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. 2019 Aug 26;18(11):2149–2164. doi: 10.1074/mcp.RA119.001625

Fig. 4.

Fig. 4.

Characteristic neutral loss of methylamine and dimethylamine allows discrimination of SDMA and ADMA spectra respectively. A, Mechanism of neutral loss for ADMA resulting in neutral loss of dimethylamine (45.058 Da). B, Mechanism of neutral loss for SDMA resulting in neutral loss of monomethylamine (31.042 Da). C, Analysis of synthetic ADMA and SDMA peptides from Zolg et al. (50) were searched for neutral losses using the Andromeda search engine. The synthetic peptides contained identical sequences and a single non-C-terminal ADMA or SDMA. Ambiguous identifications were defined as peptides who had conflicting neutral loss assignments and whose mean Andromeda scores differed by less than 30. Andromeda successfully identified 54.9% (78/142) of ADMA modifications and 73.1% (106/145) of SDMA modifications. Only one SDMA peptide was incorrectly identified as ADMA. D, MaxQuant neutral loss search applied to quantified DMA peptides from 293T cells expressing either shControl or shPRMT1 after enrichment by either SCX or ADMA/SDMA IAPs. Peptides showing neutral loss from the Andromeda search in MaxQuant were matched to their corresponding peptides in the LFQ data from Proteome Discoverer 2.2. Matching was performed by matching peptide sequence, retention time, methyl sites, and sample origin between neutral loss data and LFQ data. E, Annotated spectra of an ADMA peptide from SCX showing neutral losses of 45.058 for nine y ions. The inset peaks of each neutral loss fragment are shown with some fragments showing the isotopic envelope typical of peptides. An inset table shows the masses for each y ion and y ion neutral loss fragment, as well as the differences in mass for each pair. F, Annotated spectra of an SDMA peptide from SCX showing neutral losses of 31.042 Da for six y ions. The inset peaks of each neutral loss fragment are shown with some fragments showing the isotopic envelope typical of peptides. An inset table shows the masses for each y ion and y ion neutral loss fragment, as well as the differences in mass for each pair.