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ABSTRACT: Here we describe how a F-probe incorporated into an endogenous protein by a chemical biology method revealed protein dynamics. By explicit determination of ligand-bound and unbound structures with X-ray crystallography, the quantitative comparison of the protein's dynamics in live cells and in vitro is presented. These results clearly demonstrated the greater conformational fluctuations of the intracellular protein, partially due to macromolecular crowding effects.
Chemical Communications 03/2013; 49(27):2801-3. · 6.17 Impact Factor
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Kyohei Arita,
Shin Isogai,
Takashi Oda,
Motoko Unoki,
Kazuya Sugita,
Naotaka Sekiyama,
Keiko Kuwata,
Ryuji Hamamoto, Hidehito Tochio,
Mamoru Sato,
Mariko Ariyoshi,
Masahiro Shirakawa
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ABSTRACT: Multiple covalent modifications on a histone tail are often recognized by linked histone reader modules. UHRF1 [ubiquitin-like, containing plant homeodomain (PHD) and really interesting new gene (RING) finger domains 1], an essential factor for maintenance of DNA methylation, contains linked two-histone reader modules, a tandem Tudor domain and a PHD finger, tethered by a 17-aa linker, and has been implicated to link histone modifications and DNA methylation. Here, we present the crystal structure of the linked histone reader modules of UHRF1 in complex with the amino-terminal tail of histone H3. Our structural and biochemical data provide the basis for combinatorial readout of unmodified Arg-2 (H3-R2) and methylated Lys-9 (H3-K9) by the tandem tudor domain and the PHD finger. The structure reveals that the intermodule linker plays an essential role in the formation of a histone H3-binding hole between the reader modules by making extended contacts with the tandem tudor domain. The histone H3 tail fits into the hole by adopting a compact fold harboring a central helix, which allows both of the reader modules to simultaneously recognize the modification states at H3-R2 and H3-K9. Our data also suggest that phosphorylation of a linker residue can modulate the relative position of the reader modules, thereby altering the histone H3-binding mode. This finding implies that the linker region plays a role as a functional switch of UHRF1 involved in multiple regulatory pathways such as maintenance of DNA methylation and transcriptional repression.
Proceedings of the National Academy of Sciences 07/2012; 109(32):12950-5. · 9.68 Impact Factor
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Masatoshi Nada,
Hidenori Ohnishi, Hidehito Tochio,
Zenichiro Kato,
Takeshi Kimura,
Kazuo Kubota,
Takahiro Yamamoto,
Yuji O Kamatari,
Naotaka Tsutsumi,
Masahiro Shirakawa,
Naomi Kondo
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ABSTRACT: Toll-like receptor (TLR) signaling is initiated by the binding of various adaptor proteins through ligand-induced oligomerization of the Toll/interleukin-1 receptor (TIR) domains of the TLRs. TLR2, which recognizes peptidoglycans, lipoproteins or lipopeptides derived from Gram-positive bacteria, is known to use the TIR domain-containing adaptor proteins myeloid differentiating factor 88 (MyD88) and MyD88 adaptor-like (Mal). Molecular analyses of the binding specificity of MyD88, Mal, and TLR2 are important for understanding the initial defenses mounted against Gram-positive bacterial infections such as Streptococcus pneumoniae. However, the detailed molecular mechanisms involved in the multiple interactions of these TIR domains remain unclear. Our study demonstrates that the TIR domain proteins MyD88, Mal, TLR1, and TLR2 directly bind to each other in vitro. We have also identified two binding interfaces of the MyD88 TIR domain for the TLR2 TIR domain. A residue at these interfaces has recently been found to be mutated in innate immune deficiency patients. These novel insights into the binding mode of TIR proteins will contribute to elucidation of the mechanisms underlying innate immune deficiency diseases, and to future structural studies of hetero-oligomeric TIR-TIR complexes.
Molecular Immunology 06/2012; 52(3-4):108-16. · 2.90 Impact Factor
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ABSTRACT: Ubiquitin is a post-translational modifier that is involved in cellular functions through its covalent attachment to target proteins. Ubiquitin can also be conjugated to itself at seven lysine residues and at its amino terminus to form eight linkage-specific polyubiquitin chains for individual cellular processes. The Lys63-linked polyubiquitin chain is recognized by tandem ubiquitin-interacting motifs (tUIMs) of Rap80 for the regulation of DNA repair. To understand the recognition mechanism between the Lys63-linked diubiquitin (K63-Ub(2)) and the tUIMs in solution, we determined the solution structure of the K63-Ub(2):tUIMs complex by using NOE restraints and RDC data derived from NMR spectroscopy. The structure showed that the tUIMs adopts a nearly straight and single continuous α-helix, and the two ubiquitin units of the K63-Ub(2) separately bind to each UIM motif. The interfaces are formed between Ile44-centered patches of the two ubiquitin units and the hydrophobic residues of the tUIMs. We also showed that the linker region between the two UIM motifs possesses a random-coil conformation in the free state, but undergoes the coil-to-helix transition upon complex formation, which simultaneously fixes the relative position of ubiquitin subunits. These data suggest that the relative position of ubiquitin subunits in the K63-Ub(2):tUIMs complex is essential for linkage-specific binding of Rap80 tUIMs.
Journal of Biomolecular NMR 02/2012; 52(4):339-50. · 3.61 Impact Factor
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ABSTRACT: Three-dimensional structures of proteins are often critical in understanding proteins functions. However, structures or states of proteins in cells undergo dynamical changes in response to interactions with other proteins and/or biological molecules. In addition, post-translational modification such as phosphorylation, methylation and ubiquitination can drastically change the structure and hence the properties of proteins. Therefore, to precisely correlate structure data of proteins with cell biology data, the structure information should be collected in living cells preferably at atomic level. In addition, as numerous biomolecules are packed into limited space, the concentration of macromolecules is substantially high in cells. Such crowded environment of the cell interior can markedly change proteins behavior, affecting biochemistry and biophysics of the proteins, which is so-called "Macromolecular Crowding Effect". To figure out protein behavior inside cells, which may be missed in in vitro studies, we are developing NMR and ESR methodologies to analyze protein structure and dynamics inside eukaryotic cultured cells. In this paper, in-cell NMR/ESR studies performed on HeLa cells and Xenopus oocytes are presented.
YAKUGAKU ZASSHI 01/2012; 132(2):185-93. · 0.37 Impact Factor
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Hidenori Ohnishi, Hidehito Tochio,
Zenichiro Kato,
Norio Kawamoto,
Takeshi Kimura,
Kazuo Kubota,
Takahiro Yamamoto,
Tatsuyoshi Funasaka,
Hiroshi Nakano,
Richard W Wong,
Masahiro Shirakawa,
Naomi Kondo
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ABSTRACT: MyD88, a Toll/interleukin-1 receptor homology (TIR) domain-containing adaptor protein, mediates signals from the Toll-like receptors (TLR) or IL-1/IL-18 receptors to downstream kinases. In MyD88-dependent TLR4 signaling, the function of MyD88 is enhanced by another TIR domain-containing adaptor, Mal/TIRAP, which brings MyD88 to the plasma membrane and promotes its interaction with the cytosolic region of TLR4. Hence, Mal is recognized as the "sorting adaptor" for MyD88. In this study, a direct interaction between MyD88-TIR and another membrane-sorting adaptor, TRAM/TICAM-2, was demonstrated in vitro. Cell-based assays including RNA interference experiments and TRAM deficient mice revealed that the interplay between MyD88 and TRAM in cells is important in mediating IL-18 signal transduction. Live cell imaging further demonstrated the co-localized accumulation of MyD88 and TRAM in the membrane regions in HEK293 cells. These findings suggest that TRAM serves as the sorting adaptor for MyD88 in IL-18 signaling, which then facilitates the signal transduction. The binding sites for TRAM are located in the TIR domain of MyD88 and actually overlap with the binding sites for Mal. MyD88, the multifunctional signaling adaptor that works together with most of the TLR members and with the IL-1/IL-18 receptors, can interact with two distinct sorting adaptors, TRAM and Mal, in a conserved manner in a distinct context.
PLoS ONE 01/2012; 7(6):e38423. · 4.09 Impact Factor
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Shin Isogai,
Daichi Morimoto,
Kyohei Arita,
Satoru Unzai,
Takeshi Tenno,
Jun Hasegawa,
Yu-shin Sou,
Masaaki Komatsu,
Keiji Tanaka,
Masahiro Shirakawa, Hidehito Tochio
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ABSTRACT: p62/SQSTM1/A170 is a multimodular protein that is found in ubiquitin-positive inclusions associated with neurodegenerative
diseases. Recent findings indicate that p62 mediates the interaction between ubiquitinated proteins and autophagosomes, leading
these proteins to be degraded via the autophagy-lysosomal pathway. This ubiquitin-mediated selective autophagy is thought
to begin with recognition of the ubiquitinated proteins by the C-terminal ubiquitin-associated (UBA) domain of p62. We present
here the crystal structure of the UBA domain of mouse p62 and the solution structure of its ubiquitin-bound form. The p62
UBA domain adopts a novel dimeric structure in crystals, which is distinctive from those of other UBA domains. NMR analyses
reveal that in solution the domain exists in equilibrium between the dimer and monomer forms, and binding ubiquitin shifts
the equilibrium toward the monomer to form a 1:1 complex between the UBA domain and ubiquitin. The dimer-to-monomer transition
is associated with a structural change of the very C-terminal end of the p62 UBA domain, although the UBA fold itself is essentially
maintained. Our data illustrate that dimerization and ubiquitin binding of the p62 UBA domain are incompatible with each other.
These observations reveal an autoinhibitory mechanism in the p62 UBA domain and suggest that autoinhibition plays a role in
the function of p62.
Journal of Biological Chemistry 09/2011; 286(36):31864-31874. · 4.77 Impact Factor
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Shin Isogai,
Daichi Morimoto,
Kyohei Arita,
Satoru Unzai,
Takeshi Tenno,
Jun Hasegawa,
Yu-shin Sou,
Masaaki Komatsu,
Keiji Tanaka,
Masahiro Shirakawa, Hidehito Tochio
[show abstract]
[hide abstract]
ABSTRACT: p62/SQSTM1/A170 is a multimodular protein that is found in ubiquitin-positive inclusions associated with neurodegenerative diseases. Recent findings indicate that p62 mediates the interaction between ubiquitinated proteins and autophagosomes, leading these proteins to be degraded via the autophagy-lysosomal pathway. This ubiquitin-mediated selective autophagy is thought to begin with recognition of the ubiquitinated proteins by the C-terminal ubiquitin-associated (UBA) domain of p62. We present here the crystal structure of the UBA domain of mouse p62 and the solution structure of its ubiquitin-bound form. The p62 UBA domain adopts a novel dimeric structure in crystals, which is distinctive from those of other UBA domains. NMR analyses reveal that in solution the domain exists in equilibrium between the dimer and monomer forms, and binding ubiquitin shifts the equilibrium toward the monomer to form a 1:1 complex between the UBA domain and ubiquitin. The dimer-to-monomer transition is associated with a structural change of the very C-terminal end of the p62 UBA domain, although the UBA fold itself is essentially maintained. Our data illustrate that dimerization and ubiquitin binding of the p62 UBA domain are incompatible with each other. These observations reveal an autoinhibitory mechanism in the p62 UBA domain and suggest that autoinhibition plays a role in the function of p62.
Journal of Biological Chemistry 06/2011; 286(36):31864-74. · 4.77 Impact Factor
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ABSTRACT: "In-cell nuclear magnetic resonance (NMR)" is a unique method for characterization of conformation, interaction and dynamics of proteins inside living cells at atomic level. Since the method was proposed by Dötch and co-workers in 2001 [1], its application had been limited to bacterial cells and oocytes of Xenopus laevis [2]. Recently, we reported a method for efficient delivery of (15)N-labeled proteins into human HeLa cells using cell-penetrating peptides, and measured high-resolution two-dimensional (1)H-(15)N correlation spectra of proteins in the cells. The in-cell NMR spectroscopy in human cells is capable of analyzing structures, interactions, dynamics and stability of proteins inside cells. Of its possible applications, we propose that in-cell NMR spectroscopy can be utilized as an effective step in protein-targeted drug development process, by demonstrating that interaction of FKBP12 with immunosuppressants administered extracellularly was successfully observed in living cells. This observation suggests that drug delivery and capability of target proteins inside cells for interaction with drugs can be investigated by in-cell NMR spectroscopy. More recently, an alternative way for intracellular delivery of labeled proteins for in-cell NMR was reported on 293F cells by Shimada and co-workers. Here, we review recent technical developments of in-cell NMR spectroscopy, and discuss potential usefulness for protein chemistry and drug screening process.
Current topics in medicinal chemistry 01/2011; 11(1):68-73. · 4.47 Impact Factor
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ABSTRACT: “In-cell nuclear magnetic resonance (NMR)” is a unique method for characterization of conformation, interaction and dynamics of proteins inside living cells at atomic level. Since the method was proposed by Dotch and co-workers in 2001(1), its application had been limited to bacterial cells and oocytes of Xenopus laevis(2). Recently, we reported the method for efficient delivery of 15N-labeled proteins into human HeLa cells by using cell-penetrating peptides, and measured high-resolution two-dimensional 1H-15N correlation spectra of proteins in the cells. The in-cell NMR spectroscopy in human cells is capable of analyzing structures, interactions, dynamics and stability of proteins inside cells. Of its possible applications, we propose that in-cell NMR spectroscopy can be utilized as an effective step in protein-targeted drug development process, by demonstrating that interaction of FKBP12 with immunosuppressants administered extracellularly was successfully observed in living cells. This observation suggests that drug delivery and capability of target proteins inside cells for interaction with drugs can be investigated by in-cell NMR spectroscopy. More recently, an alternative way for intracellular delivery of labeled proteins for in-cell NMR was reported on 293F cells by Shimada and co-workers. Here, we review recent technical development of in-cell NMR spectroscopy, and discuss potential usefulness for protein chemistry and drug screening process.
Current Topics in Medicinal Chemistry 12/2010; 11(1):68-73. · 4.17 Impact Factor
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ABSTRACT: Post-translational modification of proteins by covalent attachment of ubiquitin regulates diverse cellular events. A Lys48-linked polyubiquitin chain is formed via an isopeptide bond between Lys48 and the C-terminal Gly76 of different ubiquitin molecules. The chain is attached to a lysine residue of a substrate protein, which leads to proteolytic degradation of the protein by the 26S proteasome. In order to reveal the chain-length-dependent higher order structures of polyubiquitin chains, Lys48-linked polyubiquitin chains were synthesized enzymatically on a large scale and the chains were separated according to chain length by cation-exchange column chromatography. Subsequently, crystallization screening was performed using the hanging-drop vapour-diffusion method, from which crystals of tetraubiquitin, hexaubiquitin and octaubiquitin chains were obtained. The crystals of the tetraubiquitin and hexaubiquitin chains diffracted to 1.6 and 1.8 A resolution, respectively. The tetraubiquitin crystals belonged to space group C222(1), with unit-cell parameters a = 58.795, b = 76.966, c = 135.145 A. The hexaubiquitin crystals belonged to space group P2(1), with unit-cell parameters a = 51.248, b = 102.668, c = 51.161 A. Structural analysis by molecular replacement is in progress.
Acta Crystallographica Section F Structural Biology and Crystallization Communications 07/2010; 66(Pt 7):834-7. · 0.51 Impact Factor
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ABSTRACT: DEER (double electron-electron resonance) enables the observation of long-range dipole interactions (1.5-8 nm) between electron spin centers and has become a unique method for structural analysis of site-directed spin-labeled (SDSL) proteins. The method was applied to proteins inside eukaryotic cells, Xenopus laevis oocytes. DEER measurements of the oocytes, into which SDSL-ubiquitin derivates were injected, gave rise to interpretable signals and allowed us to perform in situ analyses of the interspin distances of the proteins.
Journal of the American Chemical Society 06/2010; 132(24):8228-9. · 9.91 Impact Factor
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Naoko Iwaya,
Yohta Kuwahara,
Yoshie Fujiwara,
Natsuko Goda,
Takeshi Tenno,
Kohei Akiyama,
Shogo Mase, Hidehito Tochio,
Takahisa Ikegami,
Masahiro Shirakawa,
Hidekazu Hiroaki
[show abstract]
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ABSTRACT: Katanin p60 (kp60), a microtubule-severing enzyme, plays a key role in cytoskeletal reorganization during various cellular
events in an ATP-dependent manner. We show that a single domain isolated from the N terminus of mouse katanin p60 (kp60-NTD)
binds to tubulin. The solution structure of kp60-NTD was determined by NMR. Although their sequence similarities were as low
as 20%, the structure of kp60-NTD revealed a striking similarity to those of the microtubule interacting and trafficking (MIT)
domains, which adopt anti-parallel three-stranded helix bundle. In particular, the arrangement of helices 2 and 3 is well
conserved between kp60-NTD and the MIT domain from Vps4, which is a homologous protein that promotes disassembly of the endosomal
sorting complexes required for transport III membrane skeleton complex. Mutation studies revealed that the positively charged
surface formed by helices 2 and 3 binds tubulin. This binding mode resembles the interaction between the MIT domain of Vps4
and Vps2/CHMP1a, a component of endosomal sorting complexes required for transport III. Our results show that both the molecular
architecture and the binding modes are conserved between two AAA-ATPases, kp60 and Vps4. A common mechanism is evolutionarily
conserved between two distinct cellular events, one that drives microtubule severing and the other involving membrane skeletal
reorganization.
Journal of Biological Chemistry 05/2010; 285(22):16822-16829. · 4.77 Impact Factor
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[show abstract]
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ABSTRACT: In eukaryotes, DNA replication is fired once in a single cell cycle before cell division starts to maintain stability of the
genome. This event is tightly controlled by a series of proteins. Cdt1 is one of the licensing factors and is involved in
recruiting replicative DNA helicase Mcm2–7 proteins into the pre-replicative complex together with Cdc6. In Cdt1, the C-terminal
region serves as a binding site for Mcm2–7 proteins, although the details of these interactions remain largely unknown. Here,
we report the structure of the region and the key residues for binding to Mcm proteins. We determined the solution structure
of the C-terminal fragment, residues 450–557, of mouse Cdt1 by NMR. The structure consists of a winged-helix domain and shows
unexpected similarity to those of the C-terminal domain of Cdc6 and the central fragment of Cdt1, thereby implying functional
and evolutionary relationships. Structure-based mutagenesis and an in vitro binding assay enabled us to pinpoint the region that interacts with Mcm proteins. Moreover, by performing in vitro binding and budding yeast viability experiments, we showed that ∼45 residues located in the N-terminal direction of the structural
region are equally crucial for recognizing Mcm proteins. Our data suggest the possibility that winged-helix domain plays a
role as a common module to interact with replicative helicase in the DNA replication-licensing process.
Journal of Biological Chemistry 05/2010; 285(21):15931-15940. · 4.77 Impact Factor
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ABSTRACT: Post-translational modification by small ubiquitin-like modifier (SUMO) provides an important regulatory mechanism in diverse cellular processes. Modification of SUMO has been shown to target proteins involved in systems ranging from DNA repair pathways to the ubiquitin-proteasome degradation system by the action of SUMO-targeted ubiquitin ligases (STUbLs). STUbLs recognize target proteins modified with a poly-SUMO chain through their SUMO-interacting motifs (SIMs). STUbLs are also associated with RENi family proteins, which commonly have two SUMO-like domains (SLD1 and SLD2) at their C terminus. We have determined the crystal structures of SLD2 of mouse RENi protein, Nip45, in a free form and in complex with a mouse E2 sumoylation enzyme, Ubc9. While Nip45 SLD2 shares a beta-grasp fold with SUMO, the SIM interaction surface conserved in SUMO paralogues does not exist in SLD2. Biochemical data indicates that neither tandem SLDs or SLD2 of Nip45 bind to either tandem SIMs from either mouse STUbL, RNF4 or to those from SUMO-binding proteins, whose interactions with SUMO have been well characterized. On the other hand, Nip45 SLD2 binds to Ubc9 in an almost identical manner to that of SUMO and thereby inhibits elongation of poly-SUMO chains. This finding highlights a possible role of the RENi proteins in the modulation of Ubc9-mediated poly-SUMO formation.
Proteins Structure Function and Bioinformatics 05/2010; 78(6):1491-502. · 3.39 Impact Factor
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Naoko Iwaya,
Yohta Kuwahara,
Yoshie Fujiwara,
Natsuko Goda,
Takeshi Tenno,
Kohei Akiyama,
Shogo Mase, Hidehito Tochio,
Takahisa Ikegami,
Masahiro Shirakawa,
Hidekazu Hiroaki
[show abstract]
[hide abstract]
ABSTRACT: Katanin p60 (kp60), a microtubule-severing enzyme, plays a key role in cytoskeletal reorganization during various cellular events in an ATP-dependent manner. We show that a single domain isolated from the N terminus of mouse katanin p60 (kp60-NTD) binds to tubulin. The solution structure of kp60-NTD was determined by NMR. Although their sequence similarities were as low as 20%, the structure of kp60-NTD revealed a striking similarity to those of the microtubule interacting and trafficking (MIT) domains, which adopt anti-parallel three-stranded helix bundle. In particular, the arrangement of helices 2 and 3 is well conserved between kp60-NTD and the MIT domain from Vps4, which is a homologous protein that promotes disassembly of the endosomal sorting complexes required for transport III membrane skeleton complex. Mutation studies revealed that the positively charged surface formed by helices 2 and 3 binds tubulin. This binding mode resembles the interaction between the MIT domain of Vps4 and Vps2/CHMP1a, a component of endosomal sorting complexes required for transport III. Our results show that both the molecular architecture and the binding modes are conserved between two AAA-ATPases, kp60 and Vps4. A common mechanism is evolutionarily conserved between two distinct cellular events, one that drives microtubule severing and the other involving membrane skeletal reorganization.
Journal of Biological Chemistry 03/2010; 285(22):16822-9. · 4.77 Impact Factor
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[show abstract]
[hide abstract]
ABSTRACT: In eukaryotes, DNA replication is fired once in a single cell cycle before cell division starts to maintain stability of the genome. This event is tightly controlled by a series of proteins. Cdt1 is one of the licensing factors and is involved in recruiting replicative DNA helicase Mcm2-7 proteins into the pre-replicative complex together with Cdc6. In Cdt1, the C-terminal region serves as a binding site for Mcm2-7 proteins, although the details of these interactions remain largely unknown. Here, we report the structure of the region and the key residues for binding to Mcm proteins. We determined the solution structure of the C-terminal fragment, residues 450-557, of mouse Cdt1 by NMR. The structure consists of a winged-helix domain and shows unexpected similarity to those of the C-terminal domain of Cdc6 and the central fragment of Cdt1, thereby implying functional and evolutionary relationships. Structure-based mutagenesis and an in vitro binding assay enabled us to pinpoint the region that interacts with Mcm proteins. Moreover, by performing in vitro binding and budding yeast viability experiments, we showed that approximately 45 residues located in the N-terminal direction of the structural region are equally crucial for recognizing Mcm proteins. Our data suggest the possibility that winged-helix domain plays a role as a common module to interact with replicative helicase in the DNA replication-licensing process.
Journal of Biological Chemistry 03/2010; 285(21):15931-40. · 4.77 Impact Factor
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[show abstract]
[hide abstract]
ABSTRACT: Myeloid differentiating factor 88 (MyD88) is one of a critical adaptor molecule in the Toll-like receptor (TLR) signaling pathway. The TIR domain of MyD88 serves as a protein-protein interaction module and interacts with other TIR-containing proteins such as Mal (MyD88 adaptor-like) and Toll-like receptor 4 to form signal initiation complexes. Here we report the (15)N, (13)C, and (1)H chemical shift assignments of the TIR domain of MyD88. The resonance assignments obtained in this work will contribute to the study of heteromeric TIR-TIR interactions between MyD88 and TIR-containing receptors or adaptors.
Biomolecular NMR Assignments 03/2010; 4(2):123-5. · 0.72 Impact Factor
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ABSTRACT: Poly-ADP-ribosylation is a unique post-translational modification that controls various nuclear events such as repair of DNA single-strand breaks. Recently, the protein containing the poly-ADP-ribose (pADPr)-binding zinc-finger (PBZ) domain was shown to be a novel AP endonuclease and involved in a cell cycle checkpoint. Here, we determined the three-dimensional structure of the PBZ domain from Drosophila melanogaster CG1218-PA using NMR spectroscopy. The domain folds into a C2H2-type zinc-finger structure in an S configuration, containing a characteristic loop between the zinc-coordinating cysteine and histidine residues. This is distinct from the structure of other C2H2-type zinc fingers. NMR signal changes that occur when pADPr binds to the PBZ domains from CG1218-PA and human checkpoint with FHA (forkhead-associated) and ring finger (CHFR) and mutagenesis suggest that a surface relatively well conserved among PBZ domains may serve as a major interface with pADPr.
Genes to Cells 02/2010; 15(2):101-10. · 2.68 Impact Factor
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ABSTRACT: Magnetic resonance imaging (MRI) is one of the most promising techniques for the non-invasive visualization of biomarkers and biologically relevant species, both in vivo and ex vivo. Although (1)H MRI with paramagnetic contrast agents, such as Gd(3+) complexes and iron oxide, is widely used, it often suffers from low contrast because of the large background signals caused by the abundant distribution of protons in biological samples. Here we report the use of supramolecular organic nanoparticles to detect specific proteins by (19)F-based MRI in an off/on mode. In NMR spectroscopy these designed probes are silent when aggregated, but in the presence of a target protein they disassemble to produce a sharp signal. This 'turn-on' response allowed us to visualize clearly the protein within live cells by (19)F MRI and construct an in-cell inhibitor assay. This recognition-driven disassembly of nanoprobes for a turn-on (19)F signal is unprecedented and may extend the use of (19)F MRI for specific protein imaging.
Nature Chemistry 10/2009; 1(7):557-61. · 20.52 Impact Factor
