| First Author | Ajam T | Year | 2020 |
| Journal | Sci Rep | Volume | 10 |
| Issue | 1 | Pages | 9828 |
| PubMed ID | 32555289 | Mgi Jnum | J:296958 |
| Mgi Id | MGI:6452061 | Doi | 10.1038/s41598-020-66795-z |
| Citation | Ajam T, et al. (2020) Alternative catalytic residues in the active site of Esco acetyltransferases. Sci Rep 10(1):9828 |
| abstractText | Cohesin is a protein complex whose core subunits, Smc1, Smc3, Scc1, and SA1/SA2 form a ring-like structure encircling the DNA. Cohesins play a key role in the expression, repair, and segregation of eukaryotic genomes. Following a catalytic mechanism that is insufficiently understood, Esco1 and Esco2 acetyltransferases acetylate the cohesin subunit Smc3, thereby inducing stabilization of cohesin on DNA. As a prerequisite for structure-guided investigation of enzymatic activity, we determine here the crystal structure of the mouse Esco2/CoA complex at 1.8 A resolution. We reconstitute cohesin as tri- or tetrameric assemblies and use those as physiologically-relevant substrates for enzymatic assays in vitro. Furthermore, we employ cell-based complementation studies in mouse embryonic fibroblast deficient for Esco1 and Esco2, as a means to identify catalytically-important residues in vivo. These analyses demonstrate that D567/S566 and E491/S527, located on opposite sides of the murine Esco2 active site cleft, are critical for catalysis. Our experiments support a catalytic mechanism of acetylation where residues D567 and E491 are general bases that deprotonate the epsilon-amino group of lysine substrate, also involving two nearby serine residues - S566 and S527- that possess a proton relay function. |
