Linking Intestinal Microorganisms to their Diet-derived Growth Substrates Using Protein Stable Isotope Fingerprinting (Protein-SIF)

The aim of this study was to directly link intestinal microorganisms to their diet-derived in vivo growth substrates using protein stable isotope fingerprinting (“Protein-SIF”)(Kleiner et al. 2018, PNAS 115(24)). We conducted studies in gnotobiotic mice (C57BL/6J) colonized with a community of 13 human-derived bacterial strains (Desai et al. 2016, Cell 167(5)). There were two groups of mice: group 1 consisted of 5 mice, and group 2 of 6 mice. All mice were fed defined diets modeled on the AIN-93G (EnvigoTeklad) varying in one ingredient source, and a “standard chow” (LabDiet 5010) as a baseline diet before and after the defined diets. Group 1 mice were fed diets differing in protein source (casein, egg whites, soy protein). Group 2 mice were fed diets differing in fiber source (cellulose, inulin, corn fiber), and fat source (corn oil, soybean oil, sunflower oil). Stable carbon isotope ratios for each dietary component were measured using IRMS (table with values is included in submission). Each diet was introduced for 7 days, and fecal samples were collected on the 7th day. We processed samples from the group 1 mice and the group 2 mice as two separate batches. We analyzed the samples by LC-MS/MS. Each sample was run in four consecutive technical replicates to increase the available number of MS1 spectra for SIF calculations. Data were analyzed with Proteome Discoverer followed by the calis-p software for Protein-SIF (https://sourceforge.net/projects/calis-p/). Additionally, this dataset includes diet samples and purified dietary components that were extracted using the same approach as the fecal samples, as well as LC-MS/MS runs of human hair derived peptides used for SIF calibration.