Erce MA, Abeygunawardena D, Low JK, Hart-Smith G, Wilkins MR

Protein-protein interactions can be modulated by the methylation of arginine residues. As a means of testing this, we recently described a conditional two-hybrid (C2H) system, based on the bacterial adenylate cyclase (BACTH) system. Here, we have used this C2H system to explore the effect of arginine methylation in modulating protein-protein ...

Read More

interactions in a subset of the Saccharomyces cerevisiae arginine methylproteome network. Interactions between the yeast hub protein Npl3 and yeast proteins Air2, Ded1, Gbp2, Snp1 and Yra1 were first validated in the absence of methylation. The major yeast arginine methyltransferase Hmt1 was subsequently included in the C2H assay, initially to determine the degree of methylation which occurs. Proteins Snp1 and Yra1 were confirmed as Hmt1 substrates, with five and two novel arginine methylation sites mapped by ETD LC-MS/MS on these proteins, respectively. Proteins Ded1, and Gbp2, previously predicted but not confirmed as substrates of Hmt1, were also found to be methylated with 5 and 7 sites mapped respectively. Air2 was found to be a novel substate of Hmt1 with 2 sites mapped. Finally, we investigated the interactions of Npl3 with the five interaction partners in the presence of active Hmt1 and in the presence of Hmt1 with a G68R inactivation mutation. We found that the interaction between Npl3 and Air2, and Npl3 and Ded1, were significantly increased in the presence of active Hmt1; the interaction of Npl3 and Snp1 showed a similar degree of increase in interaction but this was not statistically significant. The interactions of Npl3 and Gbp2, along with Npl3 and Yra1, were not significantly increased or decreased by methylation. We conclude that methylarginine may be a widespread means by which the interactions of proteins are modulated.

Date: Aug. 05, 2013

View in: Pubmed Google Scholar

157369