title: Mapping of POP1-binding site on pyrin domain of ASC.
abstract: Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is an essential adaptor protein in the formation of a multiprotein complex that activates procaspase-1. ASC is also known as a modulator of NF-kappaB activation pathways. ASC has a bipartite domain structure, consisting of an N-terminal pyrin domain (PYD) and a C-terminal caspase-recruitment domain. The PYD of ASC (ASC_PYD) is known to interact with various PYD-containing intracellular danger signal sensors and PYD-only proteins. Using purified proteins, we characterized the in vitro interaction of ASC_PYD with PYD-only protein 1 (POP1). POP1 specifically interacts with ASC_PYD with a dissociation constant of 4.08 +/- 0.52 microm but does not interact with Cryopyrin. NMR and mutagenesis experiments show that a negative electrostatic potential surface patch (EPSP) on ASC_PYD, consisting of the first (H1) and fourth (H4) helices, is essential in the interaction with POP1. A positive EPSP on POP1, consisting of the second (H2) and third (H3) helices, is a counterpart of this interaction. The interaction between ASC_PYD and POP1 is similar to the interaction between caspase recruitment domains of Apaf-1 and procaspase-9. In addition, we present evidence that conformational changes at the long loop of ASC_PYD between the H2 and H3 helices can affect its interaction with POP1. Based on our observations, we propose that the positive EPSP of ASC_PYD, including the H2 and H3 helices, may be the binding site for Cryopyrin, and the interaction with Cryopyrin may induce the dissociation of POP1 from ASC_PYD.
authors: Basic Science, Fox Chase Cancer Center, 333 Cottman Avenue, Philadelphia, PA 19111, USA.
authors: Thiagarajan
authors: T
authors: Srimathi
authors: Sheila L
authors: SL
authors: Robbins
authors: Rachel L
authors: RL
authors: Dubas
authors: Helen
authors: H
authors: Chang
authors: Hong
authors: H
authors: Cheng
authors: Heinrich
authors: H
authors: Roder
authors: Young Chul
authors: YC
authors: Park
mesh_terms: D051017:Apoptosis Regulatory Proteins / Q000737:chemistry* / Q000378:metabolism; D053477:Apoptotic Protease-Activating Factor 1 / Q000378:metabolism; D001665:Binding Sites / Q000502:physiology; D053468:CARD Signaling Adaptor Proteins; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D020170:Caspase 1 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D006801:Humans; D046912:Multiprotein Complexes / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; D010449:Peptide Mapping / Q000379:methods; D011485:Protein Binding / Q000502:physiology; D017433:Protein Structure, Secondary / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D000071198:Pyrin; D012261:Ribonucleoproteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: C506592:APAF1 protein, human; D051017:Apoptosis Regulatory Proteins; D053477:Apoptotic Protease-Activating Factor 1; D053468:CARD Signaling Adaptor Proteins; D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; C000604192:MEFV protein, human; D046912:Multiprotein Complexes; D016328:NF-kappa B; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C453881:NLRP3 protein, human; C490350:POP1 protein, human; C402202:PYCARD protein, human; D000071198:Pyrin; D012261:Ribonucleoproteins; D053453:Caspase 9; D020170:Caspase 1