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Helical Protein (helical + protein)
Selected AbstractsConformational and functional analysis of the lipid binding protein Ag-NPA-1 from the parasitic nematode Ascaridia galliFEBS JOURNAL, Issue 1 2005Rositsa Jordanova Ag-NPA-1 (AgFABP), a 15 kDa lipid binding protein (LBP) from Ascaridia galli, is a member of the nematode polyprotein allergen/antigen (NPA) family. Spectroscopic analysis shows that Ag-NPA-1 is a highly ordered, ,-helical protein and that ligand binding slightly increases the ordered secondary structure content. The conserved, single Trp residue (Trp17) and three Tyr residues determine the fluorescence properties of Ag-NPA-1. Analysis of the efficiency of the energy transfer between these chromophores shows a high degree of Tyr-Trp dipole-dipole coupling. Binding of fatty acids and retinol was accompanied by enhancement of the Trp emission, which allowed calculation of the affinity constants of the binary complexes. The distance between the single Trp of Ag-NPA-1 and the fluorescent fatty acid analogue 11-[(5-dimethylaminonaphthalene-1- sulfonyl)amino]undecanoic acid (DAUDA) from the protein binding site is 1.41 nm as estimated by fluorescence resonance energy transfer. A chemical modification of the Cys residues of Ag-NPA-1 (Cys66 and Cys122) with the thiol reactive probes 5-({[(2-iodoacetyl)amino]ethyl}amino) naphthalene-1-sulfonic acid (IAEDANS) and N,N,-dimethyl- N -(iodoacetyl)- N,-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)ethylenediamine (IANBD), followed by MALDI-TOF analysis showed that only Cys66 was labeled. The observed similar affinities for fatty acids of the modified and native Ag-NPA-1 suggest that Cys66 is not a part of the protein binding pocket but is located close to it. Ag-NPA-1 is one of the most abundant proteins in A. galli and it is distributed extracellularly mainly as shown by immunohistology and immunogold electron microscopy. This suggests that Ag-NPA-1 plays an important role in the transport of fatty acids and retinoids. [source] Stability of the hydration layer of tropocollagen: A QM studyJOURNAL OF COMPUTATIONAL CHEMISTRY, Issue 4 2010K. Pálfi Abstract Collagen is a triple helical protein, highly hydrated in nature. Bella and Berman (J Mol Biol 1996, 264, 734) have reported the structure of the first hydration layer. Water molecules form bridges of different length around the POG repeats and self assemble into left-handed helical water threads. To explore the stability of these specifically hydrated places, we have designed suitable QM models: each comprises a triple helix formed by 18 residues surrounded by 8 to 12 explicit waters. Two sets of amino acids were used, one standing for the core structural subunit of tropocollagen (POG-model) and one for its natural enzyme recognition sites (AAG-model). We have determined the stability order of the water binding places, the strongest being ,8.1 kcal mol,1, while the weakest ,6.1 kcal mol,1 per hydrogen bond. In X-ray structures, each triplet of tropocollagen is shielded by six to nine water molecules. Beside the mandatory six, the "surplus" three water molecules further strengthen the binding of all the others. However, the displacement of selected water molecules turns out to be energy neutral. These water binding places on the surface of the triple helix can provide explanation on how an almost liquid-like hydration environment exists between the closely packed tropocollagens (Henkelman et al., Magn Reson Med 1994, 32, 592). It seems that these water reservoirs or buffers can provide space for "hole conduction" of water molecules and thus contribute to the elasticity of collagen. © 2009 Wiley Periodicals, Inc. J Comput Chem, 2010 [source] Maximum-likelihood density modification using pattern recognition of structural motifsACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2001Thomas C. Terwilliger The likelihood-based approach to density modification [Terwilliger (2000), Acta Cryst. D56, 965,972] is extended to include the recognition of patterns of electron density. Once a region of electron density in a map is recognized as corresponding to a known structural element, the likelihood of the map is reformulated to include a term that reflects how closely the map agrees with the expected density for that structural element. This likelihood is combined with other aspects of the likelihood of the map, including the presence of a flat solvent region and the electron-density distribution in the protein region. This likelihood-based pattern-recognition approach was tested using the recognition of helical segments in a largely helical protein. The pattern-recognition method yields a substantial phase improvement over both conventional and likelihood-based solvent-flattening and histogram-matching methods. The method can potentially be used to recognize any common structural motif and incorporate prior knowledge about that motif into density modification. [source] Structure of jack bean chitinaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2000M. Hahn The structure of jack bean chitinase was solved at 1.8,Å resolution by molecular replacement. It is an ,-helical protein with three disulfide bridges. The active site is related in structure to animal and viral lysozymes. However, unlike in lysozyme, the architecture of the active site suggests a single-step cleavage. According to this mechanism, Glu68 is the proton donor and Glu90 assists in the reaction by moving towards the substrate and recruiting a water molecule that acts as the nucleophile. In this model, a water molecule was found in contact with Glu90 O,1 and Thr119,O, at a distance of 3.0 and 2.8,Å, respectively. The model is in accordance with the observed inversion mechanism. [source] Structure of mouse ADP-ribosylhydrolase 3 (mARH3)ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2008Christoph Mueller-Dieckmann ADP-ribosylation is a reversible and covalent post-translational modification in which the attachment of ADP-ribose is catalyzed by ADP-ribosyltransferases and the removal of ADP-ribose is catalyzed by ADP-ribosylhydrolases. ADP-ribosylhydrolase 3 from mouse, consisting of 347 amino-acid residues, has been cloned, purified and crystallized. The three-dimensional structure has been resolved at a resolution of 1.8,Å. The structure constitutes a compact all-,-helical protein with two Mg2+ ions located in the active-site crevice. A structural comparison of mouse ADP-ribosylhydrolase 3 with its human orthologue shows a high degree of structural similarity. Furthermore, four prokaryotic proteins deposited in the PDB could be identified as being structurally related. [source] Probing the ,-Helical Structural Stability of Stapled p53 Peptides: Molecular Dynamics Simulations and AnalysisCHEMICAL BIOLOGY & DRUG DESIGN, Issue 4 2010Zuojun Guo Reactivation of the p53 cell apoptosis pathway through inhibition of the p53-hDM2 interaction is a viable approach to suppress tumor growth in many human cancers and stabilization of the helical structure of synthetic p53 analogs via a hydrocarbon cross-link (staple) has been found to lead to increased potency and inhibition of protein,protein binding (J. Am. Chem. Soc. 129: 5298). However, details of the structure and dynamic stability of the stapled peptides are not well understood. Here, we use extensive all-atom molecular dynamics simulations to study a series of stapled ,-helical peptides over a range of temperatures in solution. The peptides are found to exhibit substantial variations in predicted ,-helical propensities that are in good agreement with the experimental observations. In addition, we find significant variation in local structural flexibility of the peptides with the position of the linker, which appears to be more closely related to the observed differences in activity than the absolute ,-helical stability. These simulations provide new insights into the design of ,-helical stapled peptides and the development of potent inhibitors of ,-helical protein,protein interfaces. [source] Beta-helix model for the filamentous haemagglutinin adhesin of Bordetella pertussis and related bacterial secretory proteinsMOLECULAR MICROBIOLOGY, Issue 2 2001Andrey V. Kajava Bordetella pertussis establishes infection by attaching to epithelial cells of the respiratory tract. One of its adhesins is filamentous haemagglutinin (FHA), a 500-Å-long secreted protein that is rich in ,-structure and contains two regions, R1 and R2, of tandem 19-residue repeats. Two models have been proposed in which the central shaft is (i) a hairpin made up of a pairing of two long antiparallel ,-sheets; or (ii) a ,-helix in which the polypeptide chain is coiled to form three long parallel ,-sheets. We have analysed a truncated variant of FHA by electron microscopy (negative staining, shadowing and scanning transmission electron microscopy of unstained specimens): these observations support the latter model. Further support comes from detailed sequence analysis and molecular modelling studies. We applied a profile search method to the sequences adjacent to and between R1 and R2 and found additional ,covert' copies of the same motifs that may be recognized in overt form in the R1 and R2 sequence repeats. Their total number is sufficient to support the tenet of the ,-helix model that the shaft domain , a 350 Å rod , should consist of a continuous run of these motifs, apart from loop inserts. The N-terminus, which does not contain such repeats, was found to be weakly homologous to cyclodextrin transferase, a protein of known immunoglobulin-like structure. Drawing on crystal structures of known ,-helical proteins, we developed structural models of the coil motifs putatively formed by the R1 and R2 repeats. Finally, we applied the same profile search method to the sequence database and found several other proteins , all large secreted proteins of bacterial provenance , that have similar repeats and probably also similar structures. [source] Stoichiometry of lipid interactions with transmembrane proteins,Deduced from the 3D structuresPROTEIN SCIENCE, Issue 5 2006Tibor Páli Abstract The stoichiometry of the first shell of lipids interacting with a transmembrane protein is defined operationally by the population of spin-labeled lipid chains whose motion is restricted directly by the protein. Interaction stoichiometries have been determined experimentally for a wide range of ,-helical integral membrane proteins by using spin-label ESR spectroscopy. Here, we determine the spatially defined number of first-shell lipids at the hydrophobic perimeter of integral membrane proteins whose 3D structure has been determined by X-ray crystallography and lipid,protein interactions characterized by spin-labeling. Molecular modeling is used to build a single shell of lipids surrounding transmembrane structures derived from the PDB. Constrained energy optimization of the protein,lipid assemblies is performed by molecular mechanics. For relatively small proteins (up to 7,12 transmembrane helices), the geometrical first shell corresponds to that defined experimentally by perturbation of the lipid-chain dynamics. For larger, multi-subunit ,-helical proteins, the lipids perturbed directly by the protein may either exceed or be less in number than those that can be accommodated at the intramembranous perimeter. In these latter cases, the motionally restricted spin-labeled lipids can be augmented by intercalation, or can correspond to a specific subpopulation at the protein interface, respectively. For monomeric ,-barrel proteins, the geometrical lipid stoichiometry corresponds to that determined from lipid mobility for a 22-stranded barrel, but fewer lipids are motionally restricted than can be accommodated around an eight-stranded barrel. Deviations from the geometrical first shell, in the ,-barrel case, are for the smaller protein with a highly curved barrel. [source] How do helix,helix interactions help determine the folds of membrane proteins?PROTEIN SCIENCE, Issue 4 2003Perspectives from the study of homo-oligomeric helical bundles FRET, fluorescence resonance energy transfer; NBD, 7-nitrobenz-2-oxa-1,3-diazole; C-14 betaine, N -tetradecyl- N,N -dimethyl-3-ammonio-1-propanesulfonate; MF, mole fraction Abstract The final, structure-determining step in the folding of membrane proteins involves the coalescence of preformed transmembrane helices to form the native tertiary structure. Here, we review recent studies on small peptide and protein systems that are providing quantitative data on the interactions that drive this process. Gel electrophoresis, analytical ultracentrifugation, and fluorescence resonance energy transfer (FRET) are useful methods for examining the assembly of homo-oligomeric transmembrane helical proteins. These methods have been used to study the assembly of the M2 proton channel from influenza A virus, glycophorin, phospholamban, and several designed membrane proteins,all of which have a single transmembrane helix that is sufficient for association into a transmembrane helical bundle. These systems are being studied to determine the relative thermodynamic contributions of van der Waals interactions, conformational entropy, and polar interactions in the stabilization of membrane proteins. Although the database of thermodynamic information is not yet large, a few generalities are beginning to emerge concerning the energetic differences between membrane and water-soluble proteins: the packing of apolar side chains in the interior of helical membrane proteins plays a smaller, but nevertheless significant, role in stabilizing their structure. Polar, hydrogen-bonded interactions occur less frequently, but, nevertheless, they often provide a strong driving force for folding helix,helix pairs in membrane proteins. These studies are laying the groundwork for the design of sequence motifs that dictate the association of membrane helices. [source] End-to-end and end-to-middle interhelical interactions: new classes of interacting helix pairs in protein structuresACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2009Tarini Shankar Ghosh Helix,helix interactions are important for the structure, stability and function of ,-helical proteins. Helices that either cross in the middle or show extensive contacts between each other, such as coiled coils, have been investigated in previous studies. Interactions between two helices can also occur only at the terminal regions or between the terminal region of one helix and the middle region of another helix. Examples of such helix pairs are found in aquaporin, H+/Cl, transporter and Bcl-2 proteins. The frequency of the occurrence of such `end-to-end' (EE) and `end-to-middle' (EM) helix pairs in protein structures is not known. Questions regarding the residue preferences in the interface and the mode of interhelical interactions in such helix pairs also remain unanswered. In this study, high-resolution structures of all-, proteins from the PDB have been systematically analyzed and the helix pairs that interact only in EE or EM fashion have been extracted. EE and EM helix pairs have been categorized into five classes (N,N, N,C, C,C, N,MID and C,MID) depending on the region of interaction. Nearly 13% of 5725 helix pairs belonged to one of the five classes. Analysis of single-residue propensities indicated that hydrophobic and polar residues prefer to occur in the C-terminal and N-terminal regions, respectively. Hydrophobic C-terminal interacting residues and polar N-terminal interacting residues are also highly conserved. A strong correlation exists between some of the residue properties (surface area/volume and length of side chains) and their preferences for occurring in the interface of EE and EM helix pairs. In contrast to interacting non-EE/EM helix pairs, helices in EE and EM pairs are farther apart. In these helix pairs, residues with large surface area/volume and longer side chains are preferred in the interfacial region. [source] Sequence-specific recognition of DNA by hydrophobic, alanine-rich mutants of the basic region/leucine zipper motif investigated by fluorescence anisotropyBIOPOLYMERS, Issue 1 2002Gregory H. Bird Abstract We generated minimalist proteins capable of sequence-specific, high-affinity binding of DNA to probe how proteins are used and can be used to recognize DNA. In order to quantify binding affinities and specificities in our protein,DNA system, we used fluorescence anisotropy to measure in situ the thermodynamics of binding of alanine-rich mutants of the GCN4 basic region/leucine zipper (bZIP) domain to DNA duplexes containing target sites AP-1 (5,-TGACTCA-3,) or ATF/CREB (5,-TGACGTCA-3,). We simplified the ,-helical bZIP molecular recognition scaffold by alanine substitution: 4A, 11A, and 18A contain four, eleven, and eighteen alanine mutations in their DNA-binding basic regions, respectively. DNase I footprinting analysis demonstrates that all bZIP mutants retain the sequence-specific DNA-binding function of native GCN4 bZIP. Titration of fluorescein-labeled oligonucleotide duplexes with increasing amounts of protein yielded low nanomolar dissociation constants for all bZIP mutants in complex with the AP-1 and ATF/CREB sites: binding to the nonspecific control duplex was > 1000-fold weaker. Remarkably, the most heavily mutated protein 18A, containing 24 alanines in its 27-residue basic region, still binds AP-1 and ATF/CREB with dissociation constants of 15 and 7.8 nM, respectively. Similarly, wild-type bZIP binds these sites with Kd values of 9.1 and 14 nM. 11A also displays low nanomolar dissociation constants for AP-1 and ATF/CREB, while 4A binds these sites with , 10-fold weaker Kd values. Thus, both DNA-binding specificity and affinity are maintained in all our bZIP derivatives. This Ala-rich scaffold may be useful in design and synthesis of small ,-helical proteins with desired DNA-recognition properties capable of serving as therapeutics targeting transcription. © 2002 Wiley Periodicals, Inc. Biopolymers 65: 10,20, 2002 [source] | |||||||||||||||||||||||||