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1.
Carbohydr Polym ; 304: 120490, 2023 Mar 15.
Article in English | MEDLINE | ID: mdl-36641177

ABSTRACT

With an increase in the severity of environmental pollution caused by microbeads, the development of biodegradable microcapsules that can be applied in diverse fields has attracted significant attention. The degradation processes are directly related to biodegradable microcapsule creation with high stability and persistence. In this study, biodegradable microcapsules are synthesized via a complex coacervation approach using gelatin and alginate as the capsule main wall materials; additionally, enzyme-induced decomposition mechanisms are proposed by observing spectral changes in proton nuclear magnetic resonance (1H NMR) analyses. Additional analytical techniques confirm the chemical structure, morphology, and size distribution of the synthesized capsules; these uniform spherical microcapsules are 20-30 µm in size and possess a smooth surface. In addition to characterization, the microcapsules were exposed to targeted enzymes to investigate enzymatic effects using short-term and long-term degradation kinetics. Close inspection reveals that determination of the degradation rate constant of the major components in the capsule is feasible, and suggests two types of 4-stage degradation mechanisms that are enzyme-specific. These investigations demonstrate that capsule degradation can be explored in detail using 1H NMR spectroscopy to provide a viable strategy for monitoring degradation properties in the development of new biodegradable polymers.


Subject(s)
Alginates , Gelatin , Capsules/chemistry , Alginates/chemistry , Kinetics , Magnetic Resonance Spectroscopy
2.
Sci Rep ; 12(1): 1260, 2022 01 24.
Article in English | MEDLINE | ID: mdl-35075213

ABSTRACT

Middle East respiratory syndrome coronavirus (MERS-CoV) is a zoonotic virus, responsible for outbreaks of a severe respiratory illness in humans with a fatality rate of 30%. Currently, there are no vaccines or United States food and drug administration (FDA)-approved therapeutics for humans. The spike protein displayed on the surface of MERS-CoV functions in the attachment and fusion of virions to host cellular membranes and is the target of the host antibody response. Here, we provide a molecular method for neutralizing MERS-CoV through potent antibody-mediated targeting of the receptor-binding subdomain (RBD) of the spike protein. The structural characterization of the neutralizing antibody (KNIH90-F1) complexed with RBD using X-ray crystallography revealed three critical epitopes (D509, R511, and E513) in the RBD region of the spike protein. Further investigation of MERS-CoV mutants that escaped neutralization by the antibody supported the identification of these epitopes in the RBD region. The neutralizing activity of this antibody is solely provided by these specific molecular structures. This work should contribute to the development of vaccines or therapeutic antibodies for MERS-CoV.


Subject(s)
Antibodies, Monoclonal/chemistry , Antibodies, Neutralizing/chemistry , Antibodies, Viral/chemistry , Middle East Respiratory Syndrome Coronavirus/chemistry , Crystallography, X-Ray , Humans , Protein Domains
3.
Eur J Med Chem ; 226: 113819, 2021 Dec 15.
Article in English | MEDLINE | ID: mdl-34536669

ABSTRACT

The inhibition of glutaminyl cyclase (QC) may provide a promising strategy for the treatment of early Alzheimer's disease (AD) by reducing the amount of the toxic pyroform of ß-amyloid (AßΝ3pE) in the brains of AD patients. In this work, we identified potent QC inhibitors with subnanomolar IC50 values that were up to 290-fold higher than that of PQ912, which is currently being tested in Phase II clinical trials. Among the tested compounds, the cyclopentylmethyl derivative (214) exhibited the most potent in vitro activity (IC50 = 0.1 nM), while benzimidazole (227) showed the most promising in vivo efficacy, selectivity and druggable profile. 227 significantly reduced the concentration of pyroform Aß and total Aß in the brain of an AD animal model and improved the alternation behavior of mice during Y-maze tests. The crystal structure of human QC (hQC) in complex with 214 indicated tight binding at the active site, supporting that the specific inhibition of QC results in potent in vitro and in vivo activity. Considering the recent clinical success of donanemab, which targets AßΝ3pE, small molecule-based QC inhibitors may also provide potential therapeutic options for early-stage AD treatment.


Subject(s)
Alzheimer Disease/drug therapy , Aminoacyltransferases/antagonists & inhibitors , Benzimidazoles/pharmacology , Cyclopentanes/pharmacology , Enzyme Inhibitors/pharmacology , Neuroprotective Agents/pharmacology , Alzheimer Disease/metabolism , Aminoacyltransferases/metabolism , Amyloid beta-Peptides/antagonists & inhibitors , Amyloid beta-Peptides/metabolism , Animals , Benzimidazoles/chemical synthesis , Benzimidazoles/chemistry , Cyclopentanes/chemical synthesis , Cyclopentanes/chemistry , Dose-Response Relationship, Drug , Drug Discovery , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Humans , Male , Mice , Mice, Inbred ICR , Molecular Structure , Neuroprotective Agents/chemical synthesis , Neuroprotective Agents/chemistry , Structure-Activity Relationship
4.
Bioorg Med Chem Lett ; 29(16): 2275-2282, 2019 08 15.
Article in English | MEDLINE | ID: mdl-31253533

ABSTRACT

As a potential treatment of type 2 diabetes, a novel PPARγ non-TZD full agonist, compound 18 (BR102375) was identified from the original lead BR101549 by the SAR efforts of the labile metabolite control through bioisosteres approach. In vitro assessments of BR102375 demonstrated its activating potential of PPARγ comparable to Pioglitazone as well as the induction of related gene expressions. Further in vivo evaluation of BR102375 in diabetic rodent models successfully proved its glucose lowering effect as a potential antidiabetic agent, but the anticipated suppression of weight gain was not evident. The X-ray co-crystal analysis of BR102375-PPARγ LBD unexpectedly revealed binding modes totally different from those of BR101549, which was found, instead, closely resembled to those of TZD full agonists.


Subject(s)
Diabetes Mellitus, Type 2/drug therapy , Drug Discovery , Hypoglycemic Agents/pharmacology , Oxadiazoles/pharmacology , PPAR gamma/agonists , Crystallography, X-Ray , Diabetes Mellitus, Type 2/metabolism , Dose-Response Relationship, Drug , Humans , Hypoglycemic Agents/chemical synthesis , Hypoglycemic Agents/chemistry , Models, Molecular , Molecular Structure , Oxadiazoles/chemistry , PPAR gamma/metabolism , Structure-Activity Relationship
5.
Bioorg Med Chem Lett ; 29(4): 631-637, 2019 02 15.
Article in English | MEDLINE | ID: mdl-30594432

ABSTRACT

The new class of PPARgamma non-TZD agonist originally derived from the backbone of anti-hypertensive Fimasartan, BR101549, was identified as a potential lead for anti-diabetic drug development. The X-ray crystallography of BR101549 with PPARgamma ligand binding domain (LBD) revealed unique binding characteristics versus traditional TZD full agonists. The lead candidate, BR101549, has been found activating PPARgamma to the level of Pioglitazone in vitro and indeed has demonstrated its effects on blood glucose control in mouse proof-of-concept evaluation. The attempts to improve its metabolic stability profile through follow-up SAR including deuterium incorporation have been also described.


Subject(s)
Diabetes Mellitus, Type 2/drug therapy , Hypoglycemic Agents/therapeutic use , Oxadiazoles/therapeutic use , PPAR gamma/agonists , Pyrimidines/therapeutic use , Pyrimidinones/therapeutic use , 3T3-L1 Cells , Animals , Humans , Mice , Proof of Concept Study , Pyrimidinones/pharmacology , Structure-Activity Relationship
6.
Nat Commun ; 9(1): 4867, 2018 11 19.
Article in English | MEDLINE | ID: mdl-30451826

ABSTRACT

Osm1 and Frd1 are soluble fumarate reductases from yeast that are critical for allowing survival under anaerobic conditions. Although they maintain redox balance during anaerobiosis, the underlying mechanism is not understood. Here, we report the crystal structure of a eukaryotic soluble fumarate reductase, which is unique among soluble fumarate reductases as it lacks a heme domain. Structural and enzymatic analyses indicate that Osm1 has a specific binding pocket for flavin molecules, including FAD, FMN, and riboflavin, catalyzing their oxidation while reducing fumarate to succinate. Moreover, ER-resident Osm1 can transfer electrons from the Ero1 FAD cofactor to fumarate either by free FAD or by a direct interaction, allowing de novo disulfide bond formation in the absence of oxygen. We conclude that soluble eukaryotic fumarate reductases can maintain an oxidizing environment under anaerobic conditions, either by oxidizing cellular flavin cofactors or by a direct interaction with flavoenzymes such as Ero1.


Subject(s)
Flavin Mononucleotide/chemistry , Flavin-Adenine Dinucleotide/chemistry , Glycoproteins/chemistry , Oxidoreductases Acting on Sulfur Group Donors/chemistry , Riboflavin/chemistry , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae/enzymology , Succinate Dehydrogenase/chemistry , Anaerobiosis/genetics , Binding Sites , Cloning, Molecular , Crystallography, X-Ray , Escherichia coli/enzymology , Escherichia coli/genetics , Flavin Mononucleotide/metabolism , Flavin-Adenine Dinucleotide/metabolism , Gene Expression , Genetic Vectors/chemistry , Genetic Vectors/metabolism , Glycoproteins/genetics , Glycoproteins/metabolism , Kinetics , Molecular Docking Simulation , Oxidation-Reduction , Oxidoreductases Acting on Sulfur Group Donors/genetics , Oxidoreductases Acting on Sulfur Group Donors/metabolism , Protein Binding , Protein Conformation, alpha-Helical , Protein Conformation, beta-Strand , Protein Interaction Domains and Motifs , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Riboflavin/metabolism , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolism , Shewanella/enzymology , Shewanella/genetics , Substrate Specificity , Succinate Dehydrogenase/genetics , Succinate Dehydrogenase/metabolism , Triazines/chemistry , Triazines/metabolism
7.
Nucleic Acids Res ; 46(22): 11776-11788, 2018 12 14.
Article in English | MEDLINE | ID: mdl-30321390

ABSTRACT

Modification of chromatin and related transcription factors by histone deacetylases (HDACs) is one of the major strategies for controlling gene expression in eukaryotes. The HDAC domains of class IIa HDACs repress the respective target genes by interacting with the C-terminal region of the silencing mediator for retinoid and thyroid receptor (SMRT) repression domain 3 (SRD3c). However, latent catalytic activity suggests that their roles as deacetylases in gene regulation are unclear. Here, we found that two conserved GSI-containing motifs of SRD3c are critical for HDAC4 binding. Two SMRT peptides including these motifs commonly form a ß-hairpin structure in the cleft and block the catalytic entry site of HDAC4. They interact mainly with class IIa HDAC-specific residues of HDAC4 in a closed conformation. Structure-guided mutagenesis confirmed critical interactions between the SMRT peptides and HDAC4 and -5 as well as the contribution of the Arg1369 residue in the first motif for optimal binding to the two HDACs. These results indicate that SMRT binding does not activate the cryptic deacetylase activity of HDAC4 and explain how class IIa HDACs and the SMRT-HDAC3 complex are coordinated during gene regulation.


Subject(s)
Histone Deacetylases/metabolism , Nuclear Receptor Co-Repressor 2/metabolism , Repressor Proteins/metabolism , Amino Acid Motifs , Arginine/chemistry , Catalytic Domain , HEK293 Cells , Humans , Microscopy, Confocal , Mutagenesis , Mutagenesis, Site-Directed , Mutation , Peptides/chemistry , Protein Binding , Thermodynamics
8.
Am J Orthod Dentofacial Orthop ; 154(1): 108-114, 2018 Jul.
Article in English | MEDLINE | ID: mdl-29957307

ABSTRACT

INTRODUCTION: The 2-fold purpose of this study was to evaluate the effects of acid etching and calcium chloride immersion on removal torque and the bone-cutting ability of orthodontic mini-implants (OMIs). METHODS: For the removal torque part of the study, 3 types of OMIs (titanium alloy) were evaluated in a rabbit model: OMIs with acid surface etching with and without calcium chloride immersion (ECG and EG, respectively) and a control group (CG), in which the OMIs had an untreated, machined surface. We inserted 126 OMIs (42 OMIs per type) into both tibias of 21 male rabbits (5 months of age) with body weights of 3.0 to 3.5 kg. Removal torque was evaluated after 1, 4, and 7 weeks. To determine the OMIs' bone-cutting ability, total insertion time to place an OMI 6 mm into artificial bone was measured (6 OMIs per group). RESULTS: Removal torque values for the EG (3.97 ± 0.52 Ncm) and ECG (4.21 ± 0.44 Ncm) were statistically and significantly higher than those of the CG (3.02 ± 0.53 Ncm) 1 week after implantation (P <0.05). The ECG (6.54 ± 0.50, 6.61 ± 0.66 Ncm) showed the highest removal torque value followed by the EG (5.68 ± 0.58, 5.89 ± 0.70 Ncm) and CG (3.43 ± 0.62, 3.38 ± 0.54 Ncm) at 4 and 7 weeks after implantation (P <0.05). Removal torque did not change over time with the CG, but with the ECG and EG, it was significantly higher in weeks 4 and 7 than in week 1 (P <0.05). Total insertion time was significantly greater for the EG than for the ECG and CG (P <0.05). CONCLUSIONS: Treating OMIs with a calcium chloride solution improved the initial bone reaction by preventing contamination of the implant surface, and increasing the surface roughness of OMIs by acid etching enhanced their stability without decreasing the bone-cutting ability compared with OMIs without surface treatment.


Subject(s)
Acid Etching, Dental , Calcium Chloride/pharmacology , Dental Implants , Animals , Device Removal , Male , Orthodontic Anchorage Procedures , Rabbits , Surface Properties , Torque
9.
Int J Med Robot ; 13(4)2017 Dec.
Article in English | MEDLINE | ID: mdl-28371219

ABSTRACT

BACKGROUND: Despite its evident clinical benefits, single-incision laparoscopic surgery (SILS) imposes inherent limitations of collision between external arms and inadequate triangulation because multiple instruments are inserted through a single port at the same time. METHODS: A robot platform appropriate for SILS was developed wherein an elbowed instrument can be equipped to easily create surgical triangulation without the interference of robot arms. A novel joint mechanism for a surgical instrument actuated by a rigid link was designed for high torque transmission capability. RESULTS: The feasibility and effectiveness of the robot was checked through three kinds of preliminary tests: payload, block transfer, and ex vivo test. Measurements showed that the proposed robot has a payload capability >15 N with 7 mm diameter. CONCLUSIONS: The proposed robot is effective and appropriate for SILS, overcoming inadequate triangulation and improving workspace and traction force capability.


Subject(s)
Robotic Surgical Procedures/instrumentation , Robotic Surgical Procedures/methods , Robotics/instrumentation , Biomechanical Phenomena , Equipment Design , Hand/physiology , Humans , Laparoscopy/instrumentation , Stainless Steel
10.
FEBS Lett ; 591(2): 442-447, 2017 01.
Article in English | MEDLINE | ID: mdl-27990640

ABSTRACT

Stability of green fluorescent protein (GFP) is sometimes important for a proper practical application of this protein. Random mutagenesis and targeted mutagenesis have been used to create better-folded variants of GFP, including recently reported extra-superfolder GFP. Our aim was to determine the crystal structure of extra-superfolder GFP, which is more robustly folded and stable than GFP and superfolder GFP. The structural and structure-based mutagenesis analyses revealed that some of the mutations that created extra-superfolder GFP (F46L, E126K, N149K, and S208L) contribute to folding robustness by stabilizing extra-superfolder GFP with various noncovalent bonds.


Subject(s)
Green Fluorescent Proteins/chemistry , Green Fluorescent Proteins/metabolism , Amino Acid Sequence , Crystallography, X-Ray , Models, Molecular , Protein Stability , Protein Structure, Secondary , Structural Homology, Protein , Temperature
11.
Sci Rep ; 6: 31198, 2016 08 09.
Article in English | MEDLINE | ID: mdl-27502430

ABSTRACT

Caspase-2 is known to be involved in oxidative-stress mediated neuronal cell death. In this study, we demonstrated that rotenone-induced neuronal cell death is mediated by caspase-2 activation via PIDDosome formation. Our newly designed TAT-fused peptides, which contains wild-type helix number3 (H3) from RAIDD and PIDD, blocked the PIDDosome formation in vitro. Furthermore, peptides inhibited rotenone-induced caspase-2-dependent apoptosis in neuronal cells. These results suggest that PIDD- or RAIDD-targeted peptides might be effective at protecting against rotenone-induced neurotoxicity. Our peptides are novel neuronal cell apoptosis inhibitors that might serve as a prototype for development of drugs for the treatment of neurodegenerative diseases.


Subject(s)
Apoptosis/drug effects , CRADD Signaling Adaptor Protein/chemistry , Cell-Penetrating Peptides , Death Domain Receptor Signaling Adaptor Proteins/chemistry , Neurons/metabolism , Animals , Caspase 2/metabolism , Cell-Penetrating Peptides/chemistry , Cell-Penetrating Peptides/pharmacology , Cysteine Endopeptidases/metabolism , Humans , Neurons/cytology , PC12 Cells , Rats
12.
Appl Biochem Biotechnol ; 179(4): 650-8, 2016 Jun.
Article in English | MEDLINE | ID: mdl-26906118

ABSTRACT

TFCP2L1 is a transcription factor that facilitates establishment and maintenance of pluripotency in embryonic stem cells by forming a complex transcriptional network with other transcription factors (OCT4, SOX2, and NANOG). TFCP2L1 contains two distinct domains, the CP2-like domain at the N-terminus and the SAM-like domain at the C-terminus. In this study, we found that TFCP2L1 is hexamerized in solution via the C-terminal SAM-like domain. We also found that homo-oligomerization of SAM-like domain is dependent on the concentration of the proteins. Finally, we found that TFCP2L1 binds directly to DNA via the N-terminal CP2-like domain.


Subject(s)
Cell Differentiation/genetics , DNA-Binding Proteins/chemistry , Embryonic Stem Cells/metabolism , Repressor Proteins/chemistry , Animals , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Embryonic Stem Cells/chemistry , Mice , Nanog Homeobox Protein/genetics , Nanog Homeobox Protein/metabolism , Octamer Transcription Factor-3/genetics , Octamer Transcription Factor-3/metabolism , Protein Domains/genetics , Protein Multimerization , Repressor Proteins/genetics , Repressor Proteins/metabolism , SOXB1 Transcription Factors/genetics , SOXB1 Transcription Factors/metabolism
13.
Protein Pept Lett ; 23(1): 55-62, 2016.
Article in English | MEDLINE | ID: mdl-26548862

ABSTRACT

Toll-like receptors (TLRs) are evolutionarily conserved receptors with trimodular structure to respond to endogenous ligands and exogenous ligands from microbial pathogens. The highly conserved cytoplasmic C-terminal Toll/interleukin-1 receptor (TIR) domain of TLRs plays a crucial role in inflammatory reactions. In myeloid differentiation primary-response protein 88 (MyD88)- dependent signaling pathway, the interaction of TLRsTIR with cytosolic adaptor protein, MyD88TIR recruits IL-1R-associated kinases (IRAK) for subsequent activation of transcription factors nuclear factor kB (NF-kB) and activation protein 1 (AP-1) and other effector molecules. In the present investigation, TLR5TIR, TLR6TIR and MyD88TIR genes were subcloned and overexpressed in bacterium Escherichia coli strain BL- 21 (DE3). The purification and biochemical characterization of TLR5TIR and TLR6TIR&, and MyD88TIR proteins were also performed. The protein-protein interactions between TIR domains of TLR5 and TLR6 with MyD88, respectively, were evaluated in vitro at physiological pH and salt concentration. The in vitro reconstitution results showed that under physiological pH and salt concentration, MyD88TIR interacted with TLR5TIR, and did not interact with TLR6TIR protein. Both TIR domain-containing TLR5 and TLR6 proteins were prone to aggregation in a temperature-dependent manner at room temperature. At normal physiological pH and salt concentration, with the addition of binding partner MyD88TIR to TLR5/6TIR, time-dependent aggregation was not observed in both TLRsTIR at both room temperature and 4 ºC for 2 d, influencing the solubility of TLR5/6TIR. Moreover, TLR5TIR alone exhibited increase in solubility of the protein with increase in the salt concentration of the buffered solution from 0.025 M to 1.25 M at room temperature.


Subject(s)
Escherichia coli/genetics , Myeloid Differentiation Factor 88/chemistry , Toll-Like Receptor 5/chemistry , Toll-Like Receptor 6/chemistry , Escherichia coli/metabolism , Humans , Hydrogen-Ion Concentration , In Vitro Techniques , Models, Molecular , Myeloid Differentiation Factor 88/genetics , Myeloid Differentiation Factor 88/metabolism , Protein Aggregates , Protein Binding , Protein Structure, Tertiary , Salinity , Temperature , Toll-Like Receptor 5/genetics , Toll-Like Receptor 5/metabolism , Toll-Like Receptor 6/genetics , Toll-Like Receptor 6/metabolism
15.
Sci Rep ; 5: 9847, 2015 Jun 03.
Article in English | MEDLINE | ID: mdl-26038885

ABSTRACT

Apoptosis repressor with caspase recruiting domain (ARC) is a multifunctional inhibitor of apoptosis that is unusually over-expressed or activated in various cancers and in the state of the pulmonary hypertension. Therefore, ARC might be an optimal target for therapeutic intervention. Human ARC is composed of two distinct domains, N-terminal caspase recruiting domain (CARD) and C-terminal P/E (proline and glutamic acid) rich domain. ARC inhibits the extrinsic apoptosis pathway by interfering with DISC formation. ARC CARD directly interacts with the death domains (DDs) of Fas and FADD, as well as with the death effector domains (DEDs) of procaspase-8. Here, we report the first crystal structure of the CARD domain of ARC at a resolution of 2.4 Å. Our structure was a dimer with novel homo-dimerization interfaces that might be critical to its inhibitory function. Interestingly, ARC did not exhibit a typical death domain fold. The sixth helix (H6), which was detected at the typical death domain fold, was not detected in the structure of ARC, indicating that H6 may be dispensable for the function of the death domain superfamily.


Subject(s)
Apoptosis Regulatory Proteins/chemistry , Apoptosis , CARD Signaling Adaptor Proteins/chemistry , Models, Molecular , Protein Conformation , Amino Acid Sequence , Apoptosis Regulatory Proteins/metabolism , Binding Sites , CARD Signaling Adaptor Proteins/metabolism , Conserved Sequence , Molecular Sequence Data , Protein Binding , Protein Multimerization , Sequence Alignment , Structure-Activity Relationship
16.
Biochem Biophys Res Commun ; 460(4): 957-63, 2015 May 15.
Article in English | MEDLINE | ID: mdl-25839653

ABSTRACT

Inflammatory caspases, such as caspase-1, which is critical for the innate immune response, are activated upon the formation of a molecular complex called the inflammasome. The inflammasome is composed of three proteins, the Nod-like receptor (NLRP, NLRC or AIM2), apoptosis associated speck-loke protein containing a caspase-recruitment domain (ASC), and caspase-1. ASC is an adaptor molecule that contains an N-terminal PYD domain and a C-terminal CARD domain for interaction with other proteins. Upon activation, the N-terminal PYD of ASC homotypically interacts with the PYD domain of the Nod-like receptor, while its C-terminal CARD homotypically interacts with the CARD domain of caspase-1. PYD only protein 1 (POP1) negatively regulates inflammatory response by blocking the formation of the inflammasome. POP1 directly binds to ASC via a PYD:PYD interaction, thereby preventing ASC recruitment to Nod-like receptor NLRPs. POP1-mediated regulation of inflammation is of great biological importance. Here, we report the crystal structure of human POP1 and speculate about the inhibitory mechanism of POP1-mediated inflammasome formation based on the current structure.


Subject(s)
Apoptosis Regulatory Proteins/chemistry , Ribonucleoproteins/chemistry , Amino Acid Sequence , Caspase 1/chemistry , Crystallography, X-Ray , Humans , Models, Molecular , Molecular Sequence Data , Protein Conformation , Sequence Homology, Amino Acid
17.
Korean J Orthod ; 44(6): 294-303, 2014 Nov.
Article in English | MEDLINE | ID: mdl-25473645

ABSTRACT

OBJECTIVE: The purpose of this study was to examine whether orthodontic treatment experience affects the individual's perception of smile esthetics and to evaluate differences among orthodontically treated laypersons, non-treated laypersons, and dentists by using computerized image alterations. METHODS: A photograph of a woman's smile was digitally altered using a software image editing program. The alterations involved gingival margin height, crown width and length, incisal plane canting, and dental midline of the maxillary anterior teeth. Three groups of raters (orthodontically treated laypersons, non-treated laypersons, and dentists) evaluated the original and altered images using a visual analog scale. RESULTS: The threshold for detecting changes in maxillary central incisor gingival margin height among laypersons was 1.5 mm; the threshold of dentists, who were more perceptive, was 1.0 mm. For maxillary lateral incisor crown width and height, the threshold of all three groups was 3.0 mm. Canting of the incisal plane was perceived when the canting was 3.0 mm among non-treated laypersons, 2.0 mm among treated laypersons, and 1.0 mm among dentists. Non-treated laypersons could not perceive dental midline shifts; however, treated laypersons and dentists perceived them when the shift was ≥ 3.0 mm. CONCLUSIONS: Laypersons with and without orthodontic treatment experience and dentists have different perceptions of smile esthetics. Orthodontically treated laypersons were more critical than non-treated laypersons regarding incisal plane canting and dental midline shifts. Based on these findings, it is suggested that orthodontic treatment experience improved the esthetic perceptions of laypersons.

18.
PLoS One ; 9(9): e107005, 2014.
Article in English | MEDLINE | ID: mdl-25192068

ABSTRACT

Transglutaminase2 (TG2) is a multi-functional protein involved in various cellular processes, including apoptosis, differentiation, wound healing, and angiogenesis. The malfunction of TG2 causes many human disease including inflammatory disease, celiac disease, neurodegenerative diseases, tissue fibrosis, and cancers. Protein cross-linking activity, which is representative of TG2, is activated by calcium ions and suppressed by GTP. Here, we elucidated the structure of TG2 in complex with its endogenous inhibitor, GTP. Our structure showed why GTP is the optimal nucleotide for interacting with and inhibiting TG2. In addition, sequence comparison provided information describing the evolutionary scenario of GTP usage for controlling the activity of TG2.


Subject(s)
GTP-Binding Proteins/chemistry , GTP-Binding Proteins/metabolism , Guanosine Triphosphate/metabolism , Protein Interaction Domains and Motifs/genetics , Transglutaminases/chemistry , Transglutaminases/metabolism , Amino Acid Sequence , Binding Sites/genetics , Crystallography, X-Ray , Evolution, Molecular , Guanosine Triphosphate/chemistry , Humans , Models, Molecular , Molecular Sequence Data , Protein Binding , Protein Glutamine gamma Glutamyltransferase 2 , Protein Structure, Quaternary , Protein Structure, Secondary , Sequence Homology, Amino Acid
19.
Acta Crystallogr F Struct Biol Commun ; 70(Pt 8): 1053-5, 2014 Aug.
Article in English | MEDLINE | ID: mdl-25084380

ABSTRACT

Toll-like receptor (TLR) proteins have been identified and shown to play a role in the innate immune response. TLR6 associated with TLR2 can recognize diacylated lipoprotein. In this study, the human TLR6 TIR domain corresponding to amino acids 640-796 was overexpressed in Escherichia coli using engineered C-terminal His tags. The TLR6 TIR domain was then purified to homogeneity and crystallized at 20°C. Finally, X-ray diffraction data were collected to a resolution of 2.2 Šfrom a crystal belonging to space group C2, with unit-cell parameters a = 127.60, b = 44.20, c = 75.72 Å, ß = 118.89°


Subject(s)
Toll-Like Receptor 6/chemistry , Base Sequence , Crystallography, X-Ray , DNA Primers , Humans , Polymerase Chain Reaction , Protein Conformation
20.
Biochemistry ; 53(33): 5424-31, 2014 Aug 26.
Article in English | MEDLINE | ID: mdl-25119434

ABSTRACT

Receptor interaction protein kinase 1 (RIP1) is a molecular cell-fate switch. RIP1, together with Fas-associated protein with death domain (FADD) and caspase-8, forms the RIPoptosome that activates apoptosis. RIP1 also associates with RIP3 to form the necrosome that triggers necroptosis. The RIPoptosome assembles through interactions between the death domains (DDs) of RIP1 and FADD and between death effector domains (DEDs) of FADD and caspase-8. In this study, we analyzed the overall structure of the RIP1 DD/FADD DD complex, the core of the RIPoptosome, by negative-stain electron microscopy and modeling. The results show that RIP1 DD and FADD DD form a stable complex in vitro similar to the previously described Fas DD/FADD DD complex, suggesting that the RIPoptosome and the Fas death-inducing signaling complex share a common assembly mechanism. Both complexes adopt a helical conformation that requires type I, II, and III interactions between the death domains.


Subject(s)
Caspase 8/chemistry , Fas-Associated Death Domain Protein/chemistry , GTPase-Activating Proteins/chemistry , Multiprotein Complexes/chemistry , Amino Acid Sequence , Caspase 8/metabolism , Fas-Associated Death Domain Protein/genetics , Fas-Associated Death Domain Protein/metabolism , GTPase-Activating Proteins/genetics , GTPase-Activating Proteins/metabolism , Microscopy, Electron/methods , Models, Molecular , Molecular Sequence Data , Mutagenesis, Site-Directed , Protein Conformation , Protein Structure, Tertiary , Sequence Homology, Amino Acid , fas Receptor/chemistry
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