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Full Description
This volume is comprised of a collection of experimental protocols for common techniques and strategies used to study the biogenesis of b-barrel outer membrane proteins in Gram-negative bacteria. The BAM Complex: Methods and Protocols guides readers through methods on the function of the BAM complex, the roles played by each of the individual components, the expression and purification of the components, crystallization and structure determination of the components, and how the individual Bam components may assemble into a functional complex. 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, The BAM Complex: Methods and Protocols will serve as an invaluable reference for those interested in studying the BAM complex.
Contents
The β-barrel Assembly Machinery Comple.- Yeast Mitochondria as a Model System to Study the Biogenesis of Bacterial β-barrel Proteins.- Experimental Methods for Studying the BAM Complex in Neisseria meningitides.- Heat Modifiability of Outer Membrane Proteins from Gram-Negative Bacteria.- The Role of a Destabilized Membrane for OMP Insertion.- Treponema pallidum in Gel Microdroplets: A Method for Topological Analysis of BamA (TP0326) and Localization of Rare Outer Membrane Proteins.- Analyzing the Role of Periplasmic Folding Factors in the Biogenesis of OMPs and Members of the Type V Secretion System.- An In Vitro Assay for Substrate Translocation by FhaC in Liposomes.- Measuring Cell-cell Binding Using Flow-Cytometry.- Methods to Characterize Folding and Function of BamA Crosslink Mutants.- Small Angle X-ray Scattering (SAXS) Characterization of the POTRA Domains of BamA.- Assessing the Outer Membrane Insertion and Folding of Multimeric Transmembrane β-barrel Proteins.- The Expression, Purification, and Structure Determination of BamA from E. coli.- Expression and Purification of the Individual Bam Components BamB-E.- Structure Determination of the BAM Complex Accessory Lipoproteins BamB-E.- An In Vitro Assay for Outer Membrane Protein Assembly by the BAM complex.- Identification of BamC on the Surface of E. coli.- Construction and Characterization of an E. coli bamD Depletion Strain.- Expression, Purification and Screening of BamE, a Component of the BAM Complex, for Structural Characterization.- Purification and Bicelle Crystallization for Structure Determination of the E. coli Outer Membrane Protein TamA.- Strategies for the Analysis of Bam Recognition Motifs in Outer Membrane Proteins.- Summary and Future Directions.
