Analysis of the Proteome of Saccharomyces cerevisiae for Methylarginine

Arginine methylation
is a post-translational modification that
has been implicated in a plethora of cellular processes. In the present
manuscript, using two antimethylarginine antibodies and combinatorial
deletion mutants of arginine methyltransferases, we found evidence
of widespread arginine methylation in the Saccharomyces
cerevisiae proteome. Immunoprecipitation was used
for enrichment of methylarginine-containing proteins, which were identified
via tandem mass spectrometry. From this, we identified a total of
90 proteins, of which 5 were previously known to be methylated. The
proteins identified were involved in known methylarginine-associated
biological functions such as RNA processing, nuclear transport, carbohydrate
metabolic process, GMP biosynthetic process and protein folding. Through
in vivo methylation by the incorporation of [3H]-methyl
groups, we validated the methylation of 7 proteins (Ded1, Imd4, Lhp1,
Nop1, Cdc11, Gus1, Pob3). By LC–MS/MS, we then confirmed a
total of 15 novel methylarginine sites on 5 proteins (Ded1, Lhp1,
Nop1, Pab1, and Ugp1). By examination of methylation on proteins from
the triple knockout of methyltransferases Hmt1, Hsl7, Rmt2, we present
evidence for the existence of additional unidentified arginine methyltransferases
in the Saccharomyces cerevisiae proteome.