5h6l
From Proteopedia
DNA targeting ADP-ribosyltransferase Pierisin-1 in complex with beta-NAD+
Structural highlights
FunctionPRSN_PIERA ADP-ribosylates double-stranded DNA by targeting the N2 amino group of dG residues; 10-fold less active on tRNA. Induces apoptosis in a range of human cell lines (PubMed:10485873, PubMed:11592983). May play a role in removing unnecessary or harmful cells during pupation (PubMed:15528160) (Probable). May play a role in defense against parasitic wasps; its habitual predator Cotesia glomerata is only susceptible after removal of its eggshell or the surface of the larval caudal vesicle, while other wasps are damaged by this protein (PubMed:23637752) (Probable). The wild-type catalytic domain (residues 1-233) cannot be expressed in E.coli as it is unstable probably due to its toxicity. A catalytic domain fragment (Glu-165 mutated to Gln) binds dsDNA but not ssDNA; the presence of the linker inhibits DNA-binding (PubMed:28765284).[1] [2] [3] [4] [5] Publication Abstract from PubMedADP-ribosyltransferases transfer the ADP-ribose moiety of betaNAD+ to an acceptor molecule, usually a protein that modulates the function of the acceptor. Pierisin-1 is an ADP-ribosyltransferase from the cabbage butterfly Pieris rapae and is composed of N-terminal catalytic and C-terminal ricin B-like domains. Curiously, it ADP-ribosylates the DNA duplex, resulting in apoptosis of various cancer cells, which has raised interest in pierisin-1 as an anti-cancer agent. However, both the structure and the mechanism of DNA ADP-ribosylation are unclear. Here, we report the crystal structures of the N-terminal catalytic domain of pierisin-1, its complex with betaNAD+, and the catalytic domain with the linker connecting it to the ricin B-like domains. We found that the catalytic domain possesses a defined, positively charged region on the molecular surface but that its overall structure is otherwise similar to those of protein-targeting ADP-ribosyltransferases. Electrophoretic mobility shift assays and site-directed mutagenesis indicated that pierisin-1 binds double-stranded but not single-stranded DNA and that Lys122, Lys123, and Lys124, which are found in a loop, and Arg181 and Arg187, located in a basic cleft near the loop, are required for DNA binding. Furthermore, the structure of the catalytic domain with the linker revealed an autoinhibitory mechanism in which the linker occupies and blocks both the betaNAD+- and DNA-binding sites, suggesting that proteolytic cleavage to remove the linker is necessary for enzyme catalysis. Our study provides a structural basis for the DNA-acceptor specificity of pierisin-1 and reveals that a self-regulatory mechanism is required for its activity. Structural basis of autoinhibition and activation of the DNA-targeting ADP-ribosyltransferase pierisin-1.,Oda T, Hirabayashi H, Shikauchi G, Takamura R, Hiraga K, Minami H, Hashimoto H, Yamamoto M, Wakabayashi K, Shimizu T, Sato M J Biol Chem. 2017 Sep 15;292(37):15445-15455. doi: 10.1074/jbc.M117.776641. Epub , 2017 Aug 1. PMID:28765284[6] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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Categories: Large Structures | Pieris rapae | Hashimoto H | Hirabayashi H | Hiraga K | Minami H | Oda T | Sato M | Shikauchi G | Shimizu T | Sugimura T | Takamura R | Wakabayashi K | Yamamoto M
