A Simple Light Isotope Metabolic Labeling (SLIM-labeling) Strategy: A Powerful Tool to Address the Dynamics of Proteome Variations In Vivo

Many quantitative proteomics strategies rely on in vivo metabolic incorporation of amino acids with modified stable isotope profiles into proteins. These methods give rise to multiple ions for each peptide, with possible distortion of the isotopolog distribution, making the overall analytical process complex. We validated an alternative strategy, simple light isotope metabolic labeling (SLIM-labeling), which alleviates many of these problems. SLIM-labeling is based on the in vivo reduction of the isotopic composition of proteins using metabolic precursors with a unique light isotope composition to label all amino acids. This brings a new dimension to in-depth, high resolution MS-based quantitative proteomics. Here, we describe a C-12-based SLIM-labeling strategy using U-[C-12]-glucose as the metabolic precursor of all amino acids in the pathogenic yeast Candida albicans. Monoisotopic ion intensity increased exponentially following C-12 enrichment, substantially improving peptide identification scores and protein sequence coverage in bottom-up analyses. Multiplexing samples of C-12 composition varying from natural abundance (98.93%) to 100% makes it possible to address relative quantification issues, keeping all the critical information for each peptide within a single isotopolog cluster. We applied this method to measure, for the first time, protein turnover at the proteome scale in Candida albicans and its modulation by inhibitors of the proteasome and vacuolar protein degradation systems.