Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q86TM3 (cage)
 29,987 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ferritin is a protein of 24 subunits which assemble into a shell with 432 point symmetry. It can be denatured reversibly in acidic guanidine hydrochloride, with the formation of poorly populated renaturation intermediates. In order to increase the accumulation of intermediates and to study the mechanism of ferritin renaturation, we analysed variants of the human ferritin H-chain altered at the N-terminus (delta(1-13)), near the 4-fold axis (Leu-169 --> Arg), the 3-fold axis (Asp-131 --> Ile + Glu-134 --> Phe) or the 2-fold axis (Ile-85 --> Cys). We also carried out specific chemical modifications of Cys-130 (near the 3-fold axis) and Cys-85 (near the 2-fold axis). Renaturation of the modified ferritins yielded assembly intermediates that differed in size and physical properties. Alterations of residues around the 2-, 4- and 3-fold axes produced subunit monomers, dimers and higher oligomers respectively. All these intermediates could be induced to assemble into ferritin 24-mers by concentrating them or by co-renaturing them with wild-type H-ferritin. The results support the hypothesis that the symmetric subunit dimers are the building blocks of ferritin assembly, and are consistent with a reassembly pathway involving the coalescence of dimers, probably around the 4-fold axis, followed by stepwise addition of dimers until the 24-mer cage is completed. In addition they show that assembly interactions are responsible for the large hysteresis of folding and unfolding plots. The implications of the studies for in vivo heteropolymer formation in vertebrates, which have two types of ferritin chain (H and L), are discussed.
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PMID:Effects of modifications near the 2-, 3- and 4-fold symmetry axes on human ferritin renaturation. 906 64

Thrombin-treated tumor cells induce a metastatic phenotype in experimental pulmonary murine metastasis. Thrombin binds to a unique protease-activated receptor (PAR-1) that requires N-terminal proteolytic cleavage for activation by its tethered end. A 14-mer thrombin receptor activation peptide (TRAP) of the tethered end induces the same cellular changes as thrombin. Four murine tumor cells (Lewis lung, CT26 colon CA, B16F10 melanoma, and CCL163 fibroblasts) contain PAR-1, as detected by reverse transcriptase-polymerase chain reaction (RT-PCR). B16F10 cells did not contain the two other thrombin receptors, PAR-3 and glycoprotein Ib. TRAP-treated B16F10 tumor cells enhance pulmonary metastasis 41- to 48-fold (n = 17). Thrombin-treated B16F10 cells transfected with full-length murine PAR-1 sense cDNA (S6, S7, S14, and S22) enhanced their adhesion to fibronectin 1.5- to 2.4-fold (n = 5, P <.04), whereas thrombin-treated wild-type cells do not. S6 (adhesion index, 1.5-fold) and S14 (index, 2.4-fold) when examined by RT-PCR and Northern analysis showed minimal expression of PAR-1 for S6 over wild-type and considerable expression for S14. Immunohistochemistry showed greater expression of PAR-1 for S14 compared with wild-type or empty-plasmid transfected cells. In vivo experiments with the thrombin-treated S14 transfectant showed a fivefold to sixfold increase in metastases compared with empty-plasmid transfected thrombin-treated naive cells or S6 cells (n = 20, P =.0001 to .02). Antisense had no effect on thrombin-stimulated tumor mass. Thus, PAR-1 ligation and expression enhances and regulates tumor metastasis.
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PMID:Protease-activated receptor 1 (PAR-1) is required and rate-limiting for thrombin-enhanced experimental pulmonary metastasis. 980 63

Ferritin is characterized by a highly conserved architecture that comprises 24 subunits assembled into a spherical cage with 432 symmetry. The only known exception is the dodecameric ferritin from Listeria innocua. The structure of Listeria ferritin has been determined to a resolution of 2.35 A by molecular replacement, using as a search model the structure of Dps from Escherichia coli. The Listeria 12-mer is endowed with 23 symmetry and displays the functionally relevant structural features of the ferritin 24-mer, namely the negatively charged channels along the three-fold symmetry axes that serve for iron entry into the cavity and a negatively charged internal cavity for iron deposition. The electron density map shows 12 iron ions on the inner surface of the hollow core, at the interface between monomers related by two-fold axes. Analysis of the nature and stereochemistry of the iron-binding ligands reveals strong similarities with known ferroxidase sites. The L. innocua ferritin site, however, is the first described so far that has ligands belonging to two different subunits and is not contained within a four-helix bundle.
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PMID:The dodecameric ferritin from Listeria innocua contains a novel intersubunit iron-binding site. 1062 25

Oxidative addition of aliphatic alcohols to (C(8)H(14))IrCl(PMe(3))(3) in benzene yields the cis-hydrido-alkoxo products mer-cis-HIr(OR)Cl(PMe(3))(3) (R = Me, Et, 1-pentyl, 2-propyl). The analogous hydroxo complex is prepared by oxidative addition of water in THF. The addition rate depends on the nature of the alcohol (methanol > 1-pentanol >> 2-propanol and methanol > water). The reaction is retarded in polar media but accelerated by protic cosolvents. Anionic ligand redistribution involving chloride and alkoxide (or hydroxide) competes with the oxidative addition reaction. A detailed kinetic study suggests that the 16-electron IrCl(PMe(3))(3) is the species undergoing the oxidative addition, and mer-cis-HIr(OR)Cl(PMe(3))(3) is the kinetic product. The reaction proceeds by a single-step nucleophilic attack of the metal on the O-H proton. Pi-donation by chloride stabilizes the transition state and governs the stereochemical course of the reaction. Protic solvent aggregation in the transition state in an apolar medium is suggested. mer-cis-HIr(OH)Cl(PEt(3))(3), obtained by water addition to IrCl(PEt(3))(3), was crystallographically characterized, showing an unusual hydrophobic cage around the hydride ligand.
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PMID:Oxidative addition of water and aliphatic alcohols by IrCl(trialkylphosphine)(3). 1223 60

The use of 1,3,5-triaminocyclohexane (tach) as a capping ligand in generating metal-cyanide cage clusters with accessible cavities is demonstrated. The precursor complexes [(tach)M(CN)(3)] (M = Cr, Fe, Co) are synthesized by methods similar to those employed in preparing the analogous 1,4,7-triazacyclononane (tacn) complexes. Along with [(tach)Fe(CN)(3)](1)(-), the latter two species are found to adopt low-spin electron configurations. Assembly reactions between [(tach)M(CN)(3)] (M = Fe, Co) and [M'(H(2)O)(6)](2+) (M' = Ni, Co) in aqueous solution afford the clusters [(tach)(4)(H(2)O)(12)Ni(4)Co(4)(CN)(12)](8+), [(tach)(4)(H(2)O)(12)Co(8)(CN)(12)](8+), and [(tach)(4)(H(2)O)(12)Ni(4)Fe(4)(CN)(12)](8+), each possessing a cubic arrangement of eight metal ions linked through edge-spanning cyanide bridges. This geometry is stabilized by hydrogen-bonding interactions between tach and water ligands through an intervening solvate water molecule or bromide counteranion. The magnetic behavior of the Ni(4)Fe(4) cluster indicates weak ferromagnetic coupling (J = 5.5 cm(-)(1)) between the Ni(II) and Fe(III) centers, leading to an S = 6 ground state. Solutions containing [(tach)Fe(CN)(3)] and a large excess of [Ni(H(2)O)(6)](2+) instead yield a trigonal pyramidal [(tach)(H(2)O)(15)Ni(3)Fe(CN)(3)](6+) cluster, in which even weaker ferromagnetic coupling (J = 1.2 cm(-)(1)) gives rise to an S = (7)/(2) ground state. Paralleling reactions previously performed with [(Me(3)tacn)Cr(CN)(3)], [(tach)Cr(CN)(3)] reacts with [Ni(H(2)O)(6)](2+) in aqueous solution to produce [(tach)(8)Cr(8)Ni(6)(CN)(24)](12+), featuring a structure based on a cube of Cr(III) ions with each face centered by a square planar [Ni(CN)(4)](2)(-) unit. The metal-cyanide cage differs somewhat from that of the analogous Me(3)tacn-ligated cluster, however, in that it is distorted via compression along a body diagonal of the cube. Additionally, the compact tach capping ligands do not hinder access to the sizable interior cavity of the molecule, permitting host-guest chemistry. Mass spectrometry experiments indicate a 1:1 association of the intact cluster with tetrahydrofuran (THF) in aqueous solution, and a crystal structure shows the THF molecule to be suspended in the middle of the cluster cavity. Addition of THF to an aqueous solution containing [(tach)Co(CN)(3)] and [Cu(H(2)O)(6)](2+) templates the formation of a closely related cluster, [(tach)(8)(H(2)O)(6)Cu(6)Co(8)(CN)(24) superset THF](12+), in which paramagnetic Cu(II) ions with square pyramidal coordination are situated on the face-centering sites. Reactions intended to produce the cubic [(tach)(4)(H(2)O)(12)Co(8)(CN)(12)](8+) cluster frequently led to an isomeric two-dimensional framework, [(tach)(H(2)O)(3)Co(2)(CN)(3)](2+), exhibiting mer rather than fac stereochemistry at the [Co(H(2)O)(3)](2+) subunits. Attempts to assemble larger edge-bridged cubic clusters by reacting [(tach)Cr(CN)(3)] with [Ni(cyclam)](2+) (cyclam = 1,4,8,11-tetraazacyclotetradecane) complexes instead generated extended one- or two-dimensional solids. The magnetic properties of one of these solids, two-dimensional [(tach)(2)(cyclam)(3)Ni(3)Cr(2)(CN)(6)]I(2), suggest metamagnetic behavior, with ferromagnetic intralayer coupling and weak antiferromagnetic interactions between layers.
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PMID:High-nuclearity metal-cyanide clusters: synthesis, magnetic properties, and inclusion behavior of open-cage species incorporating [(tach)M(CN)3] (M = Cr, Fe, Co) complexes. 1261 5

Reaction of the bis-bidentate bridging ligand L(3), in which two bidentate chelating 3(2-pyridyl)pyrazole units are separated by a 3,3'-biphenyl spacer, with Co(II) salts affords tetranuclear cage complexes of composition [Co(4)(L(3))(6)]X(8)(X =[BF(4)](-), [ClO(4)](-), [PF(6)](-) or I(-)) in which four 6-coordinate Co(II) ions in an approximately tetrahedral array are connected by six bis-bidentate bridging ligands, one spanning each of the six edges of the Co(4) tetrahedron. In every case, X-ray crystallography reveals that the 'apical' Co(II) ion has a fac tris-chelate geometry, whereas the other three Co(II) ions have mer tris-chelate geometries, resulting in (non-crystallographic)C(3) symmetry for the cages; that this structure is retained in solution is confirmed by (1)H NMR spectroscopy of the paramagnetic cages. In every case one of the anions is located inside the central cavity of the cage, with the remaining seven outside. We found no clear evidence for an anion-based templating effect. The cage superstructure is sufficiently large to leave gaps in the centres of the faces through which the internal and external anions can exchange. Variable-temperature (19)F NMR spectroscopy was used to investigate the dynamic behaviour of the cages with X =[BF(4)](-) and [PF(6)](-) in MeCN solution: in both cases two separate signals, corresponding to external and internal anions, are clear at 233 K which have coalesced to a single signal at room temperature. Analysis of the linewidth of the minor signal (for the internal anion) at various temperatures below coalescence gave an activation energy for anion exchange of ca. 50 kJ mol(-1) in each case, a figure which suggests that anion exchange can occur via a conformational rearrangement of the cage superstructure in solution rather than opening of the cavity by cleavage of metal-ligand bonds.
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PMID:Structures and anion-binding properties of M4L6 tetrahedral cage complexes with large central cavities. 1551 Feb 62

This paper describes a systematic study on the clathrate structure of (H+)(H2O)21 using tandem mass spectrometry, vibrational predissociation spectroscopy, Monte Carlo simulations, and density functional theory calculations. We produced (H+)(H2O)n from a continuous corona-discharged supersonic expansion and observed three anomalies simultaneously at the cluster temperature near 150 K, including (1) the peak at n=21 is more intense than its neighboring ions in the mass spectrum, (2) the size-dependent dissociation fractions show a distinct drop for the 21-mer, and (3) the infrared spectrum of (H+)(H2O)21 exhibits only a single feature at 3699 cm(-1), corresponding to the free-OH stretching of three-coordinated water molecules. Interestingly, the anomalies appear or disappear together with cluster temperature, indicating close correlation of these three observations. The observations, together with Monte Carlo simulations and density functional theory calculations, corroborate the notion for the formation of a distorted pentagonal dodecahedral (5(12)) cage with a H2O molecule in the cage and a H3O+ ion on the surface for this "magic number" water cluster ion. The dodecahedral cage melts at higher temperatures, as evidenced by the emergence of a free-OH stretching feature at 3717 cm(-1) for the two-coordinated water in (H+)(H2O)21 produced in a warmer molecular beam. Extension of this study to larger clusters strongly suggests that the experimentally observed isomer of (H+)(H2O)28 is most likely to consist of a distorted protonated pentakaidecahedral (5(12)6(3)) cage enclosing two neutral water molecules.
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PMID:Protonated clathrate cages enclosing neutral water molecules: (H+)(H2O)21 and (H+)(H2O)28. 1574 40

Phage libraries displaying linear or disulfide-constrained peptides often yield weak binders, upon screening against a target, and must be optimized to improve affinity. The disadvantages of libraries based on larger complex proteins, such as single chain antibodies, have stimulated interest in the development of smaller nonimmunoglobulin protein scaffolds. A promising candidate is the Trp cage motif, a 20-residue C-terminal sequence of exendin-4. Amino acid substitution within the Trp cage resulted in a 20-mer peptide recognized as an ultrafast cooperative folding miniprotein, with ideal characteristics for the discovery of small structured nonimmunoglobulin motifs having a stable tertiary structure. Although we were unable to display the Trp cage on M13 phage, successful display was achieved using the lytic T7 phage. Interestingly, mutations were observed at a frequency dependent on display valency. A Trp cage library designed with randomized amino acids at seven solvent-exposed positions was developed from 1.6 x 10(9) primary clones in T7Select10-3b. DNA sequencing of 109 library clones revealed 38% mutants and 16% truncations by TAG codons at randomized positions. Amino acid frequencies were largely within expected bounds and DIVAA analysis revealed that the library had an average diversity of 0.67. Utility of the library was demonstrated by identification of HPQ containing Trp cage miniproteins, which bound streptavidin, and AAADPYAQWLQSMGPHSGRPPPR, which bound to human bronchial epithelial cells. A high complexity library based on the Trp cage miniprotein has demonstrated potential for identifying novel cell and protein binding peptides that could be used for the delivery of therapeutic molecules or as target-specific therapeutic agents.
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PMID:The Trp cage motif as a scaffold for the display of a randomized peptide library on bacteriophage T7. 1726 74

The Escherichia coli HtrA protein is a periplasmic protease/chaperone that is upregulated under stress conditions. The protease and chaperone activities of HtrA eliminate or refold damaged and unfolded proteins in the bacterial periplasm that are generated upon stress conditions. In the absence of substrates, HtrA oligomerizes into a hexameric cage, but binding of misfolded proteins transforms the hexamers into bigger 12-mer and 24-mer cages that encapsulate the substrates for degradation or refolding. HtrA also undergoes partial degradation as a consequence of self-cleavage of the mature protein, producing short-HtrA protein (s-HtrA). The aim of this study was to examine the physiological role of this self-cleavage process. We found that the only requirement for self-cleavage of HtrA into s-HtrA in vitro was the hydrolysis of protein substrates. In fact, peptides resulting from the hydrolysis of the protein substrates were sufficient to induce autocleavage. However, the continuous presence of full-length substrate delayed the process. In addition, we observed that the hexameric cage structure is required for autocleavage and that s-HtrA accumulates only late in the degradation reaction. These results suggest that self-cleavage occurs when HtrA reassembles back into the resting hexameric structure and peptides resulting from substrate hydrolysis are allosterically stimulating the HtrA proteolytic activity. Our data support a model in which the physiological role of the self-cleavage process is to eliminate the excess of HtrA once the stress conditions cease.
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PMID:Characterization of the autocleavage process of the Escherichia coli HtrA protein: implications for its physiological role. 1910 20

Protein cages have been the focus of studies across multiple scientific disciplines. They have been used to deliver drugs, as templates for nanostructured materials, as substrates in the development of bio-orthogonal chemistry, and to restrict diffusion to study spatially confined reactions. Although their monomers fold into four-helix bundle structures, two cage proteins, DPS and BFR, self-assemble to form a 12-mer with tetrahedral symmetry and an octahedrally symmetric 24-mer, respectively. These monomers share strong similarities of both sequence and tertiary structure. However, they differ in the presence of a short additional helix. In BFR, the fifth helix is at the C-terminus and is positioned along the 4-fold symmetry axis, whereas with DPS, an extra helix helps to define the 2-fold axis in the cage and is located between the second and third helices in the monomer bundle. In an attempt to investigate if these short helices govern protein assembly, mutants were designed and produced that delete and swap these minidomains. All mutants form highly helical structures that unfold cooperatively as evidenced by thermal melting followed by circular dichroism. Dynamic light scattering, size exclusion chromatography, and sedimentation equilibrium experiments demonstrated that although many of the BFR mutants do not self-assemble and form lower-order complexes, many DPS mutants could form cages despite their unnatural design. Taken together, our data indicate that the BC helix is less important than the E helix for overall cage self-assembly, suggesting that dimerization may not play a role in nanostructure formation that is as key as previously assumed. Additionally, we found that fusing the minidomain from BFR onto DPS results in a mutant that assembles into a homogeneous population of a novel protein oligomer. This assembled cage while still formed from 12 subunits is larger in overall shape than that of the native protein.
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PMID:A helix swapping study of two protein cages. 1940 43


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