Proteome scale turnover analysis in live animals using stable isotope metabolic labeling. Compartment modeling for mammalian protein turnover studies by stable isotope metabolic labeling. Protein turnover: measurement of proteome dynamics by whole animal metabolic labelling with stable isotope labelled amino acids. Heterogenous turnover of sperm and seminal vesicle proteins in the mouse revealed by dynamic metabolic labeling. Proteome dynamics: revisiting turnover with a global perspective. Identification of long-lived synaptic proteins by proteomic analysis of synaptosome protein turnover. Identification of long-lived proteins reveals exceptional stability of essential cellular structures. Extremely long-lived nuclear pore proteins in the rat brain. 15N metabolic labeling of mammalian tissue with slow protein turnover. Analysis of proteome dynamics in the mouse brain. C., Guan, S., Burlingame, A., Prusiner, S. Measurement of protein turnover rates by heavy water labeling of nonessential amino acids. Mitochondrial protein turnover: Methods to measure turnover rates on a large scale. Precisely measured protein lifetimes in the mouse brain reveal differences across tissues and subcellular fractions. The Dynamic State of Body Constituents (Harvard University Press, 1946).įornasiero, E. A practical recipe for stable isotope labeling by amino acids in cell culture (SILAC). The biology of proteostasis in aging and disease. Overall, the workflow presented here comprises several improvements in the determination of protein lifetimes with respect to other available methods, including a minimally invasive labeling strategy and a robust interpretation of MS results, thus enhancing reproducibility across laboratories. We also provide all the scripts needed for the interpretation of the MS results and for comparing turnover across different conditions. Finally, we present a simple experimental workflow that enables measurement of precise turnover rates in a time frame of ~4–5 weeks, including the labeling time. We then cover in detail the practical aspects of metabolic labeling and explain both the experimental and computational steps that must be taken to obtain accurate in vivo results. We start by discussing fundamental concepts of protein turnover, including different methodological approaches. Here, we describe a robust protocol for quantitative proteome turnover analysis in mice that is based on a commercially available diet for stable isotope labeling of amino acids in mammals (SILAM). Previous approaches to evaluating turnover in vivo have required custom labeling approaches, involved complex mass spectrometry (MS) analyses, or used comparative strategies that do not allow direct quantitative measurements. Despite its importance, protein turnover has been difficult to measure in vivo. Proteins are continually produced and degraded, to avoid the accumulation of old or damaged molecules and to maintain the efficiency of physiological processes.
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |