SUMO Protocols

ISBN-13: 9781934115800 / Angielski / Twarda / 2008 / 324 str.

Helle D. Ulrich

Opis

Post-translational protein modifications by members of the ubiquitin family are widely recognized as important regulatory control systems for a variety of biological pathways. Their influence on eukaryotic cellular metabolism is comparable to that of other modifi- tions such as phosphorylation, acetylation and methylation. The small ubiquitin-related modifier SUMO uses a conjugation and de-conjugation system closely related to that of ubiquitin itself; yet, the functions of the SUMO system are highly diverse and largely independent of the ubiquitin system. SUMO modification controls the activity of tr- scription factors and can influence protein stability, but it also contributes to nucleo-cy- plasmic transport, chromosome segregation and DNA damage repair. As a consequence, the SUMO system pervades virtually all areas of basic molecular and cell biology, and scientists from different backgrounds, including medical researchers, are likely to enco- ter SUMO in the course of their studies. This volume, SUMO Protocols, therefore aims at presenting a collection of methods relevant to SUMO research in order to make these tools available to biochemists, molecular and cell biologists as well as research-oriented clinicians not yet familiar with the system. In contrast to the ubiquitin system, which has been the subject of several reviews and methods collections, no practical compendium entirely devoted to SUMO has been p- lished.

Recenzje

From the reviews:

"The modification of proteins by small ubiquitin-like modification (SUMO) is relevant to virtually all areas of molecular and cell biology. SUMO Protocols is a quite comprehensive volume describing various in vivo and in vitro techniques ... . The target audience is likely to be postdoctoral fellows and PhD students in laboratories not only dedicated to SUMO pathways but those beginning their initial forays in the field ... and regulated by SUMO. ... for those pursuing SUMO-specific modification this is a worthwhile addition to their bookshelves." (Peter O'Hare, Microbiology Today, July, 2009)

Spis treści

Part I. Introduction 1 The SUMO System: An Overview Helle D. Ulrich Part II. Identification of SUMO Targets 2 Identification of SUMO Target Proteins by Quantitative Proteomics Jens S. Andersen, Ivan Matic and Alfred C. O. Vertegaal 3 Identification of SUMO-Conjugated Proteins and their SUMO Attachment Sites Using Proteomic Mass Spectrometry James A. Wohlschlegel 4 Identification of SUMO Targets through In Vitro Expression Cloning Christian B. Gocke and Hongtao Yu 5 Enhanced Detection of In Vivo SUMO Conjugation by Ubc9 Fusion-Dependent Sumoylation (UFDS) Rainer Niedenthal 6 In Vivo Detection and Characterization of Sumoylation Targets in Saccharomyces cerevisiae Helle D. Ulrich and Adelina A. Davies Part III. Identification of SUMO Interactors 7 Identification of SUMO-Interacting Proteins by Yeast Two-Hybrid Analysis Mary B. Kroetz and Mark Hochstrasser 8 Identification of SUMO Binding Motifs by NMR Candace S. Seu and Yuan Chen Part IV. Functional Analysis of Sumoylation In Vivo 9 Regulation of Transcription Factor Activity by SUMO Modification Jian Ouyang, Alvaro Valin and Grace Gill 10 Characterization of the Effects and Functions of Sumoylation Through Rapamycin-Mediated Heterodimerization Shanshan Zhu and Michael J. Matunis Part V. Biochemical Studies of SUMO Conjugation 11 Purification of SUMO Conjugating Enzymes and Kinetic Analysis of Substrate Conjugation Ali A. Yunus and Christopher D. Lima 12 Performing In Vitro Sumoylation Reactions Using Recombinant Enzymes Andreas Werner, Marie-Christine Moutty, Ulrike Möller and Frauke Melchior 13 Preparation of Sumoylated Substrates for Biochemical Analysis Puck Knipscheer, Helene Klug, Titia K.Sixma and Andrea Pichler 14 Strategies for the Expression of SUMO-Modified Target Proteins in Escherichia coli Hisato Saitoh, Junsuke Uwada and Azusa Kawasaki Part VI. Biochemical Studies of SUMO Deconjugation 15 Preparation of SUMO Proteases and Kinetic Analysis Using Endogenous Substrates David Reverter and Christopher D. Lima 16 An In Vitro FRET-Based Assay for the Analysis of SUMO Conjugation and Isopeptidase Cleavage Nicolas Stankovic-Valentin, Lukasz Kozaczkiewicz, Katja Curth and Frauke Melchior 17 FRET-Based In Vitro Assays for the Analysis of SUMO Protease Activities Michael H. Tatham and Ronald T. Hay 18 Detection and Characterization of SUMO Protease Activity Using a Sensitive Enzyme-Based Reporter Assay Craig A. Leach, Xufan Tian, Michael R. Mattern and Benjamin Nicholson Part VII. Manipulation and Alternative Uses of the SUMO System 19 Inhibition of the SUMO Pathway by Gam1 Mariaelena Pozzebon, Chiara V. Segre and Susanna Chiocca 20 SUMO Fusion Technology for Enhanced Protein Production in Prokaryotic and Eukaryotic Expression Systems Tadas Panavas, Carsten Sanders and Tauseef R. Butt

Blurb

While the small ubiquitin-related modifier SUMO uses a conjugation and de-conjugation system closely related to that of ubiquitin itself, its functions are highly diverse and largely independent of the ubiquitin system. In SUMO Protocols, leading experts present a collection of methods intended to highlight important features unique to the biology of SUMO in order to support future SUMO research. The first half of the volume focuses on the identification and functional characterization of SUMO targets and interactors, while the second half covers essential biochemical methods for the manipulation of the enzymatic components of the SUMO system. Written in the highly successful Methods in Molecular Biology™ series format, the chapters include brief introductions to the subject, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and the Notes sections, which provide tips on troubleshooting and avoiding known pitfalls.

Unique and cutting-edge, SUMO Protocols provides the necessary tools for biochemists, molecular and cellular biologists as well as research-oriented clinicians not yet familiar with this vital system.