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In recent years, a number of groundbreaking structural and mechanistic studies deepened our understanding of helicase mechanisms and established new approaches for their analyses. Many fundamental mechanistic questions ranging from the mechanism of force generation, mechanochemical coupling to distinct mechanisms by which the same enzyme translocates on DNA removing obstacles, unwinds DNA and/or remodels nucleoprotein complexes, however, remain to be answered. It is even less understood how the helicase motors are incorporated into a wide range of genome maintenance and repair machines. The field has reached a stage when the studies of molecular mechanisms and basic biology of helicases can and shall be integrated with the studies of development, cancer and longevity. The objective of this book is to provide the first systematic overview of structure, function and regulation of DNA helicases and related molecular motors. By integrating the knowledge obtained through the diverse technical approaches ranging from single-molecule biophysics to cellular and molecular biological studies the editors aim to provide a unified view on how helicases function in the cell, are regulated in response to different cellular stresses and are integrated into large macromolecular assemblies to form a complex and adaptive living system.
Helicases are the proteins that bind to double- or single-stranded DNA and/or RNA chains to unwind higher order structures, usually consuming energy from the hydrolysis of ATP molecules. The biological roles of helicases are associated with a variety of DNA and/or RNA metabolisms, including DNA-replication, -repair, -recombination, RNA processing, and transcription. Dysfunctions of helicases cause various diseases, such as xeroderma pigmentosum (XP), premature aging syndrome, cancer and immunodeficiency, in humans. Moreover, recent genetic analyses revealed that mutations in helicase-encoding genes are frequently found in patients of specific diseases. Some helicases regulate cellular senescence by controlling integrity of genomes, and others play a role in neuromuscular functions presumably by modulating processing of mRNAs. However, the molecular mechanisms of how helicases are regulated in order to maintain our health are not yet fully understood. In this research topic, we will focus on the expression and functions of helicases and their encoding genes, reviewing recent research progresses that provide new insights into development of clinical and pharmaceutical treatments targeting helicases.
DNA Helicases—Advances in Research and Application: 2012 Edition is a ScholarlyPaper™ that delivers timely, authoritative, and intensively focused information about DNA Helicases in a compact format. The editors have built DNA Helicases—Advances in Research and Application: 2012 Edition on the vast information databases of ScholarlyNews.™ You can expect the information about DNA Helicases in this eBook to be deeper than what you can access anywhere else, as well as consistently reliable, authoritative, informed, and relevant. The content of DNA Helicases—Advances in Research and Application: 2012 Edition has been produced by the world’s leading scientists, engineers, analysts, research institutions, and companies. All of the content is from peer-reviewed sources, and all of it is written, assembled, and edited by the editors at ScholarlyEditions™ and available exclusively from us. You now have a source you can cite with authority, confidence, and credibility. More information is available at http://www.ScholarlyEditions.com/.
DNA Helicases—Advances in Research and Application: 2012 Edition is a ScholarlyPaper™ that delivers timely, authoritative, and intensively focused information about DNA Helicases in a compact format. The editors have built DNA Helicases—Advances in Research and Application: 2012 Edition on the vast information databases of ScholarlyNews.™ You can expect the information about DNA Helicases in this eBook to be deeper than what you can access anywhere else, as well as consistently reliable, authoritative, informed, and relevant. The content of DNA Helicases—Advances in Research and Application: 2012 Edition has been produced by the world’s leading scientists, engineers, analysts, research institutions, and companies. All of the content is from peer-reviewed sources, and all of it is written, assembled, and edited by the editors at ScholarlyEditions™ and available exclusively from us. You now have a source you can cite with authority, confidence, and credibility. More information is available at http://www.ScholarlyEditions.com/.
This is an introductory text and laboratory manual to be used primarily in undergraduate courses. It is also useful for graduate students and research scientists who require an introduction to the theory and methods of nanopore sequencing. The book has clear explanations of the principles of this emerging technology, together with instructional material written by experts that describes how to use a MinION nanopore instrument for sequencing in research or the classroom.At Harvard University the book serves as a textbook and lab manual for a university laboratory course designed to intensify the intellectual experience of incoming undergraduates while exploring biology as a field of concentration. Nanopore sequencing is an ideal topic as a path to encourage students about the range of courses they will take in Biology by pre-emptively addressing the complaint about having to take a course in Physics or Maths while majoring in Biology. The book addresses this complaint by concretely demonstrating the range of topics — from electricity to biochemistry, protein structure, molecular engineering, and informatics — that a student will have to master in subsequent courses if he or she is to become a scientist who truly understands what his or her biology instrument is measuring when investigating biological phenomena.
DNA replication is a fundamental part of the life cycle of all organisms. Not surprisingly many aspects of this process display profound conservation across organisms in all domains of life. The chapters in this volume outline and review the current state of knowledge on several key aspects of the DNA replication process. This is a critical process in both normal growth and development and in relation to a broad variety of pathological conditions including cancer. The reader will be provided with new insights into the initiation, regulation, and progression of DNA replication as well as a collection of thought provoking questions and summaries to direct future investigations.
In this dissertation we take advantage of the real-time manipulating, high force generating, and small displacement measuring abilities of dual-trap optical tweezers to investigate properties and mechanisms of motor enzymes that act on DNA, including the DNA translocation motor of bacteriophage lambda and the proposed ATP-driven annealing helicase HARP. Through a combined approach of genetic mutagenesis, biochemical analysis, and single-molecule analysis we reveal potential functional roles of the residues residing in the ATP phosphate binding motif (Walker A) of phage lambda's large terminase subunit. Direct DNA packaging measurements of phages with mutated residues within this motif reveal a range of impaired translocation phenotypes. Analysis of the slipping and pausing exhibited by mutant enzymes suggests that residues A78, R79, and V80 mediate coupling between ATP binding/hydrolysis and DNA binding/translocation while implicating residues A78, R79, V80, and G81 in proper ATP alignment for hydrolysis. The combined findings of each analysis also implicate residue R79 in ADP release and triggering ATP hydrolysis. In other studies, initial investigation into the mechanism of termination of DNA packaging in phage lambda provides evidence against a velocity-monitoring model. A higher resolution optical tweezers system was designed and constructed and a new and improved calibration method was developed and applied to initial high-resolution measurements of DNA translocation. The use of environmental and measurement noise reduction techniques and improved optical alignment procedures have yielded tweezers with approximately nanometer spatial resolution. The calibration technique, which optimizes fits of the worm-like chain model to DNA force-extension curves, yields calibration with at least a 7-fold improvement in accuracy over our prior method. The use of these tools in measuring lambda packaging yield preliminary evidence for discrete translocation steps. Lastly, tweezer measurements of HARP interactions with forked DNA provide supporting evidence that HARP is a forked-DNA binding protein that resists DNA unzipping and is not an unwinding helicase. In addition preliminary measurements suggest that this protein forcibly reanneals DNA, prevents reannealing, resists high forces, slows reannealing in an ATP dependent manner, and is recruited to bind to forked DNA with replication protein A coated single-stranded DNA.
This book focuses on the application of fluorescence to study motor proteins (myosins, kinesins, DNA helicases and RNA polymerases). It is intended for a large community of biochemists, biophysicists and cell biologists who study a diverse collection of motor proteins. It can be used by researchers to gain an insight into their first experiments, or by experienced researchers who are looking to expand their research to new areas. Each chapter provides valuable advice for executing the experiments, along with detailed background knowledge in order to develop own experiments.
Our understanding of the mechanisms regulating gene expression, which determine the patterns of growth and development in all living organisms, ultimately involves the elucidation of the detailed and dy namic interactions of proteins with nucleic acids -both DNA and RNA. Until recently the commonly presented view of the DNA double helix as visualized on the covers of many textbooks and journals - was as a monotonous static straight rod incapable in its own right of directing the processes necessary for the conservation and selective reading of genetic information. This view, although perhaps extreme, was reinforced by the necessary linearity of genetic maps. The reality is that the biological functions of both DNA and RNA are dependent on complex, and sometimes transient, three-dimensional nucleoprotein structures in which genetically distant elements are brought into close spatial proximity. It is in such structures that the enzymatic manipulation of DNA in the essential biological processes as DNA replication, transcription and recombination are effected - the complexes are the mediators of the 'DNA transactions' of Hatch Echols.