ABSTRACT
We determined a crystal structure of bovine Arp2/3 complex, an assembly of seven proteins that initiates actin polymerization in eukaryotic cells, at 2.0 angstrom resolution. Actin-related protein 2 (Arp2) and Arp3 are folded like actin, with distinctive surface features. Subunits ARPC2 p34 and ARPC4 p20 in the core of the complex associate through long carboxyl-terminal alpha helices and have similarly folded amino-terminal alpha/beta domains. ARPC1 p40 is a seven-blade beta propeller with an insertion that may associate with the side of an actin filament. ARPC3 p21 and ARPC5 p16 are globular alpha-helical subunits. We predict that WASp/Scar proteins activate Arp2/3 complex by bringing Arp2 into proximity with Arp3 for nucleation of a branch on the side of a preexisting actin filament.
Subject(s)
Actin Cytoskeleton/chemistry , Actin Cytoskeleton/metabolism , Actins/chemistry , Actins/metabolism , Cytoskeletal Proteins , Actin-Related Protein 2 , Actin-Related Protein 3 , Adenosine Triphosphate/metabolism , Animals , Cattle , Crystallography, X-Ray , Macromolecular Substances , Models, Biological , Models, Molecular , Muscle, Skeletal , Protein Structure, Quaternary , Protein Structure, Secondary , Protein Structure, Tertiary , Protein Subunits , Static Electricity , Thymus GlandABSTRACT
The Wiskott-Aldrich-syndrome protein (WASP) regulates polymerization of actin by the Arp2/3 complex. Here we show, using fluorescence anisotropy assays, that the carboxy-terminal WA domain of WASP binds to a single actin monomer with a Kd of 0.6 microM in an equilibrium with rapid exchange rates. Both WH-2 and CA sequences contribute to actin binding. A favourable DeltaH of -10 kcal mol(-1) drives binding. The WA domain binds to the Arp2/3 complex with a Kd of 0.9 microM; both the C and A sequences contribute to binding to the Arp2/3 complex. Wiskott-Aldrich-syndrome mutations in the WA domain that alter nucleation by the Arp2/3 complex over a tenfold range without affecting affinity for actin or the Arp2/3 complex indicate that there may be an activation step in the nucleation pathway. Actin filaments stimulate nucleation by producing a fivefold increase in the affinity of WASP-WA for the Arp2/3 complex.
Subject(s)
Actins/metabolism , Cell Movement/physiology , Cytoskeletal Proteins , Cytoskeleton/metabolism , Proteins/metabolism , Actin Cytoskeleton/metabolism , Actin-Related Protein 2 , Actin-Related Protein 3 , Animals , Binding Sites/physiology , Cytoskeleton/ultrastructure , Fluorescence Polarization/methods , Fluorescence Polarization/statistics & numerical data , Humans , Point Mutation/physiology , Protein Structure, Tertiary/physiology , Proteins/chemistry , Proteins/genetics , Rabbits , Rhodamines , Wiskott-Aldrich Syndrome Protein , Wiskott-Aldrich Syndrome Protein FamilyABSTRACT
A decamer of a beta-amino acid analogue of L-proline, (3R)-carboxy pyrrolidine (beta-proline), was synthesized from a readily available (R)-glycidol. It was found to possess a rigid secondary structure, as evidenced by its CD spectrum. The beta-proline decamer, however, failed to bind to profilin, whereas the corresponding alpha-L-proline decamer bound tightly to this protein.
Subject(s)
Contractile Proteins , Microfilament Proteins/metabolism , Oligopeptides/chemical synthesis , Pyrrolidines/chemical synthesis , Animals , Circular Dichroism , Molecular Structure , Oligopeptides/chemistry , Oligopeptides/metabolism , Profilins , Protein Conformation , Protein Structure, Secondary , Spectrometry, FluorescenceABSTRACT
STATEMENT OF PROBLEM: Many clinical situations are suitably treated with cantilevered implant-supported cemented fixed restorations. PURPOSE: This article details the indications for the restorative dentist to use a cantilever fixed prosthesis after insertion of ITI dental implants. CONCLUSION: The use of implants to support cantilevered fixed partial dentures has been successful in selected clinical situations.
Subject(s)
Dental Prosthesis, Implant-Supported , Denture, Partial, Fixed/statistics & numerical data , Alveolar Bone Loss/rehabilitation , Bone Transplantation , Dental Restoration Failure , Denture Design , Esthetics, Dental , Humans , Malocclusion/rehabilitation , OsseointegrationABSTRACT
Despite universal acceptance for retaining endodontically treated and filled roots for vertical support of removable prostheses, there is little evidence for the use of overdenture abutments with fixed prostheses. This article presents a review of clinical situations for using these teeth to support fixed partial dentures. These teeth provide vertical support during function and preserve alveolar bone levels.
Subject(s)
Dental Abutments , Denture Retention/methods , Denture, Overlay , Denture, Partial, Fixed , Humans , Tooth Root/surgery , Tooth, NonvitalABSTRACT
STATEMENT OF PROBLEM: Teeth and implants have different mobility patterns. Thus, it has been believed that implant-supported restorations should not be connected to natural teeth. However, this is not always the case. PURPOSE: This article presents guidelines for connection of restorative components when implant abutments and natural teeth are involved. METHODS: Methods of connection are discussed. CONCLUSION: This article presents options for splinting of prosthetic components.
Subject(s)
Dental Abutments , Dental Implants , Dental Prosthesis Retention/instrumentation , Jaw, Edentulous, Partially/rehabilitation , Periodontal Splints , Tooth Mobility/therapy , Clinical Protocols , Dental Implantation, Endosseous , Humans , Jaw, Edentulous, Partially/complications , Tooth Mobility/complicationsABSTRACT
Survivin is a 16.5 kDa protein that is expressed during the G2/M phase of the cell cycle and is hypothesized to inhibit a default apoptotic cascade initiated in mitosis. This inhibitory function is coupled to survivin's localization to the mitotic spindle. To begin to address the structural basis of survivin's function, we report the X-ray crystal structure of a recombinant form of full length survivin to 2.58 A resolution. Survivin consists of two defined domains including an N-terminal Zn2+-binding BIR domain linked to a 65 A amphipathic C-terminal alpha-helix. The crystal structure reveals an extensive dimerization interface along a hydrophobic surface on the BIR domain of each survivin monomer. A basic patch acting as a sulfate/phosphate-binding module, an acidic cluster projecting off the BIR domain, and a solvent-accessible hydrophobic surface residing on the C-terminal amphipathic helix, are suggestive of functional protein-protein interaction surfaces.
Subject(s)
Apoptosis , Microtubule-Associated Proteins , Proteins/chemistry , Proteins/metabolism , Amino Acid Sequence , Amino Acid Substitution/genetics , Binding Sites , Caspase 3 , Caspase Inhibitors , Caspases/metabolism , Crystallography, X-Ray , Dimerization , Humans , Inhibitor of Apoptosis Proteins , Models, Molecular , Molecular Sequence Data , Mutation/genetics , Neoplasm Proteins , Protein Binding , Protein Structure, Quaternary , Protein Structure, Secondary , Proteins/genetics , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Sequence Alignment , Structure-Activity Relationship , SurvivinABSTRACT
Most nucleated cells crawl about by extending a pseudopod that is driven by the polymerization of actin filaments in the cytoplasm behind the leading edge of the plasma membrane. These actin filaments are linked into a network by Y-branches, with the pointed end of each filament attached to the side of another filament and the rapidly growing barbed end facing forward. Because Arp2/3 complex nucleates actin polymerization and links the pointed end to the side of another filament in vitro, a dendritic nucleation model has been proposed in which Arp2/3 complex initiates filaments from the sides of older filaments. Here we report, by using a light microscopy assay, many new features of the mechanism. Branching occurs during, rather than after, nucleation by Arp2/3 complex activated by the Wiskott-Aldrich syndrome protein (WASP) or Scar protein; capping protein and profilin act synergistically with Arp2/3 complex to favour branched nucleation; phosphate release from aged actin filaments favours dissociation of Arp2/3 complex from the pointed ends of filaments; and branches created by Arp2/3 complex are relatively rigid. These properties result in the automatic assembly of the branched actin network after activation by proteins of the WASP/Scar family and favour the selective disassembly of proximal regions of the network.
Subject(s)
Actins/metabolism , Contractile Proteins , Cytoskeletal Proteins , Proteins/metabolism , Acanthamoeba , Actin Depolymerizing Factors , Actin-Related Protein 2 , Actin-Related Protein 3 , Animals , Cattle , Destrin , Humans , Microfilament Proteins/metabolism , Muscle, Skeletal/metabolism , Nucleotides/metabolism , Profilins , Rabbits , Recombinant Proteins/metabolism , Wiskott-Aldrich Syndrome Protein , Wiskott-Aldrich Syndrome Protein FamilyABSTRACT
Centric relation can be a confusing term because it continues to evolve in meaning. This article presents a discussion of the historical aspects of centric relation. Guidelines to decide when to use centric relation in clinical dentistry are included.
Subject(s)
Centric Relation , Mandible/anatomy & histology , Dental Occlusion, Centric , Guidelines as Topic , Humans , Mandibular Condyle/anatomy & histologyABSTRACT
Restorative problems with less than desirable implant placement can be challenging. A procedure is presented for the fabrication of a surgical guide stent that dictates placement of dental implants. This surgical guide can enhance implant placement in an efficient and acceptable manner so that final restorations can be properly contoured and esthetic.
Subject(s)
Dental Implantation, Endosseous/instrumentation , Dental Implantation, Endosseous/methods , Humans , Models, Dental , Patient Care Planning , StentsABSTRACT
We used biochemical fractionation, immunoassays and microscopy of live and fixed Acanthamoeba to determine how much profilin is bound to its known ligands: actin, membrane PIP(2), Arp2/3 complex and polyproline sequences. Virtually all profilin is soluble after gentle homogenization of cells. During gel filtration of extracts on Sephadex G75, approximately 60% of profilin chromatographs with monomeric actin, 40% is free and none voids with Arp2/3 complex or other large particles. Selective monoclonal antibodies confirm that most of the profilin is bound to actin: 65% in extract immunoadsorption assays and 74-89% by fluorescent antibody staining. Other than monomeric actin, no major profilin ligands are detected in crude extracts. Profilin-II labeled with rhodamine on cysteine at position 58 retains its affinity for actin, PIP(2) and poly-L-proline. When syringe-loaded into live cells, it distributes throughout the cytoplasm, is excluded from membrane-bounded organelles, and concentrates in lamellapodia and sites of endocytosis but not directly on the plasma membrane. Some profilin fluorescence appears punctate, but since no particulate profilin is detected biochemically, these spots may be soluble profilin between organelles that exclude profilin. The distribution of profilin in fixed human A431 cells is similar to that in amoebas. Our results show that the major pool of polymerizable actin monomers is complexed with profilin and spread throughout the cytoplasm.
Subject(s)
Acanthamoeba/metabolism , Actins/metabolism , Contractile Proteins , Microfilament Proteins/metabolism , Animals , Cell Line , Cytoplasm/metabolism , Fluorescent Antibody Technique , Microscopy, Video , Profilins , Tissue FixationABSTRACT
A procedure is presented that developed posterior lateral contacts in group function, despite existence of an anterior canine disclusion. This procedure allows a smooth transition into group function as the canine is subjected to natural attrition.
Subject(s)
Dental Occlusion, Centric , Dental Occlusion, Traumatic/prevention & control , Cuspid/physiology , Dental Articulators , Dental Restoration Wear , Denture, Partial, Fixed , Humans , Patient Care Planning , Temporomandibular Joint Disorders/prevention & control , Tooth Attrition/physiopathologyABSTRACT
A dental technique is described that blocks out enlarged gingival embrasures to eliminate distortion of an impression and the resultant cast for removable partial or fixed partial dentures. This procedure consists of injecting polyvinyl siloxane impression material in embrasures to form custom blockout wedges. This technique provides a simple method for clean, customized blockout of potentially damaging undercuts that can distort impressions and casts. The ultimate accuracy of the cast results in a precise definitive prosthesis.
Subject(s)
Dental Impression Technique , Diastema , Gingiva/anatomy & histology , Crowns , Dental Impression Technique/standards , Denture, Partial, Fixed , Denture, Partial, Removable , HumansABSTRACT
This article presents a procedure for refining margins of provisional implant crowns. This procedure uses an analog of the implant and abutment. This procedure allows functional and esthetic considerations such as emergence profiles, symmetry, diastemas, embrasures, size, contours, position, angulation, and relation to other teeth to be evaluated.
Subject(s)
Crowns , Dental Prosthesis Design , Dental Prosthesis, Implant-Supported , Cementation , Dental Abutments , Dental Implants , Esthetics, Dental , Humans , Surface PropertiesABSTRACT
The Arp2/3 complex, a stable assembly of two actin-related proteins (Arp2 and Arp3) with five other subunits, caps the pointed end of actin filaments and nucleates actin polymerization with low efficiency. WASp and Scar are two similar proteins that bind the p21 subunit of the Arp2/3 complex, but their effect on the nucleation activity of the complex was not known. We report that full-length, recombinant human Scar protein, as well as N-terminally truncated Scar proteins, enhance nucleation by the Arp2/3 complex. By themselves, these proteins either have no effect or inhibit actin polymerization. The actin monomer-binding W domain and the p21-binding A domain from the C terminus of Scar are both required to activate Arp2/3 complex. A proline-rich domain in the middle of Scar enhances the activity of the W and A domains. Preincubating Scar and Arp2/3 complex with actin filaments overcomes the initial lag in polymerization, suggesting that efficient nucleation by the Arp2/3 complex requires assembly on the side of a preexisting filament-a dendritic nucleation mechanism. The Arp2/3 complex with full-length Scar, Scar containing P, W, and A domains, or Scar containing W and A domains overcomes inhibition of nucleation by the actin monomer-binding protein profilin, giving active nucleation over a low background of spontaneous nucleation. These results show that Scar and, likely, related proteins, such as the Cdc42 targets WASp and N-WASp, are endogenous activators of actin polymerization by the Arp2/3 complex.
Subject(s)
Actins/metabolism , Cytoskeletal Proteins , Proteins/metabolism , Acanthamoeba/metabolism , Actin-Related Protein 2 , Actin-Related Protein 3 , Actins/chemistry , Animals , Humans , Kinetics , Macromolecular Substances , Models, Molecular , Muscle, Skeletal/metabolism , Protein Conformation , Proteins/chemistry , Protozoan Proteins/chemistry , Protozoan Proteins/metabolism , Rabbits , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Wiskott-Aldrich Syndrome Protein FamilyABSTRACT
This article describes a method for efficient and precise tooth preparation for fixed partial dentures. Diagnostic procedures are also reviewed.
Subject(s)
Denture, Partial, Fixed , Tooth Preparation, Prosthodontic/methods , Bicuspid , Dental Abutments , Humans , MolarABSTRACT
Displacement of soft tissue adjacent to an implant abutment is arduous. Currently, gingival retraction cord is used before making an impression for cement-retained implant restoration. This article presents a new impression system for a cementable abutment/implant. The advantages of a cement-retained implant crown are described. This system provides efficient and accurate impressions.
Subject(s)
Dental Abutments , Dental Implants , Dental Impression Technique , Dental Prosthesis Retention/methods , Gingiva , Cementation , Dental Implantation, Endosseous , HumansABSTRACT
This article presents a review of the evaluation of the temporary splint leading up to the present-day acid-etched composite, wire-reinforced, internal splint called the "A-splint." Nine general uses for the A-splint in the modern dental practice and some common problems are presented.
