Protein interactomes of microbial pathogens comprise the total interactions of proteins that occur within the cell, and are roadmaps to the molecular processes and biochemical pathways that govern cellular functions and virulence. By examining isolated binary protein complexes in a cell-free environment, systems-level networks can be constructed from the bottom-up. Alternatively, by starting with the proteome, a systems-level network can be broken down from the top to identify interactions within whole cells. I explored these experimental methods with two Gram-negative pathogens, Yersinia pestis and Klebsiella pneumoniae. I used a proteome microarray of the plague bacterium Y. pestis to generate a database of>77,000 binary protein interactions. The database provided a means to understand Y. pestis proteins in the context of cognate interaction partners and a foundation to model the Y. pestis interactome from the bottom-up. I integrated sequence-based comparisons of homology, pathways analysis, protein abundance measurements, and transcript expression levels to reveal complexes and pathways involved in replication, biosynthesis, virulence, metabolism, and other diverse biological processes. To understand the proteomic response of bacteria that may circumvent antibiotic treatment with drugs that inhibit the ribosome, I developed a culture-based model of persistent infections with an invasive strain of extended-spectrum beta-lactamase producing K. pneumoniae to identify drug-specific networks from the top-down. The proteomes of cells that were tolerant to streptomycin or doxycycline, identified by label-free mass spectrometry, indicated fluctuations ({plusmn}2-fold) in the abundance of>600 proteins. I integrated pathways analysis, transcription factor binding sites, and experimental validation of predicted networks to reveal distinct bacterial responses that were exclusive of the anticipated ribosomal target and were unique to each drug. Because many hypothetical proteins were detected in the proteomic datasets of Y. pestis and K. pneumoniae, my results expand proteome annotation for these under-studied organisms. The approaches described here illustrate the application of proteome-scale interactomes for the prediction of networks that function within important human pathogens. Importantly, the interactomes of these Gram-negative pathogens are a potential source of targets for the development of new antimicrobials and vaccines, and offer considerations for understanding the emergence of multi-drug resistant bacteria.

The first of its kind, this volume presents the latest research findings on the chaperonins, the best studied family of a class of proteins known as molecular chaperones. These findings are changing our view of some fundamental cellular processes involving proteins, especially how proteins fold into their functional conformations. - Origins of the new view of protein folding - Prokaryotic chaperonins - Eukaryotic chaperonins - Evolution of the chaperonins - Refolding of denatured proteins - Organelle biosynthesis - Biomedical aspects

Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC): Methods and Protocols provides a synopsis of a large array of different SILAC methods by presenting a set of protocols that have been established by renowned scientists and their working groups. These include methods and protocols for the labeling of various model organisms as well as advanced strategies relying on SILAC, e.g. for the analysis of protein interactions, the mapping of posttranslational modifications or the characterization of subcellular proteomes. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC): Methods and Protocols will serve students and experienced scientists alike as a valuable reference of how to make use of the SILAC technology for their own research.

Vectors of Plant Pathogens is a collection of papers that discusses the interrelationship of plant pathogens with their vectors. This collection deals with the numerous vector groups associated with plant pathogens. One paper describes the biology, feeding behavior and distribution of aphids, leafhoppers, plant hoppers, mealy bugs, whiteflies, psyllids, membracids. Another paper addresses the virus transmission characteristics of the mealy bugs during preliminary fasting or feeding, acquisition access time, post-acquisition fasting or feeding, and the inoculation access time. Other papers also discuss the involvement of insects in transmitting bacterial and fungal pathogens; the authors list unresolved issues such as the role of insects in overwintering of bacterial pathogens or the association of the fungus with a particular vector. One author describes some suspected fungi transmission such as the pea stem necrosis virus, red clover necrotic mosaic virus, and the tomato bushy stunt virus. Another paper examines the fate of plant viruses in mite vectors and convectors particularly the viruses found in wheat, barley, or brome grass. Agriculturists, botanists, and researchers in the field of botany, conservation, and plant genealogy will find this book useful.

This volume details protocols that cover the broad arsenal of techniques used to study a secretion system from A to Z. Chapters focus on identifying and localizing the different subunits, defining interactions within subunits, monitoring conformational changes, purifying and imaging of large complexes, defining the assembly pathway by fluorescence microscopy and the role of energy during assembly and/or secretion, identifying secreted effectors as well as reporters to follow effector transport. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Bacterial Protein Secretion Systems: Methods and Protocol aims to provide techniques that are not restricted to the study of secretion systems but are also of specific interest for any researcher interested on multi-protein complexes of the bacterial cell envelope.

This book provides comprehensive up-to-date information on the structure and function of immunoglobulins. It describes the basic features of these molecules, which assists the reader in understanding how they function as an integral part of the immune system. The Immunoglobulins describes the localization and structure of different binding sites of immunoglobulin molecules, including the antigen-binding site, on the basis of latest x-ray crystallography studies. It discusses recently developed biotechnological methods that allow scientists to obtain fully active antibody molecules in vitro even without immunization and to construct new variants of immunoglobulins and their fragments by fusing with various other active molecules. A survey of recent knowledge on immunoglobulin-binding molecules other than antigens and on flexibility of immunoglobulin molecules concludes the discussion of functional aspects of the problem. - Describes recent reviews on the structure and function of immunoglobulin molecules of various species - Summarizes in detail recent findings on the fine structure of the antigen-combining site - Presents comparative data on the antigen-recognizing sites of other molecules such as MHC proteins and T-cell receptors - Summarizes growing data on immunoglobulin binding sites responsible for the reaction of immunoglobulins with molecules other than antigens - Explores the rapid advance of recent biotechnological methods used for the construction of antibody molecules and their fragments with new properties - Presents extensive references and is lavishly illustrated

Fungi: Biology and Applications, Second Edition provides a comprehensive treatment of fungi, covering biochemistry, genetics and the medical and economic significance of these organisms at introductory level. With no prior knowledge of the subject assumed, the opening chapters offer a broad overview of the basics of fungal biology, in particular the physiology and genetics of fungi and also a new chapter on the application of genomics to fungi. Later chapters move on to include more detailed coverage of topics such as antibiotic and chemical commodities from fungi, new chapters on biotechnological use of fungal enzymes and fungal proteomics, and fungal diseases of humans, antifungal agents for use in human therapy and fungal pathogens of plants.