Encode Bio (United States)

Previously, the group has demonstrated the feasibility of determining in‐vivo protein synthesis by utilizing deuterated water labeling [1] and in‐vitro protein synthesis with 15N‐glycine labeling [2]. The study is designed to give preliminary proof of concept that the 15N‐glycine labeling can also be applied for in‐vivo protein synthesis determination. Two mice were fed 20 g of 15N glycine/kg of chow for 7 days. The mice serum albumin was SDS‐PAGE purified, followed by in‐gel tryptic digestion, before MALDI‐TOF profiling. Mass spectra data for two specific peptide sequences with mass to charge (m/z) of 1479 and 1609 were analyzed for 15N enrichment and fraction of new synthesis. For m/z =1479, average enrichment of 15N in the peptide was found to be 6.06% for mouse 1 and 6.10% for mouse 2, with fraction of new synthesis found to be 97.52% and 96.54% respectively. The 15N enrichment for m/z =1609 was found to be 5.25% for mouse 1 and 6.31% for mouse 2, with fraction of new synthesis of 90.24% and 88.28% respectively. The mass spectral peaks of the heavier protein can be resolved from the natural protein by a mathematical algorithm. The capability of resolving overlapping spectra makes it possible to label proteins with lower enrichment of stable isotopes.