binding to the 5' UTR of mRNAs, Nicola K. Gray. Identification and Analysis of Frameshift Sites, Arunachalam Vimaladithan and Philip J. Farabaugh. Synthesis and Site-Specific Binding of Thioated tRNAs to Probe Ribosome-tRNA Interactions, Marylena Dabrowski and Knud H. Nierhaus. Index.