7ad9 | pdb_00007ad9
Structure of the Lifeact-F-actin complex
Structural highlights
FunctionAB140_YEAST S-adenosyl-L-methionine-dependent methyltransferase that mediates N(3)-methylcytidine modification of residue 32 of the tRNA anticodon loop of tRNA(Thr) and tRNA(Ser) (PubMed:21518804, PubMed:21518805, PubMed:28003514). N(3)-methylcytidine methylation of tRNA(Thr) requires the N6-threonylcarbamoylation of tRNA (t6A37) by the EKC/KEOPS complex as prerequisite (PubMed:28003514). N(3)-methylcytidine methylation of tRNA(Ser) requires the formation of N(6)-dimethylallyladenosine(37) (i6A37) by MOD5 as prerequisite (PubMed:28003514). Methylation of tRNA(Ser) is also stimulated by SES1 (PubMed:28003514). Binds F-actin and shows weak F-actin cross-linking activity (PubMed:9467951).[1] [2] [3] [4] Publication Abstract from PubMedLifeact is a short actin-binding peptide that is used to visualize filamentous actin (F-actin) structures in live eukaryotic cells using fluorescence microscopy. However, this popular probe has been shown to alter cellular morphology by affecting the structure of the cytoskeleton. The molecular basis for such artefacts is poorly understood. Here, we determined the high-resolution structure of the Lifeact-F-actin complex using electron cryo-microscopy (cryo-EM). The structure reveals that Lifeact interacts with a hydrophobic binding pocket on F-actin and stretches over 2 adjacent actin subunits, stabilizing the DNase I-binding loop (D-loop) of actin in the closed conformation. Interestingly, the hydrophobic binding site is also used by actin-binding proteins, such as cofilin and myosin and actin-binding toxins, such as the hypervariable region of TccC3 (TccC3HVR) from Photorhabdus luminescens and ExoY from Pseudomonas aeruginosa. In vitro binding assays and activity measurements demonstrate that Lifeact indeed competes with these proteins, providing an explanation for the altering effects of Lifeact on cell morphology in vivo. Finally, we demonstrate that the affinity of Lifeact to F-actin can be increased by introducing mutations into the peptide, laying the foundation for designing improved actin probes for live cell imaging. Structure of the Lifeact-F-actin complex.,Belyy A, Merino F, Sitsel O, Raunser S PLoS Biol. 2020 Nov 20;18(11):e3000925. doi: 10.1371/journal.pbio.3000925. , eCollection 2020 Nov. PMID:33216759[5] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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