Protein Protocols

This book presents modern and classic analytical approaches that are crucial for the biochemical and functional characterization of the archetypal protein, glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The distinguishing feature of the book is that it covers, in addition to other methods, some of the uncommon but valuable techniques as well. For example, in-gel visualization of enzyme activity, immunoblotting protocols for native (non-denatured) proteins, and proteins resolved by pH-gradient [IEF-isoelectrofocusing], etc. These expedient methods are relevant and vital for the verification of biochemical properties of GAPDH, or similar protein of interest.

This volume focuses on applications of split inteins, and the progress that has been made in the past 5 years on discovery and engineering of fast and more efficient split inteins. The first few chapters in Split Inteins: Methods and Protocols explore new techniques on how to use split inteins for affinity purification of overproduced proteins, and split-intein based technologies to prepare cyclic peptides and proteins.

This volume provides updated protocols for chemical protein synthesis. Chapters guide readers through development methods, strategies, and applications of protein chemical synthesis. Written in the format of the highly successful Methods in Molecular Biology series, each chapter includes an introduction to the topic, lists necessary materials and reagents, includes tips on troubleshooting and known pitfalls, and step-by-step, readily reproducible protocols.

This volume presents a collection of protocols to study effector-triggered immunity (ETI) in both plants and animals from eminent groups in the field. The chapters in this book cover topics such as genetic manipulation of plant and animal pathogens, host cells, and the analysis of key host responses; and techniques used for the analysis of inflammasome activation, cell death pathways, and mitochondria damage in response to pathogens. All of these topics cover a broad spectrum of immunological, biochemical, cell biological, and structural biology approaches to examine ETI. 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.

This volume provides a comprehensive guide on the Cyr61 (Cysteine-rich 61) (CCN) family of proteins and genes from basic research to cutting-edge methodologies and state-of-the-art techniques. Chapters details practical tips to overcome any obstacles with experimentation pertaining to chemistry, biology, physiology, pathology, medical and dental sciences and pharmacology of CCN proteins.

This volume provides readers with a comprehensive look at the latest techniques used to identify and characterize PDZ-mediated interactions. Chapters cover topics such as promiscuity, multimodularity, regulation, and viral recognition by PDZ domains. 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.

David Higgins and James Cregg compile for the first time an all-inclusive collection of key experimental procedures to manipulate Pichia pastoris for gene expression purposes. The methods-developed by world-class researchers in their own laboratories-contain detailed descriptions of available expression strains, as well as vectors and step-by-step instructions for the construction of expression strains and for their use in the production of foreign proteins from shake-flask or fermentor cultures. General procedures for the purification of foreign proteins are also provided, along with protocols for analyzing foreign protein products for glycosylation and subcellular location. Its detailed instructions will permit any researcher in molecular biology to quickly construct an expression strain that will produce a desired foreign protein, and to use this state-of-the-art molecular and cellular biological system successfully in every area of biological research today.

This detailed volume explores newly-developed methods in PIWI-interacting RNAs (piRNAs) research, methods currently applied to other ncRNAs involved in nuclear regulation which can be used to study piRNAs, and piRNA methods applied in non-classical organisms. It also includes several bioinformatic and biophysical methods related to piRNA studies, consistent with the increasing importance of high-throughput sequencing and computational methods. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials, step-by-step, readily reproducible protocols, and tips on troubleshooting and avoiding known pitfalls.