Solution structure of the isolated Pelle death domain

Moncrieffe, Martin C.; Stott, Katherine

doi:10.1016/j.febslet.2005.06.009

Cited by 5 publications

(4 citation statements)

References 45 publications

(69 reference statements)

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“…Sample Preparation-The expression and purification of Pelle DD and Tube DD has been described previously (27). The expression and purification of the death domain of Drosophila MyD88 was similar to that of Pelle DD and Tube DD with the exception that protein expression was done using Rosetta (Novagen) cells.…”

Section: Methodsmentioning

confidence: 99%

“…Regarding the DD complexes involved in Toll signaling, solution data (26,27) strongly suggest a 1:1 interaction between Pelle and Tube. The crystal structure of the Pelle-Tube complex, although prepared from a heterodimeric DD solution, revealed a tetramer in the asymmetric unit consisting of two Pelle-Tube heterodimers arranged in a linear array (P1:T1:P2: T2) (22).…”

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confidence: 99%

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Assembly of Oligomeric Death Domain Complexes during Toll Receptor Signaling

Moncrieffe

Grossmann

2008

Journal of Biological Chemistry

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The Drosophila Toll receptor is activated by the endogenous protein ligand Spätzle in response to microbial stimuli in immunity and spatial cues during embryonic development. Downstream signaling is mediated by the adaptor proteins Tube, the kinase Pelle, and the Drosophila homologue of myeloid differentiation primary response protein (dMyD88). Here we have characterized heterodimeric (dMyD88-Tube) and heterotrimeric (dMyD88-Tube-Pelle) death domain complexes. We show that both the heterodimeric and heterotrimeric complexes form kidney-shaped structures and that Tube is bivalent and has separate high affinity binding sites for dMyD88 and Pelle. Additionally we found no interaction between the isolated death domains of Pelle and dMyD88. These results indicate that the mode of assembly of the heterotrimeric dMyD88-Tube-Pelle complex downstream of the activated Toll receptor is unique. The measured dissociation constants for the interaction between the death domains of dMyD88 and Tube and of Pelle and a preformed dMyD88-Tube complex are used to propose a model of the early postreceptor events in Drosophila Toll receptor signaling.The bifunctional Toll pathway in Drosophila is indispensable for embryonic development and the innate immune response to Gram-positive bacteria and fungi (1). Toll is a Type I transmembrane receptor (2) whose architecture consists of N-terminal leucine-rich repeats, a transmembrane helix, and a C-terminal TIR 2 domain. It is activated by direct binding of the ligand Spätzle (3) that is cleaved from its inactive pro-form by the serine protease Easter during development (4) and by another serine protease, Spätzle-processing enzyme, during Toll-dependent antimicrobial and fungal responses (5). Activation of the Toll pathway during development sets in motion a series of events that culminate in the translocation of the Rel/ NF-B family member Dorsal into the nucleus and the activation of the zygotic genes twist and snail on the ventral side of the embryos and the deactivation of zerknullt and decapentaplegic on the dorsal side (6), whereas activation resulting from an immune challenge leads to the nuclear translocation of the Dorsal-related immunity factor (Dif) and the subsequent transcription of many genes including those encoding the antimicrobial peptides Drosomycin and Metchnikowin (7,8). In mammals, an immune response is mounted as a result of challenge by a variety of pathogen-associated molecular patterns with the activation of a homologous Toll-like receptor pathway of which approximately 11 distinct members have been identified (9).Signaling from the activated Toll receptor to the relevant transcription factors relies on interactions between adaptor proteins that associate with the TIR domain of Toll and subsequently influence interactions between other downstream components of the pathway. In Drosophila, the immediate post-receptor events are context-dependent; during an innate immune response, for example, the proteins involved are dMyD88, Tube, and Pelle (10), whereas an additiona...

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

Assembly of Oligomeric Death Domain Complexes during Toll Receptor Signaling

Moncrieffe

Grossmann

2008

Journal of Biological Chemistry

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“…CD spectroscopy measurements have been previously reported for both FADD-DD and CD95-DD [34], with MRE values at 222 nm of ∼−18,000 deg.cm 2 .dmol −1 respectively, in reasonable agreement to the values obtained here for FADD-DD. Similarly, the death domain of the Drosophila protein Pelle was reported to possess a CD spectrum showing double minima at 209 and 223 nm, with a MRE (223 nm) of ∼−18,000 deg cm 2 dmol −1 [35]. The observation that the absolute ellipticity for the DAPk-DD(L) is more than 50% lower than observed for the FADD-DD indicates that the DAPk-DD does not exhibit the same level of α-helical secondary structure.…”

Section: Resultsmentioning

confidence: 96%

Structural and Functional Characterization of the Recombinant Death Domain from Death-Associated Protein Kinase

2013

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Death-associated protein kinase (DAPk) is a calcium/calmodulin-regulated Ser/Thr-protein kinase that functions at an important point of integration for cell death signaling pathways. DAPk has a structurally unique multi-domain architecture, including a C-terminally positioned death domain (DD) that is a positive regulator of DAPk activity. In this study, recombinant DAPk-DD was observed to aggregate readily and could not be prepared in sufficient yield for structural analysis. However, DAPk-DD could be obtained as a soluble protein in the form of a translational fusion protein with the B1 domain of streptococcal protein G. In contrast to other DDs that adopt the canonical six amphipathic α-helices arranged in a compact fold, the DAPk-DD was found to possess surprisingly low regular secondary structure content and an absence of a stable globular fold, as determined by circular dichroism (CD), NMR spectroscopy and a temperature-dependent fluorescence assay. Furthermore, we measured the in vitro interaction between extracellular-regulated kinase-2 (ERK2) and various recombinant DAPk-DD constructs. Despite the low level of structural order, the recombinant DAPk-DD retained the ability to interact with ERK2 in a 1∶1 ratio with a K d in the low micromolar range. Only the full-length DAPk-DD could bind ERK2, indicating that the apparent ‘D-motif’ located in the putative sixth helix of DAPk-DD is not sufficient for ERK2 recognition. CD analysis revealed that binding of DAPk-DD to ERK2 is not accompanied by a significant change in secondary structure. Taken together our data argue that the DAPk-DD, when expressed in isolation, does not adopt a classical DD fold, yet in this state retains the capacity to interact with at least one of its binding partners. The lack of a stable globular structure for the DAPk-DD may reflect either that its folding would be supported by interactions absent in our experimental set-up, or a limitation in the structural bioinformatics assignment of the three-dimensional structure.

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“…The solution structure of the Pelle-DD was also determined by another group using NMR (33). This structure very much resembled the Pelle-DD in the complex crystal structure with Tube.…”

Section: Mining Drosophila For Structural Details Of the Innate Immunmentioning

confidence: 99%

Intracellular TLR Signaling: A Structural Perspective on Human Disease

Lasker¹,

Nair

2006

The Journal of Immunology

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TLRs are crucial sensors of microbial infection. Maintaining structural integrity of TLR signaling components is essential for subsequent immunological protection. Alterations to the structure of these signaling molecules are often associated with profound clinical outcomes and susceptibility to various infectious diseases. These changes in structure are sometimes the result of a single nucleotide polymorphism (SNP). Numerous SNPs have been found in components of the TLR signaling pathway. Recently, the medical consequences and effects on TLR signaling of several of these SNPs have been elucidated. In addition, there have been numerous structures solved that are important to our understanding of the TLR signaling pathway at the molecular level. The scope of this review is to tie together current structural, biochemical, and genetic information of TLR signaling.

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Solution structure of the isolated Pelle death domain

Cited by 5 publications

References 45 publications

Assembly of Oligomeric Death Domain Complexes during Toll Receptor Signaling

Assembly of Oligomeric Death Domain Complexes during Toll Receptor Signaling

Structural and Functional Characterization of the Recombinant Death Domain from Death-Associated Protein Kinase

Intracellular TLR Signaling: A Structural Perspective on Human Disease

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