Single-Molecule Enzymology: Fluorescence-Based and High-Throughput Methods

Academic Press, Oct 28, 2016 - Science - 616 pages

Single-Molecule Enzymology, Part A, the latest volume in the Methods in Enzymology series, continues the legacy of this premier serial with quality chapters authored by leaders in the field. This volume covers research methods in single-molecule enzymology, and includes sections on such topics as force-based and hybrid approaches, fluorescence, high-throughput sm enzymology, nanopores, and tethered particle motion.

• Continues the legacy of this premier serial with quality chapters authored by leaders in the field
• Covers research methods in single-molecule enzymology
• Contains sections on such topics as force-based and hybrid approaches, fluorescence, high-throughput sm enzymology, nanopores, and tethered particle motion

Contents

• SingleMolecule Imaging With One Color Fluorescence: 33
• Measuring Membrane Protein Dimerization Equilibrium in Lipid Bilayers by SingleMolecule Fluorescence Micr: 53
• Fluorescent Labeling of Proteins in Whole Cell Extracts for SingleMolecule Imaging: 83
• Quantifying the Assembly of Multicomponent Molecular Machines by SingleMolecule Total Internal Reflection: 105
• How to Measure Separations and Angles Between Intramolecular Fluorescent Markers: 147
• Precisely and Accurately Inferring SingleMolecule Rate Constants: 187
• Quantification of Functional Dynamics of Membrane Proteins Reconstituted in Nanodiscs Membranes by Single: 227
• Putting HumptyDumpty Together Clustering the Functional Dynamics of Single Biomolecular Machines Such as: 257
• SingleMolecule FRET to Measure Conformational Dynamics of DNA Mismatch Repair Proteins: 285
• SingleMolecule Confocal FRET Microscopy to Dissect Conformational Changes in the Catalytic Cycle of DNA: 317
• Probing the Conformational Landscape of DNA Polymerases Using DiffusionBased SingleMolecule FRET: 353
• Methods for Investigating DNA Accessibility with Single Nucleosomes: 379
• SingleMolecule Fluorescence Studies of Fast Protein Folding: 417
• SingleMolecule Multicolor FRET Assay for Studying Structural Dynamics of Biomolecules: 461
• A Multicolor SingleMolecule FRET Approach to Study Protein Dynamics and Interactions Simultaneously: 487
• Interferometric Scattering Microscopy for the Study of Molecular Motors: 517
• Enzyme Kinetics in Femtoliter Arrays: 541
• Author Index: 561
• Subject Index: 587
• Back Cover: 599

About the author

Graduate of Peter the Great St. Petersburg Polytechnic University, Russia (1996 MS diploma with honors (equivalent of cum laude) in physics/biophysics) and Osaka University, Japan (2000 PhD in biological sciences), Dr. Maria Spies is an Associate Professor of Biochemistry at the University of Iowa Carver College of Medicine. Spies’ research career has been focused on deciphering the intricate choreography of the molecular machines orchestrating the central steps in the homology directed DNA repair. Her doctoral research supported by the Japanese Government (MONBUSHO) Graduate Scholarship provided the first detailed biochemical characterization of archaeal recombinase RadA. In her postdoctoral work with Dr. Steve Kowalczykowski (UC Davis) supported by the American Cancer Society, Spies reconstituted at the single-molecule level the initial steps of bacterial recombination and helped to explain how this process is regulated. Spies’ laboratory at the University of Iowa emphasizes the molecular machinery of homologous recombination, how it is integrated into DNA replication, repair and recombination (the 3Rs of genome stability), and how it is misappropriated in the molecular pathways that process stalled DNA replication events and DNA breaks through highly mutagenic, genome destabilizing mechanisms. Her goal is to understand, reconstitute and manipulate an elaborate network of DNA recombination, replication and repair, and to harness this understanding for anticancer drug discovery. The Spies lab utilizes a broad spectrum of techniques from biochemical reconstitutions of the key biochemical reactions in DNA recombination, repair and replication, to structural and single-molecule analyses of the proteins and enzymes coordinating these reactions, to combined HTS/CADD campaigns targeting human DNA repair proteins. Work in Spies Lab has been funded by the American Cancer Society (ACS), Howard Hughes Medical Institute (HHMI), and is currently supported by the National Institutes of Health (NIH). She received several prestigious awards including HHMI Early Career Scientist Award and Margaret Oakley Dayhoff Award in Biophysics. She serves on the editorial board of the Journal of Biological Chemistry, and as an academic editor of the journal Plos-ONE. She is a permanent member and a chair of the American Cancer Society “DNA mechanisms in cancer review panel.

Bibliographic information

• Title: Single-Molecule Enzymology: Fluorescence-Based and High-Throughput Methods
• Contributors: Maria Spies, Yann R Chemla
• Publisher: Academic Press, 2016
• ISBN: 0128095474, 9780128095478
• Length: 616 pages
• Subjects: Science / Life Sciences / Biochemistry; Science / Life Sciences / Biophysics; Science / Life Sciences / Molecular Biology