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1.
Acta Crystallogr D Biol Crystallogr ; 57(Pt 12): 1829-35, 2001 Dec.
Article in English | MEDLINE | ID: mdl-11717495

ABSTRACT

The monoclinic crystal form of human peroxiredoxin 5 with eight molecules in the asymmetric unit was obtained under exactly the same conditions as the tetragonal form with one molecule in the asymmetric unit, except that the latter was briefly cryosoaked with halide for derivatization. A merohedral twinning was observed, which is rather unusual in the monoclinic system and only possible with particular unit-cell dimensions. After detwinning the native and a mercury derivative, the structure was solved by the SIR method with the help of the non-crystallographic symmetry. The packing of the monoclinic and tetragonal forms are compared, with special attention to the role of bromide ions in the change of space group after crystallization. The availability of nine (eight monoclinic plus one tetragonal) independent molecules allows an analysis of the mobility. The two Cys residues implicated in the peroxide-reduction mechanism are located in rigid regions but are covered by mobile loops.


Subject(s)
Peroxidases/chemistry , Crystallization , Crystallography, X-Ray , Humans , Models, Molecular , Peroxiredoxins , Protein Conformation , Quality Control
2.
Org Lett ; 3(20): 3067-70, 2001 Oct 04.
Article in English | MEDLINE | ID: mdl-11573996

ABSTRACT

[reaction: see text] Photolysis of oxaziridines a or nitrones b provides a convenient synthetic route to fused bicyclic lactams c adequately substituted on both cycles A and B as scaffolds for mimicking conformationally constrained beta-turn peptides as in the tripeptide RGD signaling motif of fibronectin.


Subject(s)
Bridged Bicyclo Compounds/chemical synthesis , Indoles/chemistry , Lactams/chemical synthesis , Nitrogen Oxides/chemistry , Oligopeptides/chemistry , Crystallography, X-Ray , Molecular Mimicry , Molecular Structure , Photochemistry
3.
J Mol Biol ; 311(4): 751-9, 2001 Aug 24.
Article in English | MEDLINE | ID: mdl-11518528

ABSTRACT

The peroxiredoxins define an emerging family of peroxidases able to reduce hydrogen peroxide and alkyl hydroperoxides with the use of reducing equivalents derived from thiol-containing donor molecules such as thioredoxin, glutathione, trypanothione and AhpF. Peroxiredoxins have been identified in prokaryotes as well as in eukaryotes. Peroxiredoxin 5 (PRDX5) is a novel type of mammalian thioredoxin peroxidase widely expressed in tissues and located cellularly to mitochondria, peroxisomes and cytosol. Functionally, PRDX5 has been implicated in antioxidant protective mechanisms as well as in signal transduction in cells. We report here the 1.5 A resolution crystal structure of human PRDX5 in its reduced form. The crystal structure reveals that PRDX5 presents a thioredoxin-like domain. Interestingly, the crystal structure shows also that PRDX5 does not form a dimer like other mammalian members of the peroxiredoxin family. In the reduced form of PRDX5, Cys47 and Cys151 are distant of 13.8 A although these two cysteine residues are thought to be involved in peroxide reductase activity by forming an intramolecular disulfide intermediate in the oxidized enzyme. These data suggest that the enzyme would necessitate a conformational change to form a disulfide bond between catalytic Cys47 and Cys151 upon oxidation according to proposed peroxide reduction mechanisms. Moreover, the presence of a benzoate ion, a hydroxyl radical scavenger, was noted close to the active-site pocket. The possible role of benzoate in the antioxidant activity of PRDX5 is discussed.


Subject(s)
Peroxidases/chemistry , Amino Acid Sequence , Animals , Binding Sites , Crystallography, X-Ray , Cysteine/metabolism , Disulfides/metabolism , Humans , Models, Molecular , Molecular Sequence Data , Molecular Weight , Oxidation-Reduction , Peroxidases/metabolism , Peroxiredoxins , Protein Structure, Secondary , Protein Structure, Tertiary , Rats , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Sequence Alignment
4.
J Med Chem ; 43(17): 3274-82, 2000 Aug 24.
Article in English | MEDLINE | ID: mdl-10966746

ABSTRACT

Cholic acid-derived 1,2,4,5-tetraoxanes were synthesized in order to explore the influence of steroid carrier on its antimalarial and antiproliferative activity in vitro. Starting with chiral ketones, cis and trans series of diastereomeric tetraoxanes were obtained, and the cis series was found to be approximately 2 times as active as the trans against Plasmodium falciparum D6 and W2 clones. The same tendency was observed against human melanoma (Fem-X) and human cervix carcinoma (HeLa) cell lines. The amide C(24) termini, for the first time introduced into the carrier molecule of a tetraoxane pharmacophore, significantly enhanced both antimalarial and antiproliferative activity, as compared to the corresponding methyl esters, with cis-bis(N-propylamide) being most efficient against the chloroquine-susceptible D6 clone (IC(50) = 9.29 nM). cis- and trans-bis(N-propylamides) were also screened against PBMC, and PHA-stimulated PBMC, showing a cytotoxicity/antimalarial potency ratio of 1/10 000.


Subject(s)
Antimalarials/chemical synthesis , Antineoplastic Agents/chemical synthesis , Cholic Acids/chemical synthesis , Spiro Compounds/chemical synthesis , Animals , Antimalarials/chemistry , Antimalarials/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Cholic Acids/chemistry , Cholic Acids/pharmacology , Drug Screening Assays, Antitumor , Humans , Magnetic Resonance Spectroscopy , Plasmodium falciparum/drug effects , Spiro Compounds/chemistry , Spiro Compounds/pharmacology , Stereoisomerism , Structure-Activity Relationship , Tumor Cells, Cultured
5.
Protein Sci ; 8(10): 2194-204, 1999 Oct.
Article in English | MEDLINE | ID: mdl-10548066

ABSTRACT

Several crystal structures of parvalbumin (Parv), a typical EF-hand protein, have been reported so far for different species with the best resolution achieving 1.5 A. Using a crystal grown under microgravity conditions, cryotechniques (100 K), and synchrotron radiation, it has now been possible to determine the crystal structure of the fully Ca2+-loaded form of pike (component pI 4.10) Parv.Ca2 at atomic resolution (0.91 A). The availability of such a high quality structure offers the opportunity to contribute to the definition of the validation tools useful for the refinement of protein crystal structures determined to lower resolution. Besides a better definition of most of the elements in the protein three-dimensional structure than in previous studies, the high accuracy thus achieved allows the detection of well-defined alternate conformations, which are observed for 16 residues out of 107 in total. Among them, six occupy an internal position within the hydrophobic core and converge toward two small buried cavities with a total volume of about 60 A3. There is no indication of any water molecule present in these cavities. It is probable that at temperatures of physiological conditions there is a dynamic interconversion between these alternate conformations in an energy-barrier dependent manner. Such motions for which the amplitudes are provided by the present study will be associated with a time-dependent remodeling of the void internal space as part of a slow dynamics regime (millisecond timescales) of the parvalbumin molecule. The relevance of such internal dynamics to function is discussed.


Subject(s)
Parvalbumins/chemistry , Binding Sites , Calcium/metabolism , Cold Temperature , Crystallography, X-Ray , Models, Molecular , Parvalbumins/metabolism , Protein Conformation
6.
Acta Crystallogr D Biol Crystallogr ; 55(Pt 2): 430-5, 1999 Feb.
Article in English | MEDLINE | ID: mdl-10089351

ABSTRACT

Until now, wild-type bacteriophage lambda lysozyme had been impossible to crystallize. This difficulty could be overcome by the replacement of the four tryptophan residues by aza-tryptophans. Analysis of the intermolecular and intramolecular contacts in this modification allows understanding of the differences in behaviour between the native and modified molecules. Furthermore, this mutation was very useful for the creation of new heavy-atom binding sites and for the solution of the non-crystallographic symmetry, which is extremely important for phase improvement. This procedure seems to be generally applicable, at least in the search for new possibilities for heavy-atom binding sites.


Subject(s)
Aza Compounds/chemistry , Bacteriophage lambda/enzymology , Muramidase/chemistry , Tryptophan/chemistry , Crystallization , Crystallography, X-Ray , Hydrogen Bonding , Protein Conformation
7.
J Mol Biol ; 276(1): 151-64, 1998 Feb 13.
Article in English | MEDLINE | ID: mdl-9514719

ABSTRACT

Like other lysozymes, the bacteriophage lambda lysozyme is involved in the digestion of bacterial walls. This enzyme is remarkable in that its mechanism of action is different from the classical lysozyme's mechanism. From the point of view of protein evolution, it shows features of lysozymes from different classes. The crystal structure of the enzyme in which all tryptophan residues have been replaced by aza-tryptophan has been solved by X-ray crystallography at 2.3 A using a combination of multiple isomorphous replacement, non-crystallographic symmetry averaging and density modification techniques. There are three molecules in the asymmetric unit. The characteristic structural elements of lysozymes are conserved: each molecule is organized in two domains connected by a helix and the essential catalytic residue (Glu19) is located in the depth of a cleft between the two domains. This cleft shows an open conformation in two of the independent molecules, while access to the cavity is much more restricted in the last one. A structural alignment with T4 lysozyme and hen egg white lysozyme allows us to superpose about 60 C alpha atoms with a rms distance close to 2 A. The best alignments concern the helix preceding the catalytic residue, some parts of the beta sheets and the helix joining the two domains. The results of sequence alignments with the V and C lysozymes, in which weak local similarities had been detected, are compared with the structural results.


Subject(s)
Bacteriophage lambda/enzymology , Muramidase/chemistry , Protein Conformation , Viral Proteins/chemistry , Amino Acid Sequence , Animals , Aza Compounds/chemistry , Bacteriophage T4/enzymology , Catalysis , Chickens/metabolism , Crystallization , Crystallography, X-Ray , Egg Proteins/chemistry , Evolution, Molecular , Models, Molecular , Molecular Sequence Data , Muramidase/classification , Sequence Alignment , Sequence Homology, Amino Acid , Tryptophan/analogs & derivatives , Tryptophan/chemistry
8.
Acta Crystallogr D Biol Crystallogr ; 53(Pt 2): 217-9, 1997 Mar 01.
Article in English | MEDLINE | ID: mdl-15299961

ABSTRACT

After many unsuccessful attempts to crystallize the bacteriophage lambda lysozyme, a mutant where all the tryptophan residues have been replaced by aza-tryptophans has been crystallized by the vapor-diffusion method. The crystals are orthorhombic and belong to space group P2(1)2(1)2(1) with cell dimensions a = 73.01, b = 78.80, c = 82.31 A. Diffraction data were collected using synchrotron radiation sources. Crystals diffract to a resolution of 2.3 A. Data from two different platinum derivatives were also recorded to 2.8 and 2.5 A, respectively.

9.
Steroids ; 61(12): 688-96, 1996 Dec.
Article in English | MEDLINE | ID: mdl-8987137

ABSTRACT

Cholestane-derived gem-dihydroperoxides and tetraoxanes were synthesized starting from 5 alpha- and 5 beta-cholestan-3-ones by acid-catalyzed addition of hydrogen peroxide to the ketone. They were characterized by IR, NMR, and mass spectroscopy analysis aided by molecular mechanics calculations, and, in the instance of 5 beta-cholestane-3 alpha,3 beta-dihydroperoxide (6), by x-ray analysis. The synthesized compounds were tested in vitro against Plasmodium falciparum Sierra Leone (D6) and Indochina (W2) malaria clones. All compounds were inactive to both clones, with the exception of tetraoxane 7a, which exhibited modest activity toward D6 clone with IC50 = 155 nM.


Subject(s)
Antimalarials/chemistry , Antimalarials/pharmacology , Cholestanes/chemistry , Cholestanes/chemical synthesis , Peroxides/chemistry , Animals , Antimalarials/chemical synthesis , Cholestanes/pharmacology , Magnetic Resonance Spectroscopy , Mass Spectrometry , Models, Molecular , Molecular Structure , Peroxides/chemical synthesis , Peroxides/pharmacology , Plasmodium falciparum/drug effects , Steroids/chemistry
10.
Acta Crystallogr D Biol Crystallogr ; 52(Pt 1): 165-9, 1996 Jan 01.
Article in English | MEDLINE | ID: mdl-15299738

ABSTRACT

A new crystal form of pike (pI 4.10) parvalbumin has been crystallized in presence of EDTA at pH 8.0. The crystals are orthorhombic, space group P2(1)2(1)2(1), with a = 51.84, b = 49.95, c = 34.96 A. Diffractometer data were collected to 1.75 A. The structure was solved by molecular replacement and refined to R = 0.168 for 7774 observed reflections [I>/= 2sigma(I)] in the range 8.0-1.75 A. In spite of the presence of EDTA, calcium ions are present in both primary binding sites. As compared to the previously reported structures, the main differences concern the conformation of the N-terminal residues and the packing in the unit cell.

11.
J Pharm Sci ; 82(12): 1262-5, 1993 Dec.
Article in English | MEDLINE | ID: mdl-8308708

ABSTRACT

The crystal structure of polymorph I of p-(1R,3S)3-thioanisoyl-1,2,2-trimethylcyclopentane carboxylic acid has been determined and is compared with that of polymorph II that was previously described. Polymorph I is very different at the level of the carboxyl group. It does not present disorder and the values found for the C-O bonds correspond exactly to single and double bond lengths. In addition, the carboxyl groups of the two molecules in the cell packing are involved in symmetric hydrogen bonds [2.662(6) A] leading to the formation of a dimer around the twofold axis following x with a shift on z. The different conformations on the carboxylic group between the two polymorphs are in good agreement with the thermodynamic study.


Subject(s)
Benzoates/chemistry , Cyclopentanes/chemistry , Crystallography, X-Ray , Hydrogen Bonding , Molecular Structure , Stereoisomerism , Thermodynamics
12.
J Mol Biol ; 223(3): 705-20, 1992 Feb 05.
Article in English | MEDLINE | ID: mdl-1542115

ABSTRACT

The three-dimensional structure of parvalbumin from leopard shark (Triakis semifasciata) with 109 amino acid residues (alpha-series) is described at 1.54 A resolution. Crystals were grown at 20 degrees C from 2.9 M-potassium/sodium phosphate solutions at pH 5.6. The space group is P3(1)21 and unit cell dimensions are a = b = 32.12 A and c = 149.0 A. The structure has been solved by the molecular replacement method using pike 4.10 parvalbumin as a model. The final structure refinement resulted in an R-factor of 17.3% for 11,363 independent reflections at 1.54 A resolution. The shark parvalbumin shows the main features of all parvalbumins: the folding of the chain including six alpha-helices, the salt bridge between Arg75 and Glu81, and the hydrophobic core. Compared to the structure of beta-parvalbumins from pike and carp, one main difference is observed: the chain is one residue longer and this additional residue, which extends the F helix, is involved through its C-terminal carboxylate group in a network of electrostatic contacts with two basic residues, His31 in the B helix and Lys36 in the BC segment. Furthermore, hydrogen bonds exist between the side-chains of Gln108 (F helix) and Tyr26 (B helix). There is therefore a "locking" of the tertiary structure through contacts between two sequentially distant regions in the protein and this is likely to contribute to making the stability of an alpha-parvalbumin higher in comparison to that of a beta-parvalbumin. The lengthening of the C-terminal F helix by one residue appears to be a major feature of alpha-parvalbumins in general, owing to the homologies of the amino acid sequences. Besides the lengthening of the C-terminal helix, the classification of the leopard shark parvalbumin in the alpha-series rests upon the observation of Lys13, Leu32, Glu61 and Val66. As this is the first crystal structure description of a parvalbumin from the alpha-phylogenetic lineage, it was hoped that it would clearly determine the presence or absence of a third cation binding site in parvalbumins belonging to the alpha-lineage. In beta-pike pI 4.10 parvalbumin, Asp61 participates as a direct ligand of a third site, the satellite of the CD site. In shark parvalbumin, as in nearly all alpha-parvalbumins, one finds Glu at position 61. Unfortunately, the conformation of the polar head of Glu61 cannot be inferred from the X-ray data.(ABSTRACT TRUNCATED AT 400 WORDS)


Subject(s)
Muscles/chemistry , Parvalbumins/chemistry , Amino Acid Sequence , Animals , Binding Sites , Cations/metabolism , Computer Simulation , Histidine/chemistry , Hydrogen Bonding , Magnetic Resonance Spectroscopy , Models, Molecular , Molecular Sequence Data , Parvalbumins/metabolism , Sharks , Solvents , X-Ray Diffraction
13.
J Mol Biol ; 220(4): 1017-39, 1991 Aug 20.
Article in English | MEDLINE | ID: mdl-1880797

ABSTRACT

The crystal structure of the Ca-loaded form of pike 4.10 parvalbumin (minor component from pike muscle belonging to the beta phylogenetic series), with both its primary sites CD and EF occupied by Ca2+ ions and its third site occupied by an ammonium ion, as previously determined at 1.93 A resolution, has now been refined to a resolution of 1.65 A. The crystallization of this parvalbumin in different ionic environments has allowed three novel non-isomorphous crystalline forms to be obtained: (1) a first form, crystallized in the presence of a mixture of ammonium sulphate and manganese sulphate, for which all the cation binding sites in the protein are occupied by Mn2+; (2) a second form crystallized in the presence of MgSO4 as the precipitating agent, only differs from the Ca/NH4 form by the occupation of the third site by Mg2+, whereas the primary sites remain occupied by Ca2+; (3) a third form, also crystallized in the presence of MgSO4, corresponds to a well-defined molecular species with both the primary EF site and the third site occupied by Mg2+, whereas the primary CD site remains occupied by CA2+. The corresponding molecular structures reported here have been determined at resolutions between 1.8 and 2.4 A. The comparison of the different crystal structures allows the structural modifications accompanying the substitution of the primary sites by cations differing significantly in their ionic radii (Ca2+, Mn2+, Mg2+) to be investigated in detail, and it also leads to a precise description of the third site in a typical beta parvalbumin. The substitution Ca2+ by Mg2+ within the primary site EF is characterized by a "contraction" of the co-ordination sphere, with a decrease of the mean oxygen-metal distance by a value of 0.25 A and a decrease of the co-ordination number from 7 to 6, as a consequence of the loss of a bidentate ligand (Glu101), which becomes a monodentate one. Such an adaptation of the co-ordination sphere around a cation of smaller size involves, among others, the transformation of the Glu101 side-chain from the stable gauche(+) form to the less stable gauche(-) form. The third site is clearly described as a satellite of the CD primary site, since both sites possess common protein ligands, such as Asp53 and Glu59. Furthermore, Asp61 appears as a specific ligand of the third site in the different environments investigated in this work. We finally discuss the relevance of the third site to parvalbumin phylogeny.


Subject(s)
Parvalbumins/ultrastructure , Ammonia/chemistry , Animals , Binding Sites , Calcium/chemistry , Calcium-Binding Proteins/chemistry , Calcium-Binding Proteins/ultrastructure , Crystallography , Fishes , Hydrogen Bonding , Magnesium/chemistry , Magnetic Resonance Spectroscopy , Manganese/chemistry , Models, Molecular , Muscles , Parvalbumins/chemistry , Protein Binding , Protein Conformation , X-Ray Diffraction
14.
Acta Crystallogr C ; 46 ( Pt 7): 1259-62, 1990 Jul 15.
Article in English | MEDLINE | ID: mdl-2222931

ABSTRACT

C29H40N6O7.3H2O, Mr = 638.7, trigonal, P3(1)21, a = 14.190 (2), c = 29.833 (4) A, V = 5202 (1) A3, Z = 6, Dx = 1.22 g cm-3, Cu K alpha, lambda = 1.54178 A, mu = 7.8 cm-1, F(000) = 2052, T = 291 K, R = 0.069 for 1942 observed reflections. The new cyclohexapeptide cleromyrine II was isolated from Clerodendrum myricoides. Its structure was established by spectroscopic and X-ray diffraction methods as cyclo(-Gly-Tyr-Gly-Pro-Leu-Pro-). The conformation essentially consists of two beta-turns including the Pro residues and one central very short antiparallel beta-sheet stabilized by two intramolecular hydrogen bonds: N(Tyr2)...O(Leu5) = 2.94 (2) A and N(Leu5)...O(Tyr2) = 3.02 (2) A.


Subject(s)
Peptides, Cyclic , Plants/analysis , Amino Acid Sequence , Crystallization , Magnetic Resonance Spectroscopy , Mass Spectrometry , Molecular Sequence Data , Molecular Structure , Peptides, Cyclic/isolation & purification , X-Ray Diffraction
15.
J Mol Biol ; 202(2): 349-53, 1988 Jul 20.
Article in English | MEDLINE | ID: mdl-3172221

ABSTRACT

The crystal and molecular structure of the minor component of pike parvalbumins has been determined at 1.93 A resolution by molecular replacement (1 A = 0.1 nm). The crystals are orthorhombic, space group P2(1)2(1)2 with a = 59.62 A, b = 59.83 A and c = 26.35 A. A location of the secondary cation binding site is proposed for this parvalbumin of the beta phylogenetic series.


Subject(s)
Muscle Proteins , Parvalbumins , Amino Acid Sequence , Animals , Binding Sites , Crystallography , Salmonidae
17.
J Pharm Sci ; 72(3): 232-5, 1983 Mar.
Article in English | MEDLINE | ID: mdl-6842373

ABSTRACT

A second polymorph of spiperone, 8-[3-(p-fluorobenzoyl)-propyl]-1-phenyl-1,3,8-triazaspiro[4,5] decan-4-one, has been isolated and characterized by thermal analysis and IR spectrometry. Its structure was solved by X-ray diffraction analysis. The results are compared with those previously obtained on spiperone, the main difference being in the conformation of the side chain and in the nature of the hydrogen bonding.


Subject(s)
Butyrophenones , Spiperone , Butyrophenones/pharmacology , Chemical Phenomena , Chemistry , Models, Molecular , Molecular Conformation , Spectrophotometry, Infrared , Spiperone/pharmacology , Temperature , Thermodynamics , X-Ray Diffraction
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