This volume provides classic and new methods to study the structure, assembly pathway, and protein synthesis ability of mitoribosomes across species. Following an introduction of fundamental concepts on the topic, method chapters present detailed protocols based on cryo-electron tomography, cryo-EM approaches, mitoribosome purification techniques, mitochondrial translation assays, and methods to study mitochondrial mRNAs that are translated on mitoribosomes. 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 methods, includes tips on troubleshooting and known pitfalls, and step-by-step, readily reproducible protocols.
Authoritative and cutting-edge, The Mitoribosome: Methods and Protocols, aims to be a comprehensive guide for researchers in the field.
Inhaltsverzeichnis
Discovery of Mitochondrial Ribosomes. - Evolution: Mitochondrial Ribosomes across Species. - Mitoribosome Biogenesis. - Translation in Mitochondrial Ribosomes. - Sample Preparation of Isolated Mitochondria for Cryoelectron Tomography and in situ Studies of Translation. - Cryo-EM for Structure Determination of Mitochondrial Ribosome Samples. - Sucrose Gradient Analysis of Human Mitochondrial Ribosomes and RNA. - Yeast Mitoribosome Purification and Analyses by Sucrose Density Centrifugation and Immunoprecipitation. - Rapid Cryo-Purification of the Yeast Mitochondrial Ribosome. - Methods to Study the Biogenesis of Mitoribosomal Proteins in Yeast. - Systematic Analysis of Assembly Intermediates in Yeast to Decipher the Mitoribosome Assembly Pathway. - Metabolic Labelling of Mitochondrial Translation Products in Whole Cells and Isolated Organelles. - Assembly of the Mitochondrial Translation Initiation Complex. - Reconstitution of Mammalian Mitochondrial Translation System Capable of Long Polypeptide Synthesis. - Human Mitoribosome Profiling: a Re-engineered Approach Tailored to Study Mitochondrial Translation. - The ARG8
m Reporter for the Study of Yeast Mitochondrial Translation. - Four Colour Super-Resolution RNA Fluorescent In Situ Hybridization and Immunocytochemistry to Visualise Mitochondrial mRNAs in Context with Mitochondrial Ribosomes. - Digital RNase Footprinting of RNA-protein Complexes and Ribosomes in Mitochondria. - Dead-seq: Discovering Synthetic Lethal Interactions from Dead Cells Genomics.
