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Synthetic trap-peptides allow the identification the first phosphatase of the mitochondrial import machinery

Sep 02, 2025

The mitochondrial translocase of the outer membrane (TOM) complex is a heterooligomeric protein machinery responsible for the import of proteins into mitochondria.  Many components of this machinery possess phosphorylation sites, some of which play a significant role in the regulation of import. While kinases responsible for phosphorylating several of these sites have already been identified, the counteracting phosphatases remain elusive.

In a recent study, Laura Scheinost and Maja Köhn (both Project AI06) together with SFB 1381 PIs Nora Vögtle (B04) and Chris Meisinger (A06) developed synthetic trap-peptides to identify phosphatases interacting with Tom6, one subunit of the TOM complex. Using trap-peptides, they successfully enriched full holoenzymes of protein phosphatases 2A (PP2A) and 4 (PP4) from yeast cytosolic extracts. Their interaction with Tom6 was mediated by the regulatory subunits Cdc55^reg (PP2A) and Psy2^reg (PP4), and in vitro assays confirmed that PP2A can dephosphorylate Ser16 on Tom6.

Overall, this approach enabled the identification of complete holoenzymes binding to the target site and revealed PP2A as the first known phosphatase of the protein import machinery.

These results have been published in the FEBS Journal.

Illustration: Laura Scheinost