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Sample records for peptide conformer acidity

  1. Entropy reduction in unfolded peptides (and proteins) due to conformational preferences of amino acid residues.

    PubMed

    Schweitzer-Stenner, Reinhard; Toal, Siobhan E

    2014-11-07

    As established by several groups over the last 20 years, amino acid residues in unfolded peptides and proteins do not exhibit the unspecific random distribution as assumed by the classical random coil model. Individual amino acid residues in small peptides were found to exhibit different conformational preferences. Here, we utilize recently obtained conformational distributions of guest amino acid residues in GxG peptides to estimate their conformational entropy, which we find to be significantly lower than the entropy of an assumed random coil like distribution. Only at high temperature do backbone entropies approach random coil like values. We utilized the obtained backbone entropies of the investigated amino acid residues to estimate the loss of conformational entropy caused by a coil → helix transition and identified two subsets of amino acid residues for which the thus calculated entropy losses correlate well with the respective Gibbs energy of helix formation obtained for alanine based host-guest systems. Calculated and experimentally derived entropic losses were found to be in good agreement. For most of the amino acid residues investigated entropic losses derived from our GxG distributions correlate very well with corresponding values recently obtained from MD simulations biased by conformational propensities derived from truncated coil libraries. Both, conformational entropy and the entropy of solvation exhibit a strong, residue specific temperature dependence, which can be expected to substantially affect the stability of unfolded states. Altogether, our results provide strong evidence for the notion that conformational preferences of amino acid residues matter with regard to the thermodynamics of peptide and protein folding.

  2. Predicting three-dimensional conformations of peptides constructed of only glycine, alanine, aspartic acid, and valine.

    PubMed

    Oda, Akifumi; Fukuyoshi, Shuichi

    2015-06-01

    The GADV hypothesis is a form of the protein world hypothesis, which suggests that life originated from proteins (Lacey et al. 1999; Ikehara 2002; Andras 2006). In the GADV hypothesis, life is thought to have originated from primitive proteins constructed of only glycine, alanine, aspartic acid, and valine ([GADV]-proteins). In this study, the three-dimensional (3D) conformations of randomly generated short [GADV]-peptides were computationally investigated using replica-exchange molecular dynamics (REMD) simulations (Sugita and Okamoto 1999). Because the peptides used in this study consisted of only 20 residues each, they could not form certain 3D structures. However, the conformational tendencies of the peptides were elucidated by analyzing the conformational ensembles generated by REMD simulations. The results indicate that secondary structures can be formed in several randomly generated [GADV]-peptides. A long helical structure was found in one of the hydrophobic peptides, supporting the conjecture of the GADV hypothesis that many peptides aggregated to form peptide multimers with enzymatic activity in the primordial soup. In addition, these results indicate that REMD simulations can be used for the structural investigation of short peptides.

  3. Predicting Three-Dimensional Conformations of Peptides Constructed of Only Glycine, Alanine, Aspartic Acid, and Valine

    NASA Astrophysics Data System (ADS)

    Oda, Akifumi; Fukuyoshi, Shuichi

    2015-06-01

    The GADV hypothesis is a form of the protein world hypothesis, which suggests that life originated from proteins (Lacey et al. 1999; Ikehara 2002; Andras 2006). In the GADV hypothesis, life is thought to have originated from primitive proteins constructed of only glycine, alanine, aspartic acid, and valine ([GADV]-proteins). In this study, the three-dimensional (3D) conformations of randomly generated short [GADV]-peptides were computationally investigated using replica-exchange molecular dynamics (REMD) simulations (Sugita and Okamoto 1999). Because the peptides used in this study consisted of only 20 residues each, they could not form certain 3D structures. However, the conformational tendencies of the peptides were elucidated by analyzing the conformational ensembles generated by REMD simulations. The results indicate that secondary structures can be formed in several randomly generated [GADV]-peptides. A long helical structure was found in one of the hydrophobic peptides, supporting the conjecture of the GADV hypothesis that many peptides aggregated to form peptide multimers with enzymatic activity in the primordial soup. In addition, these results indicate that REMD simulations can be used for the structural investigation of short peptides.

  4. Conformational studies on peptides of alpha-aminoxy acids with functionalized side-chains.

    PubMed

    Yang, Dan; Liu, Guo-Jun; Hao, Yu; Li, Wei; Dong, Ze-Min; Zhang, Dan-Wei; Zhu, Nian-Yong

    2010-06-01

    Peptides of homochiral alpha-aminoxy acids of nonpolar side chains can form a 1.8(8)-helix. In this paper, we report the conformational studies of alpha-aminoxy peptides 1-3, which have functionalized side chains, in both nonpolar and polar solvents. (1)H NMR, XRD, and FTIR absorption studies confirm the presence of the eight-membered-ring intramolecular hydrogen bonds (the N-O turns) in nonpolar solvents as well as in methanol. CD studies of peptides 1-3 in different solvents indicate that a substantial degree of helical content is retained in methanol and acidic aqueous buffers. The introduction of functionalized side chains in alpha-aminoxy peptides provides opportunities for designing biologically active peptides.

  5. Silicon-Containing Amino Acids: Synthetic Aspects, Conformational Studies, and Applications to Bioactive Peptides.

    PubMed

    Rémond, Emmanuelle; Martin, Charlotte; Martinez, Jean; Cavelier, Florine

    2016-10-12

    Unnatural α-amino acids form a family of essential molecules used for, among other applications, the synthesis of modified peptides, to improve resistance to proteolytic enzyme degradation, and to modulate physico- and biochemical properties of bioactive peptides as well as chiral inducers in asymmetric synthesis. Among them, silicon-containing unnatural amino acids are becoming an interesting new class of building blocks. The replacement of carbon atoms in bioactive substances with silicon is becoming increasingly popular. Peptides containing silyl amino acids hold great promise for maintaining or reinforcing the biological activity of active compounds, while they simultaneously enhance their resistance to enzyme degradation. In addition, the lipophilicity of the silicon atom facilitates their membrane crossing and their bioavailability. Nowadays, the interest of the pharmaceutical industry in peptide- and protein-based therapies is increasing. In this respect, silicon-containing amino acids and peptides are likely to be a significant part of future innovations in this area, and more generally in the area of biomolecules. In this process, commercial availability of silicon-containing amino acids is necessary: new syntheses have been developed, and work in this area is ongoing. This review aims to be a comprehensive and general summary of the different methods used to prepare silicon-containing amino acids and their implications on conformational structures and biological applications when they are incorporated into bioactive molecules.

  6. Conformational characterization of the 1-aminocyclobutane-1-carboxylic acid residue in model peptides.

    PubMed

    Gatos, M; Formaggio, F; Crisma, M; Toniolo, C; Bonora, G M; Benedetti, Z; Di Blasio, B; Iacovino, R; Santini, A; Saviano, M; Kamphuis, J

    1997-01-01

    A series of N- and C-protected, monodispersed homo-oligopeptides (to the dodecamer level) from the small-ring alicyclic C alpha, alpha-dialkylated glycine 1-aminocyclobutane-1-carboxylic acid (Ac4c) and two Ala/Ac4c tripeptides were synthesized by solution methods and fully characterized. The conformational preferences of all the model peptides were determined in deuterochloroform solution by FT-IR absorption and 1H-NMR. The molecular structures of the amino acid derivatives Z-Ac4c-OH and Z2-Ac4c-OH, the tripeptides Z-(Ac4c)3-OtBu, Z-Ac4c-(L-Ala)2-OMe and Z-L-Ala-Ac4c-L-Ala-OMe, and the tetrapeptide Z-(Ac4c)4-OtBu were determined in the crystal state by X-ray diffraction. The average geometry of the cyclobutyl moiety of the Ac4c residue was assessed and the tau(N-C alpha-C') bond angle was found to be significantly expanded from the regular tetrahedral value. The conformational data are strongly in favour of the conclusion that the Ac4c residue is an effective beta-turn and helix former. A comparison with the structural propensities of alpha-aminoisobutyric acid, the prototype of C alpha, alpha-dialkylated glycines, and the other extensively investigated members of the family of 1-aminocycloalkane-1-carboxylic acids (Acnc, with n = 3, 5-8) is made and the implications for the use of the Ac4c residue in conformationally constrained peptide analogues are briefly examined.

  7. Acid-base titration of melanocortin peptides: evidence of Trp rotational conformers interconversion.

    PubMed

    Fernandez, Roberto M; Vieira, Renata F F; Nakaie, Clóvis R; Lamy, M Teresa; Ito, Amando S

    2005-01-01

    Tryptophantime-resolved fluorescence was used to monitor acid-base titration properties of alpha-melanocyte stimulating hormone (alpha-MSH) and the biologically more potent analog [Nle4, D-Phe7]alpha -MSH (NDP-MSH), labeled or not with the paramagnetic amino acid probe 2,2,6,6-tetramthylpiperidine-N-oxyl-4-amino-4-carboxylic acid (Toac). Global analysis of fluorescence decay profiles measured in the pH range between 2.0 and 11.0 showed that, for each peptide, the data could be well fitted to three lifetimes whose values remained constant. The less populated short lifetime component changed little with pH and was ascribed to Trp g+ chi1 rotamer, in which electron transfer deactivation predominates over fluorescence. The long and intermediate lifetime preexponential factors interconverted along that pH interval and the result was interpreted as due to interconversion between Trp g- and trans chi1 rotamers, driven by conformational changes promoted by modifications in the ionization state of side-chain residues. The differences in the extent of interconversion in alpha-MSH and NDP-MSH are indicative of structural differences between the peptides, while titration curves suggest structural similarities between each peptide and its Toac-labeled species, in aqueous solution. Though less sensitive than fluorescence, the Toac electron spin resonance (ESR) isotropic hyperfine splitting parameter can also monitor the titration of side-chain residues located relatively far from the probe.

  8. Peptide Conformer Acidity Analysis of Protein Flexibility Monitored by Hydrogen Exchange†

    PubMed Central

    2009-01-01

    The amide hydrogens that are exposed to solvent in the high-resolution X-ray structures of ubiquitin, FK506-binding protein, chymotrypsin inhibitor 2, and rubredoxin span a billion-fold range in hydroxide-catalyzed exchange rates which are predictable by continuum dielectric methods. To facilitate analysis of transiently accessible amides, the hydroxide-catalyzed rate constants for every backbone amide of ubiquitin were determined under near physiological conditions. With the previously reported NMR-restrained molecular dynamics ensembles of ubiquitin (PDB codes 2NR2 and 2K39) used as representations of the Boltzmann-weighted conformational distribution, nearly all of the exchange rates for the highly exposed amides were more accurately predicted than by use of the high-resolution X-ray structure. More strikingly, predictions for the amide hydrogens of the NMR relaxation-restrained ensemble that become exposed to solvent in more than one but less than half of the 144 protein conformations in this ensemble were almost as accurate. In marked contrast, the exchange rates for many of the analogous amides in the residual dipolar coupling-restrained ubiquitin ensemble are substantially overestimated, as was particularly evident for the Ile 44 to Lys 48 segment which constitutes the primary interaction site for the proteasome targeting enzymes involved in polyubiquitylation. For both ensembles, “excited state” conformers in this active site region having markedly elevated peptide acidities are represented at a population level that is 102 to 103 above what can exist in the Boltzmann distribution of protein conformations. These results indicate how a chemically consistent interpretation of amide hydrogen exchange can provide insight into both the population and the detailed structure of transient protein conformations. PMID:19722680

  9. Assessing the Native State Conformational Distribution of Ubiquitin by Peptide Acidity

    PubMed Central

    Hernández, Griselda; Anderson, Janet S.; LeMaster, David M.

    2011-01-01

    At equilibrium, every energetically feasible conformation of a protein occurs with a non-zero probability. Quantitative analysis of protein flexibility is thus synonymous with determining the proper Boltzmann-weighting of this conformational distribution. The exchange reactivity of solvent-exposed amide hydrogens greatly varies with conformation, while the short-lived peptide anion intermediate implies an insensitivity to the dynamics of conformational motion. Amides that are well-exposed in model conformational ensembles of ubiquitin vary a million-fold in exchange rates which continuum dielectric methods can predict with an rmsd of 3. However, the exchange rates for many of the more rarely exposed amides are markedly overestimated in the PDB-deposited 2K39 and 2KN5 ubiquitin ensembles, while the 2NR2 ensemble predictions are largely consistent with those of the Boltzmann-weighted conformational distribution sampled at the level of 1%. The correlation between the fraction of solvent-accessible conformations for a given amide hydrogen and the exchange rate constant for that residue provides a useful monitor of the degree of completeness with which a given ensemble has sampled the energetically accessible conformational space. These exchange predictions correlate with the degree to which each ensemble deviates from a set of 46 ubiquitin X-ray structures. Kolmogorov-Smirnov analysis for the distribution of intra- and inter-ensemble pairwise structural rmsd values assisted the identification of a subensemble of 2K39 that eliminates the overestimations of hydrogen exchange rates observed for the full ensemble. The relative merits of incorporating experimental restraints into the conformational sampling process is compared to using these restraints as filters to select subpopulations consistent with the experimental data. PMID:21055867

  10. Role of enthalpy-entropy compensation interactions in determining the conformational propensities of amino acid residues in unfolded peptides.

    PubMed

    Toal, Siobhan E; Verbaro, Daniel J; Schweitzer-Stenner, Reinhard

    2014-02-06

    The driving forces governing the unique and restricted conformational preferences of amino acid residues in the unfolded state are still not well understood. In this study, we experimentally determine the individual thermodynamic components underlying intrinsic conformational propensities of these residues. Thermodynamic analysis of ultraviolet-circular dichroism (UV-CD) and (1)H NMR data for a series of glycine capped amino acid residues (i.e., G-x-G peptides) reveals the existence of a nearly exact enthalpy-entropy compensation for the polyproline II-β strand equilibrium for all investigated residues. The respective ΔHβ, ΔSβ values exhibit a nearly perfect linear relationship with an apparent compensation temperature of 295 ± 2 K. Moreover, we identified iso-equilibrium points for two subsets of residues at 297 and 305 K. Thus, our data suggest that within this temperature regime, which is only slightly below physiological temperatures, the conformational ensembles of amino acid residues in the unfolded state differ solely with respect to their capability to adopt turn-like conformations. Such iso-equilibria are rarely observed, and their existence herein indicates a common physical origin behind conformational preferences, which we are able to assign to side-chain dependent backbone solvation. Conformational effects such as differences between the number of sterically allowed side chain rotamers can contribute to enthalpy and entropy but not to the Gibbs energy associated with conformational preferences. Interestingly, we found that alanine, aspartic acid, and threonine are the only residues which do not share these iso-equilbiria. The enthalpy-entropy compensation discovered as well as the iso-equilbrium and thermodynamics obtained for each amino acid residue provide a new and informative way of identifying the determinants of amino acid propensities in unfolded and disordered states.

  11. Conformational characterization of peptides rich in the cycloaliphatic C alpha,alpha-disubstituted glycine 1-aminocyclononane-1-carboxylic acid.

    PubMed

    Gatos, M; Formaggio, F; Crisma, M; Valle, G; Toniolo, C; Bonora, G M; Saviano, M; Iacovino, R; Menchise, V; Galdiero, S; Pedone, C; Benedetti, E

    1997-01-01

    A series of N- and C-protected, monodispersed homo-oligopeptides (to the pentamer level) from the cycloaliphatic C alpha,alpha-dialkylated glycine 1-aminocyclononane-1-carboxylic acid (Ac9c) and two Ala/Ac9c tripeptides have been synthesized by solution methods and fully characterized. The conformational preferences of all the model peptides were determined in deuterochloroform solution by FT-IR absorption and 1H-NMR. The molecular structures of the amino acid derivatives mCIAc-Ac9c-OH and Z-Ac9c-OtBu, the dipeptide pBrBz-(Ac9c)2-OtBu, the tetrapeptide Z-(Ac9c)4-OtBu, and the pentapeptide Z-(Ac9c)5-OtBu were determined in the crystal state by X-ray diffraction. Based on this information, the average geometry and the preferred conformation for the cyclononyl moiety of the Ac9c residue have been assessed. The backbone conformational data are strongly in favour of the conclusion that the Ac9c residue is a strong beta-turn and helix former. A comparison with the structural propensity of alpha-aminoisobutyric acid, the prototype of C alpha,alpha-dialkylated glycines, and the other extensively investigated members of the family of 1-aminocycloalkane-1-carboxylic acids (Acnc, with n = 3-8) is made and the implications for the use of the Ac9c residue in conformationally constrained analogues of bioactive peptides are briefly examined.

  12. Amino Acid Chirality and Ferrocene Conformation Guided Self-Assembly and Gelation of Ferrocene-Peptide Conjugates.

    PubMed

    Adhikari, Bimalendu; Singh, Charanpreet; Shah, Afzal; Lough, Alan J; Kraatz, Heinz-Bernhard

    2015-08-03

    The self-assembly and gelation behavior of a series of mono- and disubstituted ferrocene (Fc)-peptide conjugates as a function of ferrocene conformation and amino acid chirality are described. The results reveal that ferrocene-peptide conjugates self-assemble into organogels by controlling the conformation of the central ferrocene core, through inter- versus intramolecular hydrogen bonding in the attached peptide chain(s). The chirality controlled assembling studies showed that two monosubstituted Fc conjugates FcCO-LFLFLA-OMe and FcCO-LFLFDA-OMe form gels with nanofibrillar network structures, whereas the other two diastereomers FcCO-DFLFLA-OMe and FcCO-LFDFLA-OMe exclusively produced straight nanorods and non-interconnected small fibers, respectively. This suggests the potential tuning of gelation behavior and nanoscale morphology by altering the chirality of constituted amino acids. The current study confirms the profound effect of diastereomerism and no influence of enantiomers on gelation. Correspondingly, the diastereomeric and enantiomeric Fc[CO-FFA-OMe]2 were constructed for the study of chirality-organized structures.

  13. A proposed bioactive conformation of Peptide T

    NASA Astrophysics Data System (ADS)

    Centeno, Nuria B.; Perez, Juan J.

    1998-01-01

    The conformational profiles of Peptide T, (5-8)Peptide T, [Abu5](4-8)Peptide T and (4-8)Peptide T were computed independently to assess the geometrical characteristics of the bioactive conformation of Peptide T. The conformational profiles of the peptides were computed within the molecular mechanics framework using an effective dielectric constant of 80. The conformational space was thoroughly sampled using an iterative simulated annealing protocol. The bioactive conformation was assessed by pairwise cross comparisons of each of the unique low energy conformations found for each of the different analogs studied. After a putative bioactive conformation was selected, in order to further validate our hypothesis the conformational profile of the potent compound cyclo(Thr-Thr-Asn-Tyr-Thr-Asp) was computed and the putative bioactive conformation was found. The conformation exhibits a pseudo β-turn involving the side chain of Thr5 and the carbonyl oxygen of Tyr7 forming a C12 ring.

  14. Side-chain conformational thermodynamics of aspartic acid residue in the peptides and achatin-I in aqueous solution.

    PubMed

    Kimura, Tomohiro; Matubayasi, Nobuyuki; Nakahara, Masaru

    2004-02-01

    Sequence-position dependence of the side-chain conformational equilibrium of aspartic acid (Asp) residue is investigated for both model Asp peptides (di- to tetra-) and neuropeptide achatin-I (Gly--Phe-Ala-Asp) in aqueous solution. The trans-to-gauche conformational changes on the dihedral angle of C-C(alpha)-C(beta)-C are analyzed in terms of the standard free energy DeltaG(0), enthalpy DeltaH(0), and entropy -TDeltaS(0). The thermodynamic quantities are obtained by measuring the dihedral-angle-dependent vicinal (1)H-(1)H coupling constants in nuclear magnetic resonance over a wide temperature range. When the carboxyl groups of Asp are ionized, DeltaG(0) in the aqueous phase depends by approximately 1-2 kJ mol(-1) on the sequence position, whereas the energy change in the gas phase (absence of solvent) depends by tens of kJ mol(-1). Therefore, the weak position dependence of DeltaG(0) is a result of the compensation for the intramolecular effect by the hydration (= DeltaG(0)-). The DeltaH(0) and -TDeltaS(0) components, on the other hand, exhibit a notable trend at the C-terminus. The C-terminal DeltaH(0) is larger than the N- and nonterminal DeltaH(0) values due to the intramolecular repulsion between alpha- and beta-. The C-terminal -TDeltaS(0) is negative and larger in magnitude than the others, and an attractive solute-solvent interaction at the C-terminus serves as a structure breaker of the water solvent.

  15. Specific Amyloid Binding of Polybasic Peptides In Vivo Is Retained by β-Sheet Conformers but Lost in the Disrupted Coil and All D-Amino Acid Variants.

    PubMed

    Wall, Jonathan S; Williams, Angela; Richey, Tina; Stuckey, Alan; Wooliver, Craig; Christopher Scott, J; Donnell, Robert; Martin, Emily B; Kennel, Stephen J

    2017-02-22

    The heparin-reactive, helical peptide p5 is an effective amyloid imaging agent in mice with systemic amyloidosis. Analogs of p5 with modified secondary structure characteristics exhibited altered binding to heparin, synthetic amyloid fibrils, and amyloid extracts in vitro. Herein, we further study the effects of peptide helicity and chirality on specific amyloid binding using a mouse model of systemic inflammation-associated (AA) amyloidosis. Peptides with disrupted helical structure [p5(coil) and p5(Pro3)], with an extended sheet conformation [p5(sheet)] or an all-D enantiomer [p5(D)], were chemically synthesized, radioiodinated, and their biodistribution studied in WT mice as well as transgenic animals with severe systemic AA amyloidosis. Peptide binding was assessed qualitatively by using small animal single-photon emission computed tomography/x-ray computed tomography imaging and microautoradiography and quantitatively using tissue counting. Peptides with reduced helical propensity, p5(coil) and p5(Pro3), exhibited significantly reduced binding to AA amyloid-laden organs. In contrast, peptide p5(D) was retained by non-amyloid-related ligands in the liver and kidneys of both WT and AA mice, but it also bound AA amyloid in the spleen. The p5(sheet) peptide specifically bound AA amyloid in vivo and was not retained by healthy tissues in WT animals. Modification of amyloid-targeting peptides using D-amino acids should be performed cautiously due to the introduction of unexpected secondary pharmacologic effects. Peptides that adopt a helical structure, to align charged amino acid side chains along one face, exhibit specific reactivity with amyloid; however, polybasic peptides with a propensity for β-sheet conformation are also amyloid-reactive and may yield a novel class of amyloid-targeting agents for imaging and therapy.

  16. Integrating the intrinsic conformational preferences of non-coded α-amino acids modified at the peptide bond into the NCAD database

    PubMed Central

    Revilla-López, Guillem; Rodríguez-Ropero, Francisco; Curcó, David; Torras, Juan; Calaza, M. Isabel; Zanuy, David; Jiménez, Ana I.; Cativiela, Carlos; Nussinov, Ruth; Alemán, Carlos

    2011-01-01

    Recently, we reported a database (NCAD, Non-Coded Amino acids Database; http://recerca.upc.edu/imem/index.htm) that was built to compile information about the intrinsic conformational preferences of non-proteinogenic residues determined by quantum mechanical calculations, as well as bibliographic information about their synthesis, physical and spectroscopic characterization, the experimentally-established conformational propensities, and applications (J. Phys. Chem. B 2010, 114, 7413). The database initially contained the information available for α-tetrasubstituted α-amino acids. In this work, we extend NCAD to three families of compounds, which can be used to engineer peptides and proteins incorporating modifications at the –NHCO– peptide bond. Such families are: N-substituted α-amino acids, thio-α-amino acids, and diamines and diacids used to build retropeptides. The conformational preferences of these compounds have been analyzed and described based on the information captured in the database. In addition, we provide an example of the utility of the database and of the compounds it compiles in protein and peptide engineering. Specifically, the symmetry of a sequence engineered to stabilize the 310-helix with respect to the α-helix has been broken without perturbing significantly the secondary structure through targeted replacements using the information contained in the database. PMID:21491493

  17. Protein biosynthesis with conformationally restricted amino acids

    SciTech Connect

    Mendel, D. Lawrence Berkeley Lab., CA ); Ellman, J.; Schultz, P.G. )

    1993-05-19

    The incorporation of conformationally constrained amino acids into peptides is a powerful approach for generating structurally defined peptides as conformational probes and bioactive agents. The ability to site-specifically introduce constrained amino acids into large polypeptide chains would provide a similar opportunity to probe the flexibility, conformation, folding and stability of proteins. To this end, we have examined the competence of the Escherichia coli protein biosynthetic machinery to incorporate a number of these unnatural amino acids into the 164 residue protein T4 lysozyme (T4L). Results clearly demonstrate that the protein biosynthetic machinery can accommodate a wide variety of conformationally constrained amino acids. The expansion of structural motifs that can be biosynthetically incorporated into proteins to include a large number of conformationally constrained amino acids significantly increases the power of mutagenesis methods as probes of protein structure and function and provides additional insights into the steric requirements of the translational machinery. 13 refs., 2 figs.

  18. Helices with additional H-bonds: crystallographic conformations of α,γ-hybrid peptides helices composed of β-hydroxy γ-amino acids (statines).

    PubMed

    Malik, Ankita; Kumar, Mothukuri Ganesh; Bandyopadhyay, Anupam; Gopi, Hosahudya N

    2017-01-01

    β-Hydroxy-γ-amino acids (Statines) are a class of naturally occurring non-ribosomal amino acids frequently found in many peptide natural products. Peptidomimetics constituted with statines have been used as inhibitors for various aspartic acid proteases. In contrast to the synthetic γ-amino acids, very little is known about the folding behavior of these naturally occurring β-hydroxy γ-amino acids. To understand the folding behavior of statines, three α,γ-hybrid peptides P1 (Ac-Aib-γPhe-Aib-(R, S)Phesta-Aib-γPhe-Aib-CONH2 ), P2 (Ac-Aib-γPhe-Aib-(S, S)Phesta-Aib-γPhe-Aib-CONH2 ), and P3 (Ac-Aib-γPhe-Aib-(S, S)Phesta-Aib-(S, S)Phesta-Aib-CONH2 ) were synthesized on solid phase and their helical conformations in single crystals were studied. Results suggest that both syn and anti diastereoisomers of statines can be accommodated into the helix without deviating overall helical conformation of α,γ-hybrid peptides. In comparison with syn diastereoisomer, the anti diastereoisomer was found to be directly involved in the intramolecular H-bonding with the backbone carbonyl groups (i to i + 3) similar to the backbone amide NHs in the helix.

  19. Side-Chain Conformational Thermodynamics of Aspartic Acid Residue in the Peptides and Achatin-I in Aqueous Solution

    PubMed Central

    Kimura, Tomohiro; Matubayasi, Nobuyuki; Nakahara, Masaru

    2004-01-01

    Sequence-position dependence of the side-chain conformational equilibrium of aspartic acid (Asp) residue is investigated for both model Asp peptides (di- to tetra-) and neuropeptide achatin-I (Gly-𝒟-Phe-Ala-Asp) in aqueous solution. The trans-to-gauche conformational changes on the dihedral angle of C–Cα–Cβ–C are analyzed in terms of the standard free energy ΔG0, enthalpy ΔH0, and entropy −TΔS0. The thermodynamic quantities are obtained by measuring the dihedral-angle-dependent vicinal 1H-1H coupling constants in nuclear magnetic resonance over a wide temperature range. When the carboxyl groups of Asp are ionized, ΔG0 in the aqueous phase depends by ∼1–2 kJ mol−1 on the sequence position, whereas the energy change \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} \\begin{equation*}{\\Delta}E_{{\\mathrm{gas}}}^{0}\\end{equation*}\\end{document} in the gas phase (absence of solvent) depends by tens of kJ mol−1. Therefore, the weak position dependence of ΔG0 is a result of the compensation for the intramolecular effect \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} \\begin{equation*}{\\Delta}E_{{\\mathrm{gas}}}^{0}\\end{equation*}\\end{document} by the hydration \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} \\begin{equation*}{\\Delta}G_{{\\mathrm{hyd}}}^{0}\\end{equation*}\\end{document} (= ΔG0–\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage

  20. Assessing the Impact of Backbone Length and Capping Agent on the Conformational Preferences of a Model Peptide: Conformation Specific IR and UV Spectroscopy of 2-AMINOISOBUTYRIC Acid

    NASA Astrophysics Data System (ADS)

    Gord, Joseph R.; Hewett, Daniel M.; Kubasik, Matthew A.; Zwier, Timothy S.

    2015-06-01

    2-Aminoisobutyric acid (Aib) is an achiral, α-amino acid having two equivalent methyl groups attached to C_α. Extended Aib oligomers are known to have a strong preference for the adoption of a 310-helical structure in the condensed phase. Here, we have taken a simplifying step and focused on the intrinsic folding propensities of Aib by looking at a series of capped Aib oligomers in the gas phase, free from the influence of solvent molecules and cooled in a supersonic expansion. Resonant two-photon ionization and IR-UV holeburning have been used to record single-conformation UV spectra using the Z-cap as the UV chromophore. Resonant ion-dip infrared (RIDIR) spectroscopy provides single-conformation IR spectra in the OH stretch and NH stretch regions. Data have been collected on a set of Z-(Aib)n-X oligomers with n = 1, 2, 4, 6 and X = -OH and -OMethyl. The impacts of these capping groups and differences in backbone length have been found to dramatically influence the conformational space accessed by the molecules studied here. Oligomers of n=4 have sufficient backbone length for a full turn of the 310-helix to be formed. Early interpretation of the data collected shows clear spectroscopic markers signaling the onset of 310-helix formation as well as evidence of structures incorporating C7 and C14 hydrogen bonded rings. Toniolo, C.; Bonora, G. M.; Barone, V.; Bavoso, A.; Benedetti, E.; Di Blasio, B.; Grimaldi, P.; Lelj, F.; Pavone, V.; Pedone, C., Conformation of Pleionomers of α-Aminoisobutyric Acid. Macromolecules 1985, 18, 895-902.

  1. Systematic conformational investigations of peptoids and peptoid-peptide chimeras.

    PubMed

    Brandt, Wolfgang; Herberg, Thomas; Wessjohann, Ludger

    2011-01-01

    Peptoids are originally defined as N-substituted oligoglycine derivatives, and in a broader definition as N-substituted peptides (peptoid-peptide chimeras). Both types were systematically investigated by force field calculations. The Merck MMFF and YASARA2 force fields were shown to be, among others, the most suitable ones for conformational investigations of peptoids with no missing parameterizations, in contrast to AMBER or CHARMM. Ramachandran-like plots were calculated for dipeptoids and chimeras using energy calculations and grid searches by varying the dihedral angels PHI and PSI in steps of 10 degrees for s-cis- and s-trans amide bonds. Barriers as well as low energy conformations are compared to peptide Ramachandran plots, showing that peptoids have both, more barriers due to additional steric interactions as well as access to minimum conformations not accessible by peptides. Low energy conformations of dimers were used as starting conformations of higher oligomers of the peptoids for extensive molecular dynamics simulations over 10 or 20 ns with the YASARA2 force field and an explicit water solvent box to evaluate their potential to form secondary structural elements. Especially peptoids with aminoisobutyric acid-like monomer units were found to form left-handed or polyproline-like helices also known from less common natural peptides. Furthermore, new secondary structures appear feasible based on stable conformations outside the allowed areas of the Ramachandran plot for peptides, but allowed for peptoids.

  2. Synthesis of conformationally preorganized and cell-permeable guanidine-based gamma-peptide nucleic acids (gammaGPNAs).

    PubMed

    Sahu, Bichismita; Chenna, Venugopal; Lathrop, Kira L; Thomas, Sufi M; Zon, Gerald; Livak, Kenneth J; Ly, Danith H

    2009-02-20

    A general method for preparing optically pure guanidine-based gamma-peptide nucleic acid (gammaGPNA) monomers for all four natural nucleobases (A, C, G, and T) is described. These second-generation gammaGPNAs differ from the first-generation GPNAs in that the guanidinium group is installed at the gamma- instead of the alpha-position of the N-(2-aminoethyl)glycine backbone unit. This positional switch enables GPNAs to be synthesized from relatively cheap L- as opposed to D-amino acids. Unlike their alpha-predecessors, which are randomly folded, gammaGPNAs prepared from L-amino acids are preorganized into a right-handed helix and bind to DNA and RNA with exceptionally high affinity and sequence selectivity and are readily taken up by mammalian cells.

  3. Synthesis of Conformationally Preorganized and Cell-Permeable Guanidine-Based γ-Peptide Nucleic Acids (γGPNAs)

    PubMed Central

    Sahu, Bichismita; Chenna, Venugopal; Lathrop, Kira L.; Thomas, Sufi M.; Zon, Gerald; Livak, Kenneth J.; Ly, Danith H.

    2009-01-01

    A general method for preparing optically-pure guanidine-based γ-peptide nucleic acid (γGPNA) monomers for all four natural nucleobases (A, C, G and T) is described. These second-generation γGPNAs differ from the first in that the guanidinium group is installed at the γ-instead of the α-position of the N-(2-aminoethyl)glycine backbone unit. This positional switch enables GPNAs to be synthesized from relatively cheap L- as opposed to D-amino acids. Unlike their α-predecessors, which are randomly-folded, γGPNAs prepared from L-amino acids are preorganized into a right-handed helix and bind to DNA and RNA with exceptionally high affinity and sequence selectivity, and are readily taken up by mammalian cells. PMID:19161276

  4. Conformational Sampling of Peptides in Cellular Environments☆

    PubMed Central

    Tanizaki, Seiichiro; Clifford, Jacob; Connelly, Brian D.; Feig, Michael

    2008-01-01

    Abstract Biological systems provide a complex environment that can be understood in terms of its dielectric properties. High concentrations of macromolecules and cosolvents effectively reduce the dielectric constant of cellular environments, thereby affecting the conformational sampling of biomolecules. To examine this effect in more detail, the conformational preference of alanine dipeptide, poly-alanine, and melittin in different dielectric environments is studied with computer simulations based on recently developed generalized Born methodology. Results from these simulations suggest that extended conformations are favored over α-helical conformations at the dipeptide level at and below dielectric constants of 5–10. Furthermore, lower-dielectric environments begin to significantly stabilize helical structures in poly-alanine at ɛ = 20. In the more complex peptide melittin, different dielectric environments shift the equilibrium between two main conformations: a nearly fully extended helix that is most stable in low dielectrics and a compact, V-shaped conformation consisting of two helices that is preferred in higher dielectric environments. An additional conformation is only found to be significantly populated at intermediate dielectric constants. Good agreement with previous studies of different peptides in specific, less-polar solvent environments, suggest that helix stabilization and shifts in conformational preferences in such environments are primarily due to a reduced dielectric environment rather than specific molecular details. The findings presented here make predictions of how peptide sampling may be altered in dense cellular environments with reduced dielectric response. PMID:17905846

  5. Analysis of residue conformations in peptides in Cambridge structural database and protein-peptide structural complexes.

    PubMed

    Raghavender, Upadhyayula Surya

    2017-03-01

    A comprehensive statistical analysis of the geometric parameters of peptide chains in a reduced dataset of protein-peptide complexes in Protein Data Bank (PDB) is presented. The angular variables describing the backbone conformations of amino acid residues in peptide chains shed insights into the conformational preferences of peptide residues interacting with protein partners. Nonparametric statistical approaches are employed to evaluate the interrelationships and associations in structural variables. Grouping of residues based on their structure into chemical classes reveals characteristic trends in parameter relationships. A comparison of canonical amino acid residues in free peptide structures in Cambridge structural database (CSD) with identical residues in PDB complexes, suggests that the information can be integrated from both the structural repositories enabling efficient and accurate modeling of biologically active peptides. © 2016 John Wiley & Sons A/S.

  6. Does L to D-amino acid substitution trigger helix→sheet conformations in collagen like peptides adsorbed to surfaces?

    PubMed

    Velmurugan, Punitha; Jonnalagadda, Raghava Rao; Sankaranarayanan, Kamatchi; Dhathathreyan, Aruna

    2015-12-01

    The present work reports on the structural order, self assembling behaviour and the role in adsorption to hydrophilic or hydrophobic solid surfaces of modified sequence from the triple helical peptide model of the collagenase cleavage site in type I collagen (Uniprot accession number P02452 residues from 935 to 970) using (D)Ala and (D)Ile substitutions as given in the models below: Model-1: GSOGADGPAGAOGTOGPQGIAGQRGVV GLOGQRGER. Model-2: GSOGADGP(D)AGAOGTOGPQGIAGQRGVVGLOGQRGER. Model-3: GSOGADGPAGAOGTOGPQG(D)IAGQRGVVGLOGQRGER. Collagenase is an important enzyme that plays an important role in degrading collagen in wound healing, cancer metastasis and even in embryonic development. However, the mechanism by which this degradation occurs is not completely understood. Our results show that adsorption of the peptides to the solid surfaces, specifically hydrophobic triggers a helix to beta transition with order increasing in peptide models 2 and 3. This restricts the collagenolytic behaviour of collagenase and may find application in design of peptides and peptidomimetics for enzyme-substrate interaction, specifically with reference to collagen and other extra cellular matrix proteins.

  7. Effect of D-amino acids at Asp{sup 23} and Ser{sup 26} residues on the conformational preference of A{beta}{sub 20-29} peptides

    SciTech Connect

    Shanmugam, Ganesh; Polavarapu, Prasad L. . E-mail: Prasad.L.Polavarapu@Vanderbilt.edu; Hallgas, Balazs; Majer, Zsuzsa

    2005-09-30

    The effects of D-amino acids at Asp{sup 23} and Ser{sup 26} residues on the conformational preference of {beta}-amyloid (A{beta}) peptide fragment (A{beta}{sub 20-29}) have been studied using different spectroscopic techniques, namely vibrational circular dichroism (VCD), vibrational absorption, and electronic circular dichroism. To study the structure of the A{beta}{sub 20-29}, [D-Asp{sup 23}]A{beta}{sub 20-29}, and [D-Ser{sup 26}]A{beta}{sub 20-29} peptides under different conditions, the spectra were measured in 10 mM acetate buffer (pH 3) and in 2,2,2-trifluoroethanol (TFE). The spectroscopic results indicated that at pH 3, A{beta}{sub 20-29} peptide takes random coil with {beta}-turn structure, while [D-Ser{sup 26}]A{beta}{sub 20-29} peptide adopts significant amount of polyproline II (PPII) type structure along with {beta}-turn contribution and D-Asp-substituted peptide ([D-Asp{sup 23}]A{beta}{sub 20-29}) adopts predominantly PPII type structure. The increased propensity for PPII conformation upon D-amino acid substitution, in acidic medium, has important biological implications. In TFE, A{beta}{sub 20-29}, [D-Asp{sup 23}]A{beta}{sub 20-29}, and [D-Ser{sup 26}]A{beta}{sub 20-29} peptides adopt 3{sub 10}-helix, {alpha}-helix, and random coil with some {beta}-turn structures, respectively. The VCD data obtained for the A{beta} peptide films suggested that the secondary structures for the peptide films are not the same as those for corresponding solution and are also different among the A{beta} peptides studied here. This observation suggests that dehydration can have a significant influence on the structural preferences of these peptides.

  8. Peptide N-Amination Supports β-Sheet Conformations.

    PubMed

    Sarnowski, Matthew P; Kang, Chang Won; Elbatrawi, Yassin M; Wojtas, Lukasz; Del Valle, Juan R

    2017-02-13

    The conformational heterogeneity of backbone N-substituted peptides limits their ability to adopt stable secondary structures. Herein, we describe a practical synthesis of backbone aminated peptides that readily adopt β-sheet folds. Data derived from model N-amino peptides suggest that extended conformations are stabilized through cooperative steric, electrostatic, and hydrogen-bonding interactions.

  9. Halogen Bonding: A Powerful Tool for Modulation of Peptide Conformation.

    PubMed

    Danelius, Emma; Andersson, Hanna; Jarvoll, Patrik; Lood, Kajsa; Gräfenstein, Jürgen; Erdélyi, Máté

    2017-06-27

    Halogen bonding is a weak chemical force that has so far mostly found applications in crystal engineering. Despite its potential for use in drug discovery, as a new molecular tool in the direction of molecular recognition events, it has rarely been assessed in biopolymers. Motivated by this fact, we have developed a peptide model system that permits the quantitative evaluation of weak forces in a biologically relevant proteinlike environment and have applied it for the assessment of a halogen bond formed between two amino acid side chains. The influence of a single weak force is measured by detection of the extent to which it modulates the conformation of a cooperatively folding system. We have optimized the amino acid sequence of the model peptide on analogues with a hydrogen bond-forming site as a model for the intramolecular halogen bond to be studied, demonstrating the ability of the technique to provide information about any type of weak secondary interaction. A combined solution nuclear magnetic resonance spectroscopic and computational investigation demonstrates that an interstrand halogen bond is capable of conformational stabilization of a β-hairpin foldamer comparable to an analogous hydrogen bond. This is the first report of incorporation of a conformation-stabilizing halogen bond into a peptide/protein system, and the first quantification of a chlorine-centered halogen bond in a biologically relevant system in solution.

  10. Conformational properties of oxazoline-amino acids

    NASA Astrophysics Data System (ADS)

    Staś, Monika; Broda, Małgorzata A.; Siodłak, Dawid

    2016-04-01

    Oxazoline-amino acids (Xaa-Ozn) occur in natural peptides of potentially important bioactivity. The conformations of the model compounds: Ac-(S)-Ala-Ozn(4R-Me), Ac-(S)-Ala-Ozn(4S-Me), and (gauche+, gauche-, anti) Ac-(S)-Val-Ozn(4R-Me) were studied at meta-hybrid M06-2X/6-311++G(d,p) method including solvent effect. Boc-L-Ala-L-Ozn-4-COOMe and Boc-L-Val-L-Ozn-4-COOMe were synthesized and studied by FT-IR and NMR-NOE methods. The conformations in crystal state were gathered from the Cambridge Structural Data Base. The main conformational feature of the oxazoline amino acids is the conformation β2 (ϕ,ψ ∼ -161°, -6°), which predominates in weakly polar environment and still is accessible in polar surrounding. The changes of the conformational preferences towards the conformations αR (ϕ,ψ ∼ -70°, -15°) and then β (ϕ,ψ ∼ -57°, -155°) are observed with increase of the environment polarity.

  11. Peptide Sequence and Conformation Strongly Influence Tryptophan Fluorescence

    PubMed Central

    Alston, Roy W.; Lasagna, Mauricio; Grimsley, Gerald R.; Scholtz, J. Martin; Reinhart, Gregory D.; Pace, C. Nick

    2008-01-01

    This article probes the denatured state ensemble of ribonuclease Sa (RNase Sa) using fluorescence. To interpret the results obtained with RNase Sa, it is essential that we gain a better understanding of the fluorescence properties of tryptophan (Trp) in peptides. We describe studies of N-acetyl-l-tryptophanamide (NATA), a tripeptide: AWA, and six pentapeptides: AAWAA, WVSGT, GYWHE, HEWTV, EAWQE, and DYWTG. The latter five peptides have the same sequence as those surrounding the Trp residues studied in RNase Sa. The fluorescence emission spectra, the fluorescence lifetimes, and the fluorescence quenching by acrylamide and iodide were measured in concentrated solutions of urea and guanidine hydrochloride. Excited-state electron transfer from the indole ring of Trp to the carbonyl groups of peptide bonds is thought to be the most important mechanism for intramolecular quenching of Trp fluorescence. We find the maximum fluorescence intensities vary from 49,000 for NATA with two carbonyls, to 24,400 for AWA with four carbonyls, to 28,500 for AAWAA with six carbonyls. This suggests that the four carbonyls of AWA are better able to quench Trp fluorescence than the six carbonyls of AAWAA, and this must reflect a difference in the conformations of the peptides. For the pentapeptides, EAWQE has a fluorescence intensity that is more than 50% greater than DYWTG, showing that the amino acid sequence influences the fluorescence intensity either directly through side-chain quenching and/or indirectly through an influence on the conformational ensemble of the peptides. Our results show that peptides are generally better models for the Trp residues in proteins than NATA. Finally, our results emphasize that we have much to learn about Trp fluorescence even in simple compounds. PMID:18065477

  12. pH-directed self-assembling helical peptide conformation

    USDA-ARS?s Scientific Manuscript database

    The beta-sheet and alpha-helix peptide conformation are two of the most fundamentally ordered secondary structures found in proteins and peptides. They also give rise to self-assembling motifs that form macromolecular channels and nanostructures. Through design these conformations can yield enhance...

  13. Molecular dynamics simulation and conformational analysis of some catalytically active peptides.

    PubMed

    Honarparvar, Bahareh; Skelton, Adam A

    2015-04-01

    The design of stable and inexpensive artificial enzymes with potent catalytic activity is a growing field in peptide science. The first step in this design process is to understand the key factors that can affect the conformational preference of an enzyme and correlate them with its catalytic activity. In this work, molecular dynamics simulations in explicit water of two catalytically active peptides (peptide 1: Fmoc-Phe1-Phe2-His-CONH2; peptide 2: Fmoc-Phe1-Phe2-Arg-CONH2) were performed at temperatures of 300, 400, and 500 K. Conformational analysis of these peptides using Ramachandran plots identified the secondary structures of the amino acid residues involved (Phe1, Phe2, His, Arg) and confirmed their conformational flexibility in solution. Furthermore, Ramachandran maps revealed the intrinsic preference of the constituent residues of these compounds for a helical conformation. Long-range interaction distances and radius of gyration (R g) values obtained during 20 ns MD simulations confirmed their tendency to form folded conformations. Results showed a decrease in side-chain (Phe1, Phe2, His ring, and Arg) contacts as the temperature was raised from 300 to 400 K and then to 500 K. Finally, the radial distribution functions (RDF) of the water molecules around the nitrogen atoms in the catalytically active His and Arg residues of peptide 1 and peptide 2 revealed that the strongest water-peptide interaction occurred with the arginine nitrogen atoms in peptide 2. Our results highlight differences in the secondary structures of the two peptides that can be explained by the different arrangement of water molecules around the nitrogen atoms of Arg in peptide 2 as compared to the arrangement of water molecules around the nitrogen atoms of His in peptide 1. The results of this work thus provide detailed insight into peptide conformations which can be exploited in the future design of peptide analogs.

  14. Conformational Variety of Polyanionic Peptides At Low Salt Concentrations

    NASA Astrophysics Data System (ADS)

    Bertrand, Marylène; Brack, André

    1997-12-01

    Cordially dedicated to Dr. Leslie Orgel on the occasion of his 70th birthday. Sequential oligo- and polypeptides based on glutamic acid and leucine residues have been synthesized. In pure water, they exhibit a random coil conformation. Addition of very small amounts of divalent metallic cations induces the formation of ordered structure in the peptides which remain in solution. Higher salt concentrations precipitate the peptides. Polypeptides with alternating glutamic acid and leucine residues undergo a coil to β-sheet transition in the presence of Ca^2+, Ba^2+, Mn^2+, Co^2+, Zn^2+ and Hg^2+. Addition of Cu^2+ or Fe^3+ induces the formation of an α-helix. Solid amorphous CdS generates water soluble β-sheets, as well. Sequential poly(Leu-Glu-Glu-Leu) adopts an α-helix in the presence of divalent cations. The sequence-dependent conformational diversity was extended to poly(Asp-Leu) and poly(Leu-Asp-Asp-Leu).

  15. Conformational study of O-glycosylated threonine containing peptide models.

    PubMed

    Maeji, N J; Inoue, Y; Chujo, R

    1987-06-01

    1H n.m.r. studies of Z-Thr-OMe, Z-Thr-Ala-OMe, Z-Ala-Thr-OMe and their glycosylated derivatives indicate the possibility of an intramolecular hydrogen bond between Thr N alpha H and the N-acetyl carbonyl of the carbohydrate moiety, 3,4,6-tri-O-acetyl-2-acetamido-2-deoxy-alpha-D-galactopyranose (AcGalNAc). This is especially true in the case of Z-Thr(AcGalNAc)-Ala-OMe, suggesting that the strength of this hydrogen bond is dependent on the neighboring amino acids on the carbonyl terminal side of Thr. The existence of such a hydrogen bond implies a conformation in which the carbohydrate moiety is restricted to an orientation with its plane roughly perpendicular to the peptide backbone. In such an orientation, steric problems will be minimized in the case of clustered O-glycosidically linked Thr(Ser) residues as found in human erythrocyte glycophorin. A locked orientation of the carbohydrate moiety with respect to the peptide backbone may also play a conformational role in antifreeze glycoproteins.

  16. Conformational and Functional Effects Induced by D- and L-Amino Acid Epimerization on a Single Gene Encoded Peptide from the Skin Secretion of Hypsiboas punctatus

    PubMed Central

    de Magalhães, Mariana T. Q.; Barbosa, Eder A.; Prates, Maura V.; Verly, Rodrigo M.; Munhoz, Victor Hugo O.; de Araújo, Ivan E.; Bloch, Carlos

    2013-01-01

    Skin secretion of Hypsiboas punctatus is the source of a complex mixture of bioactive compounds where peptides and small proteins prevail, similarly to many other amphibians. Among dozens of molecules isolated from H. punctatus in a proteomic based approach, we report here the structural and functional studies of a novel peptide named Phenylseptin (FFFDTLKNLAGKVIGALT-NH2) that was purified as two naturally occurring D- and L-Phes configurations. The amino acid epimerization and C-terminal amidation for both molecules were confirmed by a combination of techniques including reverse-phase UFLC, ion mobility mass spectrometry, high resolution MS/MS experiments, Edman degradation, cDNA sequencing and solid-phase peptide synthesis. RMSD analysis of the twenty lowest-energy 1H NMR structures of each peptide revealed a major 90° difference between the two backbones at the first four N-terminal residues and substantial orientation changes of their respective side chains. These structural divergences were considered to be the primary cause of the in vitro quantitative differences in antimicrobial activities between the two molecules. Finally, both molecules elicited equally aversive reactions in mice when delivered orally, an effect that depended entirely on peripheral gustatory pathways. PMID:23565145

  17. Conformational structure of the amphipathic peptide insecticide L-KALA

    SciTech Connect

    Nandel, F.S.; Ahluwalia, A.; Kaur, A.

    1995-07-05

    The conformation structure of the tetrapeptide Ac-Lys-Ala-Leu-Ala-OMe, the repeat unit of a 30 residue peptide YAA(KALA){sub 6}LAA, and also its analog containing an unusual amino acid, (Aib): Ac-Lys-Aib-Leu-Aib-OMe, has been investigated by the PCILO method. The global/low-energy minima for all the residues lies at {phi} = -30{degrees} and {psi} = 120{degrees}. In this conformation, the potential hydrogen-bonding sites are free, i.e., capable of intermolecular hydrogen bonding. As the angles lie closer to the collagen helical region and potential hydrogen-bonding sites are free, this structure is named the collagen-type helix. In aqueous solution, this structure is stabilized by salvation of the peptide bond. The local-energy minimum in all maps correspond to the right-handed helical region. The helix thus formed generates a pore of 3 {Angstrom} along the helix axis, with one lateral hydrophobic side and the other hydrophilic side. The pore thus formed is cation-selective and accounts for the leakage of contents from vesicles. On the basis of the length of the helix and placement of the side chains of lysine and leucine, a model has been proposed for the aggregation and fusion of the vesicles. The amphipathic basic residues interact with the head groups of the acidic liposomes by extending toward the polar face of the helix to insert their charged moieties by overcoming the electrostatic repulsive forces between the opposing vesicles. 40 refs., 6 figs.

  18. Polyproline II conformation is one of many local conformational states and is not an overall conformation of unfolded peptides and proteins

    PubMed Central

    Makowska, Joanna; Rodziewicz-Motowidło, Sylwia; Bagińska, Katarzyna; Vila, Jorge A.; Liwo, Adam; Chmurzyński, Lech; Scheraga, Harold A.

    2006-01-01

    The alanine-based peptide Ac-XX(A)7OO-NH2, referred to as XAO (where X, A, and O denote diaminobutyric acid, alanine, and ornithine, respectively), has recently been proposed to possess a well defined polyproline II (PII) conformation at low temperatures. Based on the results of extensive NMR and CD investigations combined with theoretical calculations, reported here, we present evidence that, on the contrary, this peptide does not have any significant amount of organized PII structure but exists in an ensemble of conformations with a distorted bend in the N- and C-terminal regions. The conformational ensemble was obtained by molecular dynamics/simulated annealing calculations using the amber suite of programs with time-averaged distance and dihedral-angle restraints obtained from rotating-frame nuclear Overhauser effect (ROE) volumes and vicinal coupling constants 3JHNΗα, respectively. The computed ensemble-averaged radius of gyration Rg (7.4 ± 1.0) Å is in excellent agreement with that measured by small-angle x-ray scattering (SAXS) whereas, if the XAO peptide were in the PII conformation, Rg would be 11.6 Å. Depending on the pH, peptide concentration, and temperature, the CD spectra of XAO do or do not possess the maximum with positive ellipticity in the 217-nm region, which is characteristic of the PII structure, reflecting a shifting conformational equilibrium rather than an all-or-none transition. The “PII conformation” should, therefore, be considered as one of the accessible conformational states of individual amino acid residues in peptides and proteins rather than as a structure of most of the chain in the early stage of folding. PMID:16446433

  19. Comprehensive conformational studies of five tripeptides and a deduced method for efficient determinations of peptide structures.

    PubMed

    Yu, Wenbo; Wu, Zhiqing; Chen, Huibin; Liu, Xu; MacKerell, Alexander D; Lin, Zijing

    2012-02-23

    Thorough searches on the potential energy surfaces of five tripeptides, GGG, GYG, GWG, TGG, and MGG, were performed by considering all possible combinations of the bond rotational degrees of freedom with a semiempirical and ab initio combined computational approach. Structural characteristics of the obtained stable tripeptide conformers were carefully analyzed. Conformers of the five tripeptides were found to be closely connected with conformers of their constituting dipeptides and amino acids. A method for finding all important tripeptide conformers by optimizing a small number of trial structures generated by suitable superposition of the parent amino acid and dipeptide conformers is thus proposed. Applying the method to another five tripeptides, YGG, FGG, WGG, GFA, and GGF, studied before shows that the new approach is both efficient and reliable by providing the most complete ensembles of tripeptide conformers. The method is further generalized for application to larger peptides by introducing the breeding and mutation concepts in a genetic algorithm way. The generalized method is verified to be capable of finding tetrapeptide conformers with secondary structures of strands, helices, and turns, which are highly populated in larger peptides. This show some promise for the proposed method to be applied for the structural determination of larger peptides. © 2012 American Chemical Society

  20. Comprehensive Conformational Studies of Five Tripeptides and a Deduced Method for Efficient Determinations of Peptide Structures

    PubMed Central

    Yu, Wenbo; Wu, Zhiqing; Chen, Huibin; Liu, Xu; MacKerell, Alexander D.

    2012-01-01

    Thorough searches on the potential energy surfaces of five tripeptides, GGG, GYG, GWG, TGG and MGG, were performed by considering all possible combinations of the bond rotational degrees of freedom with a semi-empirical and ab initio combined computational approach. Structural characteristics of the obtained stable tripeptide conformers were carefully analyzed. Conformers of the five tripeptides were found to be closely connected with conformers of their constituting dipeptides and amino acids. A method for finding all important tripeptide conformers by optimizing a small number of trial structures generated by suitable superposition of the parent amino acid and dipeptide conformers is thus proposed. Applying the method to another five tripeptides, YGG, FGG, WGG, GFA and GGF, studied before shows that the new approach is both efficient and reliable by providing the most complete ensembles of tripeptide conformers. The method is further generalized for application to larger peptides by introducing the breeding and mutation concepts in a genetic algorithm way. The generalized method is verified to be capable of finding tetrapeptide conformers with secondary structures of strands, helices and turns which are highly populated in larger peptides. This show some promise for the proposed method to be applied for the structural determination of larger peptides. PMID:22260814

  1. Manipulation of peptide conformations by fine-tuning of the environment and/or the primary sequence.

    PubMed

    Li, S C; Kim, P K; Deber, C M

    1995-06-01

    The widely observed phenomenon that peptides are capable of adopting multiple conformations in different environments suggests that secondary structure formation in a peptide segment is a process involving not only the peptide itself but also the surrounding solvent media. The influence of the primary sequence and the molecular environment on peptide conformations are now investigated using synthetic peptides of amino acid sequence H2N-(Ser-Lys)2-Ala-X-Gly-Ala-X-Gly-Trp-Ala-X-Gly-(Lys-Ser)3-OH, where X = Ile or Val. These two peptides, namely 3I (X = Ile) and 3V (X = Val), are found to lack defined secondary structure in aqueous buffer. However, discrete conformational states, e.g., alpha-helices and beta-sheets, are readily generated and interconverted for both peptides when the buffer is modulated with the addition of methanol, sodium dodecyl sulfate (SDS) micelles, or phospholipid vesicles. The role of the primary sequence in affecting peptide conformations is manifested in that peptides 3I and 3V, which differ respectively in their content of beta-branched Ile or Val residues, differ in their secondary structures at monomeric concentrations in 2 mM SDS and in mixed lipid vesicles of phosphatidic acid and phosphatidylcholine. The overall results suggest that peptide segments can be conformationally flexible entities poised to react to minor modulation in either the molecular environment or the primary sequence, a circumstance that may be relevant to protein functioning and folding.

  2. Fluoroolefins as peptide mimetics. 2. A computational study of the conformational ramifications of peptide bond replacement.

    PubMed

    McKinney, Brian E; Urban, Joseph J

    2010-01-21

    The design of peptide mimetic compounds is greatly facilitated by the identification of functionalities that can act as peptide replacements. The fluoroalkene moiety has recently been employed for that purpose. The purpose of this work is to examine the conformational ramifications of replacing peptide bonds with fluoroalkene moieties, thus generating peptidomimetics. The alanine dipeptide analogue (ADA) was chosen as a model compound. Three peptidomimetic systems were investigated including one generated by replacement of both peptide bonds of ADA, designated as DFA, and those generated by the single replacement of the C-terminal peptide bond and N-terminal peptide bond, designated as CFA and NFA, respectively. Conformations for all three systems were generated by exhaustive Monte Carlo searching. Relative conformational energies were calculated at the MP2/aug-cc-pVTZ/MP2/aug-cc-pVDZ (for DFA), MP2/-aug-cc-pVTZ//MP2/6-311+G(d,p), B3LYP/6-31+G(d)//B3LYP/6-31+G(d), and MMFF levels of theory. Aqueous phase conformational preferences were determined through calculations making use of continuum hydration models. The results indicate that replacement of both peptide bonds of ADA generates a peptidomimetic with conformational preferences where extended conformations are favored and the conformational profile is relatively insensitive to the nature of the surrounding medium. This is in contrast to ADA where the conformational preferences depend highly on the surrounding medium and where folded conformations with intramolecular hydrogen bonds are important in the absence of an interacting solvent. CFA and NFA are found to exhibit conformational preferences that do in some ways more closely resemble those of the alanine dipeptide analogue. This is particularly true in the case of NFA where interactions between the NH and CF groups are reminiscent of the intramolecular hydrogen bonding possible in ADA.

  3. Synthesis and characterization of conformationally preorganized, (R)-diethylene glycol-containing γ-peptide nucleic acids with superior hybridization properties and water solubility.

    PubMed

    Sahu, Bichismita; Sacui, Iulia; Rapireddy, Srinivas; Zanotti, Kimberly J; Bahal, Raman; Armitage, Bruce A; Ly, Danith H

    2011-07-15

    Developed in the early 1990s, peptide nucleic acid (PNA) has emerged as a promising class of nucleic acid mimic because of its strong binding affinity and sequence selectivity toward DNA and RNA and resistance to enzymatic degradation by proteases and nucleases; however, the main drawbacks, as compared to other classes of oligonucleotides, are water solubility and biocompatibility. Herein we show that installation of a relatively small, hydrophilic (R)-diethylene glycol ("miniPEG", R-MP) unit at the γ-backbone transforms a randomly folded PNA into a right-handed helix. Synthesis of optically pure (R-MP)γPNA monomers is described, which can be accomplished in a few simple steps from a commercially available and relatively cheap Boc-l-serine. Once synthesized, (R-MP)γPNA oligomers are preorganized into a right-handed helix, hybridize to DNA and RNA with greater affinity and sequence selectivity, and are more water soluble and less aggregating than the parental PNA oligomers. The results presented herein have important implications for the future design and application of PNA in biology, biotechnology, and medicine, as well as in other disciplines, including drug discovery and molecular engineering.

  4. Borinic acid catalysed peptide synthesis.

    PubMed

    El Dine, Tharwat Mohy; Rouden, Jacques; Blanchet, Jérôme

    2015-11-18

    The catalytic synthesis of peptides is a major challenge in the modern organic chemistry hindered by the well-established use of stoichiometric coupling reagents. Herein, we describe for the first time that borinic acid is able to catalyse this reaction under mild conditions with an improved activity compared to our recently developed thiophene-based boronic acid. This catalyst is particularly efficient for peptide bond synthesis affording dipeptides in good yields without detectable racemization.

  5. Influence of Protein Flexibility and Peptide Conformation on Reactivity of Monoclonal Anti-Peptide Antibodies with a Protein α -helix

    NASA Astrophysics Data System (ADS)

    Fieser, Terry M.; Tainer, John A.; Geysen, H. Mario; Houghten, Richard A.; Lerner, Richard A.

    1987-12-01

    Monoclonal antibodies against an α -helical region of the iron-containing, oxygen-binding protein myohemerythrin were isolated following immunization of mice with either the whole protein or a peptide homolog of the helix. Three distinct epitopes within the myohemerythrin helix were identified. The individual residues within two of these epitopes that were essential for antibody binding were determined by measuring antibody binding to a set of peptides in which each amino acid of the epitope was replaced in turn by each of the other 19 amino acids. Hydrophilic residues that are exposed in the native conformation and buried, hydrophobic residues were both shown to be irreplaceable, suggesting their direct involvement in antibody binding. The influence of antigen conformation on antibody binding to these amphipathic epitopes was assessed by measuring the relative affinities of the antibodies for peptides, intact protein, and apoprotein. All of the antibodies bound to apoprotein better than to native protein, indicating that relaxation of the native structure by removal of the iron center increases antibody affinity for myohemerythrin. However, not all of the antibodies tested bound to peptides better than to protein, suggesting that increased antigen flexibility is not always sufficient to maximize antibody binding. Antibody binding to peptides appeared to also be influenced by the ability of the peptides to attain secondary structure at the epitopes, either alone or due to carrier influences.

  6. Conformations of Cationized Peptides. Determination of Ligand Binding Geometries by Irmpd Spectroscopy

    NASA Astrophysics Data System (ADS)

    Dunbar, Robert C.; Steill, Jeffrey; Oomens, Jos; Polfer, Nick C.

    2009-06-01

    Spectroscopic study of the conformations of metalated amino acids has mapped out in some detail the preferences for canonical (charge solvated) versus zwitterionic (salt bridge) conformations. Corresponding studies of larger peptides are now possible. Here are described results for several singly and doubly charged metal ions with dipeptides and tripeptides. Factors including ion charge, size of cation, and side chain identity and sequence are found to be conformational determinants. IRMPD spectra of the ions were acquired by irradiating the cell with infrared light from the FELIX free electron laser at wavelengths in the approximate range 500 to 1900 cm^{-1}.

  7. Conformational Restriction of Peptides Using Dithiol Bis-Alkylation.

    PubMed

    Peraro, L; Siegert, T R; Kritzer, J A

    2016-01-01

    Macrocyclic peptides are highly promising as inhibitors of protein-protein interactions. While many bond-forming reactions can be used to make cyclic peptides, most have limitations that make this chemical space challenging to access. Recently, a variety of cysteine alkylation reactions have been used in rational design and library approaches for cyclic peptide discovery and development. We and others have found that this chemistry is versatile and robust enough to produce a large variety of conformationally constrained cyclic peptides. In this chapter, we describe applications, methods, mechanistic insights, and troubleshooting for dithiol bis-alkylation reactions for the production of cyclic peptides. This method for efficient solution-phase macrocyclization is highly useful for the rapid production and screening of loop-based inhibitors of protein-protein interactions. © 2016 Elsevier Inc. All rights reserved.

  8. Major histocompatibility complex conformational epitopes are peptide specific

    PubMed Central

    1992-01-01

    Serologically distinct forms of H-2Kb are stabilized by loading cells expressing "empty" class I major histocompatibility complex (MHC) molecules with different H-2Kb binding peptides. The H-2Kb epitope recognized by monoclonal antibody (mAb) 28.8.6 was stabilized by ovalbumin (OVA) (257-264) and murine cytomegalovirus (MCMV) pp89 (168- 176) peptides, but not by vesicular stomatic virus nucleoprotein (VSV NP) (52-59) and influenza NP (Y345-360) peptides. The H-2Kb epitope recognized by mAb 34.4.20 was stabilized by VSV NP (52-59) peptide but not by OVA (257-264), MCMV pp89 (168-176), or influenza NP (Y345-360) peptides. Immunoprecipitation of H-2Kb molecules from normal cells showed that 28.8.6 and 34.4.20 epitopes were only present on a subset of all conformationally reactive H-2Kb molecules. Using alanine- substituted derivatives of the VSV peptide, the 28.8.6 epitope was completely stabilized by substitution of the first residue and partially stabilized by substitution of the third or the fifth residues in the peptides. These results indicate that distinct conformational MHC epitopes are dependent on the specific peptide that occupies the antigenic peptide binding groove on individual MHC molecules. The changes in MHC epitopes observed may also be important in understanding the diversity of T cell receptors used in an immune response and the influence of peptides on development of the T cell repertoire. PMID:1281212

  9. Conformational studies of γ-turn in pseudopeptides containing α-amino acid and conformationally constrained meta amino benzoic acid/meta nitro aniline

    NASA Astrophysics Data System (ADS)

    Dutt Konar, Anita

    2013-03-01

    Reverse turns (commonly β-turns and γ-turns), a common motif in proteins and peptides, have attracted attention due to their relevance in a wide variety of biological processes. In an attempt to artificially imitate and stabilize these turns in short acyclic peptides, a series of N-terminally protected pseudopeptides comprising of an α-amino acid and conformationally constrained meta amino benzoic acid (mABA)/meta nitro aniline (mNA) (peptides I-VI) have been synthesized. The molecules were well characterized by various spectroscopic techniques and subjected to a systematic conformational analysis. Our experimental results reveal that only pseudopeptides I and II with methyl as the sidechain, tertiary butyloxy carbonyl as the N-terminal protecting group and (mABA)/(mNA) at the C-terminus adopt γ-turn conformations in solid state as well as in solution. Even slight modification of any of the stated conditions donot support the formation of this γ-turn architecture in the solid state. Interestingly, the peptides III-V which displays extended conformation in solid state forms γ-turn structure in solution. Thus this result reflects the importance of co-operative steric interactions amongst various amino acid residues in stabilizing a particular conformation in peptides in different phases (solid and solution). This report may open a new avenue in introducing γ-turn motifs within the bioactive conformation of selected peptides.

  10. The paradox of conformational constraint in the design of Cbl(TKB)-binding peptides

    NASA Astrophysics Data System (ADS)

    Kumar, Eric A.; Chen, Qianyi; Kizhake, Smitha; Kolar, Carol; Kang, Myungshim; Chang, Chia-En A.; Borgstahl, Gloria E. O.; Natarajan, Amarnath

    2013-04-01

    Solving the crystal structure of Cbl(TKB) in complex with a pentapeptide, pYTPEP, revealed that the PEP region adopted a poly-L-proline type II (PPII) helix. An unnatural amino acid termed a proline-templated glutamic acid (ptE) that constrained both the backbone and sidechain to the bound conformation was synthesized and incorporated into the pYTPXP peptide. We estimated imposing structural constraints onto the backbone and sidechain of the peptide and preorganize it to the bound conformation in solution will yield nearly an order of magnitude improvement in activity. NMR studies confirmed that the ptE-containing peptide adopts the PPII conformation, however, competitive binding studies showed an order of magnitude loss of activity. Given the emphasis that is placed on imposing structural constraints, we provide an example to support the contrary. These results point to conformational flexibility at the interface, which have implications in the design of potent Cbl(TKB)-binding peptides.

  11. The paradox of conformational constraint in the design of Cbl(TKB)-binding peptides

    PubMed Central

    Kumar, Eric A.; Chen, Qianyi; Kizhake, Smitha; Kolar, Carol; Kang, Myungshim; Chang, Chia-en A.; Borgstahl, Gloria E. O.; Natarajan, Amarnath

    2013-01-01

    Solving the crystal structure of Cbl(TKB) in complex with a pentapeptide, pYTPEP, revealed that the PEP region adopted a poly-L-proline type II (PPII) helix. An unnatural amino acid termed a proline-templated glutamic acid (ptE) that constrained both the backbone and sidechain to the bound conformation was synthesized and incorporated into the pYTPXP peptide. We estimated imposing structural constraints onto the backbone and sidechain of the peptide and preorganize it to the bound conformation in solution will yield nearly an order of magnitude improvement in activity. NMR studies confirmed that the ptE-containing peptide adopts the PPII conformation, however, competitive binding studies showed an order of magnitude loss of activity. Given the emphasis that is placed on imposing structural constraints, we provide an example to support the contrary. These results point to conformational flexibility at the interface, which have implications in the design of potent Cbl(TKB)-binding peptides. PMID:23572190

  12. Temperature selectivity effects in reversed-phase liquid chromatography due to conformation differences between helical and non-helical peptides.

    PubMed

    Chen, Yuxin; Mant, Colin T; Hodges, Robert S

    2003-08-22

    In order to characterize the effect of temperature on the retention behaviour and selectivity of separation of polypeptides and proteins in reversed-phase high-performance liquid chromatography (RP-HPLC), the chromatographic properties of four series of peptides, with different peptide conformations, have been studied as a function of temperature (5-80 degrees C). The secondary structure of model peptides was based on either the amphipathic alpha-helical peptide sequence Ac-EAEKAAKEX(D/L)EKAAKEAEK-amide, (position X being in the centre of the hydrophobic face of the alpha-helix), or the random coil peptide sequence Ac-X(D/L)LGAKGAGVG-amide, where position X is substituted by the 19 L- or D-amino acids and glycine. We have shown that the helical peptide analogues exhibited a greater effect of varying temperature on elution behaviour compared to the random coil peptide analogues, due to the unfolding of alpha-helical structure with the increase of temperature during RP-HPLC. In addition, temperature generally produced different effects on the separations of peptides with different L- or D-amino acid substitutions within the groups of helical or non-helical peptides. The results demonstrate that variations in temperature can be used to effect significant changes in selectivity among the peptide analogues despite their very high degree of sequence homology. Our results also suggest that a temperature-based approach to RP-HPLC can be used to distinguish varying amino acid substitutions at the same site of the peptide sequence. We believe that the peptide mixtures presented here provide a good model for studying temperature effects on selectivity due to conformational differences of peptides, both for the rational development of peptide separation optimization protocols and a probe to distinguish between peptide conformations.

  13. Peptide environment specifies conformation. Helicity of hydrophobic segments compared in aqueous, organic, and membrane environments.

    PubMed

    Li, S C; Deber, C M

    1993-11-05

    Transmembrane segments in integral membrane proteins exist characteristically as helices in lipid bilayers, yet are often rich in residues considered helix-destabilizing (Val, Ile, Gly) in soluble proteins. We propose that helicity of a transmembrane segment is likely to be affected by factors other than the "intrinsic" helical propensities of its component amino acids. This hypothesis is tested by comparing the conformation(s) in aqueous, organic, membrane-mimetic (micellar), and membrane (bilayer) environments of designed model peptides with systematically altered helical propensity and/or segmental hydrophobicity. Peptides of prototypic sequence NH2-(Ser-Lys)2-Ala5-Leu6-Ala7-Ala8-Leu9-Ala10-++ +Trp11-Ala12-Leu13-Ala14- (Lys-Ser)3-OH were synthesized, which incorporate a hydrophobic core "guest" segment (residues 5-14) into a water-soluble hydrophilic host matrix. Related peptides featured substitution of Leu6,9,13-->Gly, Leu6,9,13-->Ala, and Ala7,10,14-->Gly. Circular dichroism spectra revealed that algorithms for soluble proteins correctly predicted peptide helical proclivities in aqueous solutions, but peptide helicity in organic (trifluoroethanol) solvents, membrane-mimetic SDS micelles, and negatively charged lipid bilayer vesicles, was found to be governed almost exclusively by the segmental hydrophobicity of the peptide mid-hydrophobic core segment. In related Trp fluorescence studies, peptide-membrane association was similarly correlated with extent of hydrophobic interaction.

  14. Helix and hairpin nucleation in short peptides using centrally positioned conformationally constrained dipeptide segments.

    PubMed

    Chandrappa, Siddappa; Aravinda, Subrayashastry; Raghothama, Srinivasarao; Sonti, Rajesh; Rai, Rajkishor; Harini, Veldore V; Shamala, Narayanaswamy; Balaram, Padmanabhan

    2012-04-14

    The effect of incorporation of a centrally positioned Ac(6)c-Xxx segment where Xxx = (L)Val/(D)Val into a host oligopeptide composed of l-amino acid residues has been investigated. Studies of four designed octapeptides Boc-Leu-Phe-Val-Ac(6)c-Xxx-Leu-Phe-Val-OMe (Xxx = (D)Val 1, (L)Val 2) Boc-Leu-Val-Val-Ac(6)c-Xxx-Leu-Val-Val-OMe (Xxx = (D)Val 3, (L)Val 4) are reported. Diagnostic nuclear Overhouse effects characteristic of hairpin conformations are observed for Xxx = (D)Val peptides (1 and 3) while continuous helical conformation characterized by sequential N(i)H ↔ N(i+1)H NOEs are favored for Xxx = (L)Val peptides (2 and 4) in methanol solutions. Temperature co-efficient of NH chemical shifts are in agreement with distinctly different conformational preferences upon changing the configuration of the residue at position 5. Crystal structures of peptides 2 and 4 (Xxx = (L)Val) establish helical conformations in the solid state, in agreement with the structures deduced from NMR data. The results support the design principle that centrally positioned type I β-turns may be used to nucleate helices in short peptides, while type I'β-turns can facilitate folding into β-hairpins. This journal is © The Royal Society of Chemistry 2012

  15. Peptide Suboptimal Conformation Sampling for the Prediction of Protein-Peptide Interactions.

    PubMed

    Lamiable, Alexis; Thévenet, Pierre; Eustache, Stephanie; Saladin, Adrien; Moroy, Gautier; Tuffery, Pierre

    2017-01-01

    The blind identification of candidate patches of interaction on the protein surface is a difficult task that can hardly be accomplished without a heuristic or the use of simplified representations to speed up the search. The PEP-SiteFinder protocol performs a systematic blind search on the protein surface using a rigid docking procedure applied to a limited set of peptide suboptimal conformations expected to approximate satisfactorily the conformation of the peptide in interaction. All steps rely on a coarse-grained representation of the protein and the peptide. While simple, such a protocol can help to infer useful information, assuming a critical analysis of the results. Moreover, such a protocol can be extended to a semi-flexible protocol where the suboptimal conformations are directly folded in the vicinity of the receptor.

  16. Aspartate-bond isomerization affects the major conformations of synthetic peptides.

    PubMed

    Szendrei, G I; Fabian, H; Mantsch, H H; Lovas, S; Nyéki, O; Schön, I; Otvos, L

    1994-12-15

    The aspartic acid bond changes to an beta-aspartate bond frequently as a side-reaction during peptide synthesis and often as a post-translational modification of proteins. The formation of beta-asparate bonds is reported to play a major role not only in protein metabolism, activation and deactivation, but also in pathological processes such as deposition of the neuritic plaques of Alzheimer's disease. Recently, we reported how conformational changes following the aspartic-acid-bond isomerization may help the selective aggregation and retention of the amyloid beta peptide in affected brains (Fabian et al., 1994). In the current study we used circular dichroism, Fourier-transform infrared spectroscopy, and molecular modeling to characterize the general effect of the beta-aspartate-bond formation on the conformation of five sets of synthetic model peptides. Each of the non-modified, parent peptides has one of the major secondary structures as the dominant spectroscopically determined conformation: a type I beta turn, a type II beta turn, short segments of alpha or 3(10) helices, or extended beta strands. We found that both types of turn structures are stabilized by the aspartic acid-bond isomerization. The isomerization at a terminal position did not affect the helix propensity, but placing it in mid-chain broke both the helix and the beta-pleated sheet with the formation of reverse turns. The alteration of the geometry of the lowest energy reverse turn was also supported by molecular dynamics calculations. The tendency of the aspartic acid-bond isomerization to stabilize turns is very similar to the effect of incorporating sugars into synthetic peptides and suggests a common feature of these post-translational modifications in defining the secondary structure of protein fragments.

  17. Influence of glycine residues on peptide conformation in membrane environments.

    PubMed

    Li, S C; Deber, C M

    1992-01-01

    Transmembrane (TM) segments of integral membrane proteins are putatively alpha-helical in conformation, yet their primary sequences are rich in residues known in globular proteins as helix-breakers (Gly) and beta-sheet promoters (Ile, Val, Thr). To examine the specific 2 degrees structure propensities of such residues in membrane environments, we have now designed and synthesized a series of model 20-residue peptides with "guest" hydrophobia segments embedded in "host" N- and C-terminal hydrophilic matrices. Molecular design was based on the prototypical sequence NH2-(Ser-Lys)2-Ala5-Leu6-x7-Ala8-Leu9-y10-Trp 11-Ala12-Leu13-z14-(Lys-Ser)3-OH. The 10-residue hydrophobic mid-segment 5-14 is expected to act as ca. three turns of an alpha-helix. In the present work, we compare the 20-residue peptide having three "helix-forming" Ala residues [x = y = z = Ala (peptide 3A)] to the corresponding peptide 3G (x = y = z = Gly) which contains three "helix-breaking" Gly residues. Trp was inserted to provide a measure of aromatic character typical of TM segments; Ser and Lys enhanced solubility in aqueous media. Circular dichroism studies in water, in a membrane-mimetic [sodium dodecylsulfate (SDS)], medium, and in methanol solutions, demonstrated the exquisite sensitivity of the conformations of these peptides to environment, and proved that despite its backbone flexibility, Gly can be accommodated as readily as Ala into a hydrophobic alpha-helix in a membrane. Nevertheless, the relative stability of Ala- vs. Gly-containing helices emerged in methanol solvent titration and temperature dependence experiments in SDS.(ABSTRACT TRUNCATED AT 250 WORDS)

  18. Efficient conformational sampling of peptides adsorbed onto inorganic surfaces: insights from a quartz binding peptide.

    PubMed

    Wright, Louise B; Walsh, Tiffany R

    2013-04-07

    Harnessing the properties of biomolecules, such as peptides, adsorbed on inorganic surfaces is of interest to many cross-disciplinary areas of science, ranging from biomineralisation to nanomedicine. Key to advancing research in this area is determination of the peptide conformation(s) in its adsorbed state, at the aqueous interface. Molecular simulation is one such approach for accomplishing this goal. In this respect, use of temperature-based replica-exchange molecular dynamics (T-REMD) can yield enhanced sampling of the interfacial conformations, but does so at great computational expense, chiefly because of the need to include an explicit representation of water at the interface. Here, we investigate a number of more economical variations on REMD, chiefly those based on Replica Exchange with Solvent Tempering (REST), using the aqueous quartz-binding peptide S1-(100) α-quartz interfacial system as a benchmark. We also incorporate additional implementation details specifically targeted at improving sampling of biomolecules at interfaces. We find the REST-based variants yield configurational sampling of the peptide-surface system comparable with T-REMD, at a fraction of the computational time and resource. Our findings also deliver novel insights into the binding behaviour of the S1 peptide at the quartz (100) surface that are consistent with available experimental data.

  19. En route towards the peptide γ-helix: X-ray diffraction analyses and conformational energy calculations of Adm-rich short peptides.

    PubMed

    Mazzier, Daniela; Grassi, Luigi; Moretto, Alessandro; Alemán, Carlos; Formaggio, Fernando; Toniolo, Claudio; Crisma, Marco

    2017-04-01

    We performed the solution-phase synthesis of a set of model peptides, including homo-oligomers, based on the 2-aminoadamantane-2-carboxylic acid (Adm) residue, an extremely bulky, highly lipophilic, tricyclic, achiral, C(α) -tetrasubstituted α-amino acid. In particular, for the difficult peptide coupling reaction between two Adm residues, we took advantage of the Meldal's α-azidoacyl chloride approach. Most of the synthesized Adm peptides were characterized by single-crystal X-ray diffraction analyses. The results indicate a significant propensity for the Adm residue to adopt γ-turn and γ-turn-like conformations. Interestingly, we found that a -CO-(Adm)2 -NH- sequence is folded in the crystal state into a regular, incipient γ-helix, at variance with the behavior of all of the homo-dipeptides from C(α) -tetrasubstituted α-amino acids already investigated, which tend to adopt either the β-turn or the fully extended conformation. Our density functional theory conformational energy calculations on the terminally blocked homo-peptides (n = 2-8) fully confirmed the crystal-state data, strongly supporting the view that this rigid C(α) -tetrasubstituted α-amino acid residue is largely the most effective building block for γ-helix induction, although to a limited length (anti-cooperative effect). Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

  20. Conformational analysis of peptides and glycopeptides derived from the consensus sequence for β-O-glucosylation.

    PubMed

    Somovilla, Victor J; Martinez-Sáez, Nuria; Fernández-Tejada, Alberto; de la Torre, Beatríz G; Andreu, David; Jimenez-Barbero, Jesús; L Asensio, Juan; Avenoza, Alberto; Busto, Jesus H; Corzana, Francisco; Peregrina, Jesus M

    2014-01-01

    Cys-Xxx-Ser-Xxx-Pro-Cys (Xxx= any amino acid but Pro) is the most common sequence present in naturally occurring peptides and proteins glycosylated with β-O-glucose (β-O-Glc). Taking into account the lack of studies concerning the spatial disposition of this sequence, we have synthesized and analyzed, in aqueous solution, the conformational behavior of peptides and a glycopeptide derived from the particular fragment Cys-Ala-Ser-Ser-Pro-Cys. This sequence is found in the crystal structure of the complex of blood coagulation factor VIIa with soluble tissue factor. Our studies, based on the use of NOESY experiments in combination with molecular dynamics (MD) simulations, indicate that for this particular fragment, initially characterized by a type I β-turn motif, the glycosylation with β-O-Glc forces the peptide backbone into an extended conformation. This conformation is stabilized by the presence of both hydrogen bonds and water pockets between the peptide and the sugar moieties.

  1. Conformations of heterochiral and homochiral proline-pseudoproline segments in peptides: context dependent cis-trans peptide bond isomerization.

    PubMed

    Raghothama, Srinivasarao; Raghavender, Upadhyayula Surya; Aravinda, Subrayashastry; Shamala, Narayanaswamy; Balaram, Padmanabhan

    2009-01-01

    The pseudoproline residue (Psi Pro, L-2,2-dimethyl-1,3-thiazolidine-4-carboxylic acid) has been introduced into heterochiral diproline segments that have been previously shown to facilitate the formation of beta-hairpins, containing central two and three residue turns. NMR studies of the octapeptide Boc-Leu-Phe-Val-(D)Pro-Psi Pro-Leu-Phe-Val-OMe (1), Boc-Leu-Val-Val-(D)Pro-Psi Pro-Leu-Val-Val-OMe (2), and the nonapeptide sequence Boc-Leu-Phe-Val-(D)Pro-Psi Pro-(D)Ala-Leu-Phe-Val-OMe (3) established well-registered beta-hairpin structures in chloroform solution, with the almost exclusive population of the trans conformation for the peptide bond preceding the Psi Pro residue. The beta-hairpin conformation of 1 is confirmed by single crystal X-ray diffraction. Truncation of the strand length in Boc-Val-(D)Pro-Psi Pro-Leu-OMe (4) results in an increase in the population of the cis conformer, with a cis/trans ratio of 3.65. Replacement of Psi Pro in 4 by (L)Pro in 5, results in almost exclusive population of the trans form, resulting in an incipient beta-hairpin conformation, stabilized by two intramolecular hydrogen bonds. Further truncation of the sequence gives an appreciable rise in the population of cis conformers in the tripeptide Piv-(D)Pro-Psi Pro-Leu-OMe (6). In the homochiral segment Piv-Pro-Psi Pro-Leu-OMe (7) only the cis form is observed with the NMR evidence strongly supporting a type VIa beta-turn conformation, stabilized by a 4-->1 hydrogen bond between the Piv (CO) and Leu (3) NH groups. The crystal structure of the analog peptide 7a (Piv-Pro-Psi(H,CH3)Pro-Leu-NHMe) confirms the cis peptide bond geometry for the Pro-Psi(H,CH3)Pro peptide bond, resulting in a type VIa beta-turn conformation.

  2. The polyproline II conformation in short alanine peptides is noncooperative.

    PubMed

    Chen, Kang; Liu, Zhigang; Kallenbach, Neville R

    2004-10-26

    The finding that short alanine peptides possess a high fraction of polyproline II (PII) structure (Phi=-75 degrees, Psi=+145 degrees ) at low temperature has broad implications for unfolded states of proteins. An important question concerns whether or not this structure is locally determined or cooperative. We have monitored the conformation of alanine in a series of model peptides AcGGAnGGNH2 (n=1-3) over a temperature range from -10 degrees C to +80 degrees C. Use of 15N-labeled alanine substitutions makes it possible to measure 3JalphaN coupling constants accurately over the full temperature range. Based on a 1D next-neighbor model, the cooperative parameter sigma of PII nucleation is evaluated from the coupling constant data. The finding that sigma is close to unity (1 +/- 0.2) indicates a noncooperative role for alanine in PII structure formation, consistent with statistical surveys of the Protein Data Bank that suggest that most PII structure occurs in isolated residues. Lack of cooperativity in these models implies that hydration effects that influence PII conformation in water are highly localized. Using a nuclear Overhauser effect ratio strategy to define the alanine Psi angle, we estimate that, at 40 degrees C, the time-averaged alanine conformation (Phi=-80 degrees, Psi=+170 degrees ) deviates from canonical PII structure, indicating that PII melts at high temperature. Thus, the high-temperature state of short alanine peptides seems to be an unfolded ensemble with higher distribution in the extended beta structure basin, but not a coil.

  3. Influence of silaproline on peptide conformation and bioactivity.

    PubMed

    Cavelier, Florine; Vivet, Bertrand; Martinez, Jean; Aubry, André; Didierjean, Claude; Vicherat, André; Marraud, Michel

    2002-03-27

    The analogue gamma-(dimethylsila)-proline, denoted silaproline (Sip), was synthesized in both enantiomerically pure forms by diastereoselective alkylation of a chiral glycine equivalent with use of Schöllkopf's bis-lactim ether method. The effect of replacing a proline residue in model peptides by this new proline surrogate has been examined in the crystal state by X-ray diffraction and in solution by IR absorption and NMR techniques. Silaproline and proline-containing sequences exhibit very similar conformational properties. Silaproline was also substituted for proline in a neurotensin (8-13) analogue that retained biological activity and exhibited enhanced resistance to biodegradation.

  4. Conformations of amino acids in proteins.

    PubMed

    Hovmöller, Sven; Zhou, Tuping; Ohlson, Tomas

    2002-05-01

    The main-chain conformations of 237 384 amino acids in 1042 protein subunits from the PDB were analyzed with Ramachandran plots. The populated areas of the empirical Ramachandran plot differed markedly from the classical plot in all regions. All amino acids in alpha-helices are found within a very narrow range of phi, psi angles. As many as 40% of all amino acids are found in this most populated region, covering only 2% of the Ramachandran plot. The beta-sheet region is clearly subdivided into two distinct regions. These do not arise from the parallel and antiparallel beta-strands, which have quite similar conformations. One beta region is mainly from amino acids in random coil. The third and smallest populated area of the Ramachandran plot, often denoted left-handed alpha-helix, has a different position than that originally suggested by Ramachandran. Each of the 20 amino acids has its own very characteristic Ramachandran plot. Most of the glycines have conformations that were considered to be less favoured. These results may be useful for checking secondary-structure assignments in the PDB and for predicting protein folding.

  5. Conformational contribution to thermodynamics of binding in protein-peptide complexes through microscopic simulation.

    PubMed

    Das, Amit; Chakrabarti, J; Ghosh, Mahua

    2013-03-19

    We extract the thermodynamics of conformational changes in biomacromolecular complexes from the distributions of the dihedral angles of the macromolecules. These distributions are obtained from the equilibrium configurations generated via all-atom molecular dynamics simulations. The conformational thermodynamics data we obtained for calmodulin-peptide complexes using our methodology corroborate well with the experimentally observed conformational and binding entropies. The conformational free-energy changes and their contributions for different peptide-binding regions of calmodulin are evaluated microscopically.

  6. A Monte Carlo study of peptide insertion into lipid bilayers: equilibrium conformations and insertion mechanisms.

    PubMed Central

    Maddox, Michael W; Longo, Marjorie L

    2002-01-01

    The membrane insertion behavior of two peptides, Magainin2 and M2 delta, was investigated by applying the Monte Carlo simulation technique to a theoretical model. The model included many novel aspects, such as a new semi-empirical lipid bilayer model and a new set of semi-empirical transfer energies, which reproduced the experimental insertion behavior of Magainin2 and M2 delta without parameter fitting. Additionally, we have taken into account diminished internal (intramolecular) hydrogen bonding at the N- and C-termini of helical peptides. All simulations were carried out at 305 K, above the membrane thermal phase transition temperature, and at pH 7.0. The peptide equilibrium conformations are discussed for a range of bilayers with tail polarities varying from octanol-like to alkane-like. Probability distributions of the individual amino-acid-residue positions show the dynamic nature of these equilibrium conformations. Two different insertion mechanisms for M2 delta, and a translocation mechanism for Magainin2, are described. A study of the effect of bilayer thickness on M2 delta insertion suggests a critical thickness above which insertion is unfavorable. Additionally, we did not need to use an orientational potential or array of hard cylinders to persuade M2 delta to insert perpendicular to the membrane surface. Instead, we found that diminished internal hydrogen bonding in the helical conformation anchored the termini in the headgroups and resulted in a nearly perpendicular orientation. PMID:11751313

  7. Laspartomycin, an acidic lipopeptide antibiotic with a unique peptide core.

    PubMed

    Borders, Donald B; Leese, Richard A; Jarolmen, Howard; Francis, Noreen D; Fantini, Amadeo A; Falla, Tim; Fiddes, John C; Aumelas, André

    2007-03-01

    Laspartomycin was originally isolated and characterized in 1968 as a lipopeptide antibiotic related to amphomycin. The molecular weight and structure remained unknown until now. In the present study, laspartomycin was purified by a novel calcium chelate procedure, and the structure of the major component (1) was determined. The structure of laspartomycin C (1) differs from that of amphomycin and all related antibiotics as a result of its peptide region being acidic rather than amphoteric and the amino acid branching into the side chain being diaminopropionic rather than diaminobutyric. In addition, the fatty acid side chain is 2,3-unsaturated compared to 3,4-unsaturated for amphomycin and other related antibiotics. Calcium ion addition to stabilize a particular conformer was found to be important for an enzymatic deacylation of the antibiotic. A peptide resulting from the deacylation was critical for chemical structure determination by NMR studies, which also involved addition of calcium ions to stabilize a conformer.

  8. Conformational Ensembles Explored Dynamically from Disordered Peptides Targeting Chemokine Receptor CXCR4

    PubMed Central

    Vincenzi, Marian; Costantini, Susan; Scala, Stefania; Tesauro, Diego; Accardo, Antonella; Leone, Marilisa; Colonna, Giovanni; Guillon, Jean; Portella, Luigi; Trotta, Anna Maria; Ronga, Luisa; Rossi, Filomena

    2015-01-01

    This work reports on the design and the synthesis of two short linear peptides both containing a few amino acids with disorder propensity and an allylic ester group at the C-terminal end. Their structural properties were firstly analyzed by means of experimental techniques in solution such as CD and NMR methods that highlighted peptide flexibility. These results were further confirmed by MD simulations that demonstrated the ability of the peptides to assume conformational ensembles. They revealed a network of transient and dynamic H-bonds and interactions with water molecules. Binding assays with a well-known drug-target, i.e., the CXCR4 receptor, were also carried out in an attempt to verify their biological function and the possibility to use the assays to develop new specific targets for CXCR4. Moreover, our data indicate that these peptides represent useful tools for molecular recognition processes in which a flexible conformation is required in order to obtain an interaction with a specific target. PMID:26030674

  9. Conformations of Gly(n)H+ and Ala(n)H+ peptides in the gas phase.

    PubMed Central

    Hudgins, R R; Mao, Y; Ratner, M A; Jarrold, M F

    1999-01-01

    High-resolution ion mobility measurements and molecular dynamics simulations have been used to probe the conformations of protonated polyglycine and polyalanine (Gly(n)H and Ala(n)H+, n = 3-20) in the gas phase. The measured collision integrals for both the polyglycine and the polyalanine peptides are consistent with a self-solvated globule conformation, where the peptide chain wraps around and solvates the charge located on the terminal amine. The conformations of the small peptides are governed entirely by self-solvation, whereas the larger ones have additional backbone hydrogen bonds. Helical conformations, which are stable for neutral Alan peptides, were not observed in the experiments. Molecular dynamics simulations for Ala(n)H+ peptides suggest that the charge destabilizes the helix, although several of the low energy conformations found in the simulations for the larger Ala(n)H+ peptides have small helical regions. PMID:10049339

  10. Cyclic Constraints on Conformational Flexibility in γ-PEPTIDES: Conformation-Specific IR and UV Spectroscopy

    NASA Astrophysics Data System (ADS)

    Walsh, Patrick S.; Kusaka, Ryoji; Zwier, Timothy S.; Fisher, Brian F.; Gellman, Samuel H.

    2013-06-01

    Spectroscopic studies of flexible peptides in the gas phase can provide insight to their inherent structural preferences in the absence of solvent. Recently, there has been increased attention paid to synthetic foldamers containing non-natural residues that can be specifically engineered to robustly form particular secondary structures. These engineered peptides have potential in therapeutic drug design because they are resistant to enzymatic degradation. Specifically, the Gellman group has synthesized a γ-peptide with a six membered cyclic constraint in the γ^{4}-γ^{3} position and an ethyl group at the γ^{2} position (γ_{ACHC}). The three stereocenters have a well-defined chirality [S,S,S]. These two features constrain the relative orientation of adjacent amide groups, thereby favoring a particular "pitch" to the turn. Solution phase results indicate that constrained γ-peptides induce the formation of a 14-helix. Ac-γ_{ACHC}-NHBz, its monohydrate and Ac-γ_{ACHC}-γ_{ACHC}-NHBz have been studied using ultraviolet (UV) and infrared (IR) double-resonance methods to obtain conformation-specific spectra under jet-cooled conditions in the gas phase. IR spectra in the hydride stretch (3300-3750 cm^{-1}), amide I/II and OH bend (1400-1800 cm^{-1}) were recorded and compared to predictions using density functional methods (DFT) and harmonic frequency calculations. We will compare the present results on constrained γ-peptides with corresponding results on unconstrained analogs. Data obtained for the monohydrated water cluster of Ac-γ_{ACHC}-NHBz will also be presented, including assignment of the water bend fundamental, which appears in the midst of transitions due to the amide II vibrations. L. Guo, W. Zhang, A. G. Reidenbach, M. W. Giuliano, I. A. Guzei, L. C. Spencer and S. H. Gellman Angew. Chem. Int. Ed. 2011, 50, 5843-5846

  11. N-methylation of N(alpha)-acylated, fully C(alpha)-methylated, linear, folded peptides: synthetic and conformational aspects.

    PubMed

    Moretto, Alessandro; Crisma, Marco; Kaptein, Bernard; Broxterman, Quirinus B; Toniolo, Claudio

    2006-01-01

    Peptides characterized by single or multiple N-methylated, C(alpha)-trisubstituted (e.g., protein) amino acids are of great interest in medicinal chemistry. Several naturally occurring peptides, remarkably stable to enzymatic attacks, are based on N-methylated residues. The classical conditions (CH(3)I/Ag(2)O in DMF, 24 h, room temperature) for N-methylation of the peptide function are useful tools for distinguishing solvent exposed from intramolecularly H-bonded -CO-NH- groups in peptides. In this work we have extended this reaction to N(alpha)-acylated, linear peptides based exclusively on helicogenic C(alpha)-tetrasubstituted alpha-amino acids, e.g., Aib (alpha-aminoisobutyric acid) or (alphaMe)Nva (C(alpha)-methyl norvaline) residues. Under the experimental conditions used, only amide monomethylation (on the N-terminal, acylated, residue) takes place. Methylation of internal peptide groups linking two C(alpha)-tetrasubstituted residues was not observed. Our FT-IR absorption, NMR, and X-ray diffraction investigations support the view that the beta-turn and 3(10)-helical conformations preferred by the original peptides are not dramatically perturbed in the derivatives monomethylated at position 1. In particular, the tertiary amide bonds are trans. Conversely, the packing modes in the crystals are strongly influenced by the reduction of the number of H-bonding donors. The MeXxx-Xxx peptide bond is readily disrupted under mild acidic conditions. (c) 2006 Wiley Periodicals, Inc.

  12. Using Cooperatively Folded Peptides To Measure Interaction Energies and Conformational Propensities.

    PubMed

    Ardejani, Maziar S; Powers, Evan T; Kelly, Jeffery W

    2017-08-15

    The rates and equilibria of the folding of biopolymers are determined by the conformational preferences of the subunits that make up the sequence of the biopolymer and by the interactions that are formed in the folded state in aqueous solution. Because of the centrality of these processes to life, quantifying conformational propensities and interaction strengths is vitally important to understanding biology. In this Account, we describe our use of peptide model systems that fold cooperatively yet are small enough to be chemically synthesized to measure such quantities. The necessary measurements are made by perturbing an interaction or conformation of interest by mutation and measuring the difference between the folding free energies of the wild type (in which the interaction or conformation is undisturbed) and the mutant model peptides (in which the interaction has been eliminated or the conformational propensities modified). With the proper controls and provided that the peptide model system in question folds via a two-state process, these folding free energy differences can be accurate measures of interaction strengths or conformational propensities. This method has the advantage of having high sensitivity and high dynamic range because the energies of interest are coupled to folding free energies, which can be measured with precisions on the order of a few tenths of a kilocalorie by well-established biophysical methods, like chaotrope or thermal denaturation studies monitored by fluorescence or circular dichroism. In addition, because the model peptides can be chemically synthesized, the full arsenal of natural and unnatural amino acids can be used to tune perturbations to be as drastic or subtle as desired. This feature is particularly noteworthy because it enables the use of analytical tools developed for physical organic chemistry, especially linear free energy relationships, to decompose interaction energies into their component parts to obtain a deeper

  13. Gas-Phase Hydrogen-Deuterium Exchange Labeling of Select Peptide Ion Conformer Types: a Per-Residue Kinetics Analysis.

    PubMed

    Khakinejad, Mahdiar; Kondalaji, Samaneh Ghassabi; Tafreshian, Amirmahdi; Valentine, Stephen J

    2015-07-01

    The per-residue, gas-phase hydrogen deuterium exchange (HDX) kinetics for individual amino acid residues on selected ion conformer types of the model peptide KKDDDDDIIKIIK have been examined using ion mobility spectrometry (IMS) and HDX-tandem mass spectrometry (MS/MS) techniques. The [M + 4H](4+) ions exhibit two major conformer types with collision cross sections of 418 Å(2) and 446 Å(2); the [M + 3H](3+) ions also yield two different conformer types having collision cross sections of 340 Å(2) and 367 Å(2). Kinetics plots of HDX for individual amino acid residues reveal fast- and slow-exchanging hydrogens. The contributions of each amino acid residue to the overall conformer type rate constant have been estimated. For this peptide, N- and C-terminal K residues exhibit the greatest contributions for all ion conformer types. Interior D and I residues show decreased contributions. Several charge state trends are observed. On average, the D residues of the [M + 3H](3+) ions show faster HDX rate contributions compared with [M + 4H](4+) ions. In contrast the interior I8 and I9 residues show increased accessibility to exchange for the more elongated [M + 4H](4+) ion conformer type. The contribution of each residue to the overall uptake rate showed a good correlation with a residue hydrogen accessibility score model calculated using a distance from charge site and initial incorporation site for nominal structures obtained from molecular dynamic simulations (MDS).

  14. Gas-Phase Hydrogen-Deuterium Exchange Labeling of Select Peptide Ion Conformer Types: a Per-Residue Kinetics Analysis

    NASA Astrophysics Data System (ADS)

    Khakinejad, Mahdiar; Kondalaji, Samaneh Ghassabi; Tafreshian, Amirmahdi; Valentine, Stephen J.

    2015-07-01

    The per-residue, gas-phase hydrogen deuterium exchange (HDX) kinetics for individual amino acid residues on selected ion conformer types of the model peptide KKDDDDDIIKIIK have been examined using ion mobility spectrometry (IMS) and HDX-tandem mass spectrometry (MS/MS) techniques. The [M + 4H]4+ ions exhibit two major conformer types with collision cross sections of 418 Å2 and 446 Å2; the [M + 3H]3+ ions also yield two different conformer types having collision cross sections of 340 Å2 and 367 Å2. Kinetics plots of HDX for individual amino acid residues reveal fast- and slow-exchanging hydrogens. The contributions of each amino acid residue to the overall conformer type rate constant have been estimated. For this peptide, N- and C-terminal K residues exhibit the greatest contributions for all ion conformer types. Interior D and I residues show decreased contributions. Several charge state trends are observed. On average, the D residues of the [M + 3H]3+ ions show faster HDX rate contributions compared with [M + 4H]4+ ions. In contrast the interior I8 and I9 residues show increased accessibility to exchange for the more elongated [M + 4H]4+ ion conformer type. The contribution of each residue to the overall uptake rate showed a good correlation with a residue hydrogen accessibility score model calculated using a distance from charge site and initial incorporation site for nominal structures obtained from molecular dynamic simulations (MDS).

  15. Effect of Inactivating Mutations on Peptide Conformational Ensembles: The Plant Polypeptide Hormone Systemin.

    PubMed

    Chowdhury, Saikat Dutta; Sarkar, Aditya K; Lahiri, Ansuman

    2016-07-25

    As part of their basal immune mechanism against insect/herbivore attacks, plants have evolved systemic response mechanisms. Such a systemic wound response in tomato was found to involve an 18 amino acid polypeptide called systemin, the first polypeptide hormone to be discovered in plants. Systematic alanine scanning and deletion studies showed differential modulation in its activity, particularly a major loss of function due to alanine substitution at positions 13 and 17 and less extentive loss of function due to substitution at position 12. We have studied the conformational ensembles of wild-type systemin along with its 17 variants by carrying out a total of 5.76 μs of replica-exchange molecular dynamics simulation in an implicit solvent environment. In our simulations, wild-type systemin showed a lack of α-helical and β-sheet structures, in conformity with earlier circular dichroism and NMR data. On the other hand, two regions containing diproline segments showed a tendency to adopt polyproline II structures. Examination of conformational ensembles of the 17 variants revealed a change in the population distributions, suggesting a less flexible structure for alanine substitutions at positions 12 and 13 but not for position 17. Combined with the experimental observations that positions 1-14 of systemin are important for the formation of the peptide-receptor complex, this leads to the hypothesis that loss of conformational flexibility may play a role in the loss of activity of systemin due to the P12A and P13A substitutions, while T17A deactivation probably occurs for a different reason, most likely the loss of the threonine phosphorylation site. We also indicate possible structural reasons why the substitution of the prolines at positions 12 and 13 leads to a loss of conformational freedom in the peptide.

  16. Solvent and conformation dependence of amide I vibrations in peptides and proteins containing proline.

    PubMed

    Roy, Santanu; Lessing, Joshua; Meisl, Georg; Ganim, Ziad; Tokmakoff, Andrei; Knoester, Jasper; Jansen, Thomas L C

    2011-12-21

    We present a mixed quantum-classical model for studying the amide I vibrational dynamics (predominantly CO stretching) in peptides and proteins containing proline. There are existing models developed for determining frequencies of and couplings between the secondary amide units. However, these are not applicable to proline because this amino acid has a tertiary amide unit. Therefore, a new parametrization is required for infrared-spectroscopic studies of proteins that contain proline, such as collagen, the most abundant protein in humans and animals. Here, we construct the electrostatic and dihedral maps accounting for solvent and conformation effects on frequency and coupling for the proline unit. We examine the quality and the applicability of these maps by carrying out spectral simulations of a number of peptides with proline in D(2)O and compare with experimental observations.

  17. Conformation of dehydropentapeptides containing four achiral amino acid residues – controlling the role of L-valine

    PubMed Central

    Krzciuk-Gula, Joanna; Makowski, Maciej; Latajka, Rafał; Kafarski, Paweł

    2014-01-01

    Summary Structural studies of pentapeptides containing an achiral block, built from two dehydroamino acid residues (ΔZPhe and ΔAla) and two glycines, as well as one chiral L-Val residue were performed using NMR spectroscopy. The key role of the L-Val residue in the generation of the secondary structure of peptides is discussed. The obtained results suggest that the strongest influence on the conformation of peptides arises from a valine residue inserted at the C-terminal position. The most ordered conformation was found for peptide Boc-Gly-ΔAla-Gly-ΔZPhe-Val-OMe (3), which adopts a right-handed helical conformation. PMID:24778717

  18. All-atom molecular dynamics study of EAK16 peptide: the effect of pH on single-chain conformation, dimerization and self-assembly behavior.

    PubMed

    Emamyari, Soheila; Fazli, Hossein

    2014-05-01

    Single-chain equilibrium conformation and dimerization of the three types of ionic EAK16 peptide are studied under three pH conditions using all-atom molecular dynamics simulations. It is found that both the single-chain conformation and the dimerization process of EAK16-IV are considerably different from those of the two other types, EAK16-I and EAK16-II. The value of pH is found to have a stronger effect on the single-chain conformation and dimerization of EAK16-IV. It is shown that in addition to the charge pattern on the peptide chains, the size of the side chains of the charged amino acids plays role in the conformation of the peptide chains and their dimerization. The results shed light on the pH-dependent self-assembly behavior of EAK16 peptide in the bulk solution, which has been reported in the literature.

  19. Triple-helical peptides: an approach to collagen conformation, stability, and self-association.

    PubMed

    Brodsky, Barbara; Thiagarajan, Geetha; Madhan, Balaraman; Kar, Karunakar

    2008-05-01

    Peptides have been an integral part of the collagen triple-helix structure story, and have continued to serve as useful models for biophysical studies and for establishing biologically important sequence-structure-function relationships. High resolution structures of triple-helical peptides have confirmed the basic Ramachandran triple-helix model and provided new insights into the hydration, hydrogen bonding, and sequence dependent helical parameters in collagen. The dependence of collagen triple-helix stability on the residues in its (Gly-X-Y)(n) repeating sequence has been investigated by measuring melting temperatures of host-guest peptides and an on-line collagen stability calculator is now available. Although the presence of Gly as every third residue is essential for an undistorted structure, interruptions in the repeating (Gly-X-Y)(n) amino acid sequence pattern are found in the triple-helical domains of all nonfibrillar collagens, and are likely to play a role in collagen binding and degradation. Peptide models indicate that small interruptions can be incorporated into a rod-like triple-helix with a highly localized effect, which perturbs hydrogen bonds and places the standard triple-helices on both ends out of register. In contrast to natural interruptions, missense mutations which replace one Gly in a triple-helix domain by a larger residue have pathological consequences, and studies on peptides containing such Gly substitutions clarify their effect on conformation, stability, and folding. Recent studies suggest peptides may also be useful in defining the basic principles of collagen self-association to the supramolecular structures found in tissues.

  20. Multiple gas-phase conformations of proline-containing peptides: Is it always cis/trans isomerization?

    PubMed Central

    Lietz, Christopher B.; Chen, Zhengwei; Son, Chang Yun; Pang, Xueqin; Cui, Qiang; Li, Lingjun

    2016-01-01

    Ion mobility-mass spectrometry (IM-MS) is often employed to look at the secondary, tertiary, and quaternary structure of naked peptides and proteins in the gas-phase. Recently, it has offered a unique glimpse into proline-containing peptides and their cis/trans Xxx-Pro isomers. An experimental “signature” has been identified wherein a proline-containing peptide has its Pro residues substituted with another amino acid and the presence or absence of conformations in the IM-MS spectra are observed. Despite high probability that one could attribute these conformations to cis/trans isomers, it is also possible that cis/trans isomers are not the cause of the additional conformations in proline-containing peptides. However, the experimental evidence of such a system has not been demonstrated or reported. Herein, we present the IM-MS analysis of Neuropeptide Y’s wild-type (WT) signal sequence and Leu7Pro (L7P) mutant. Although comparison of arrival times and collision cross sections of [M+4H]4+ ions yield the cis/trans “signature”, molecular dynamics indicates that a cis-Pro7 is not very stable and that trans-Pro7 conformations of the same cross section arise with equal frequency. We believe this work further underscores the importance of theoretical calculations in IM-MS structural assignments. PMID:27434776

  1. Multiple gas-phase conformations of proline-containing peptides: is it always cis/trans isomerization?

    PubMed

    Lietz, Christopher B; Chen, Zhengwei; Yun Son, Chang; Pang, Xueqin; Cui, Qiang; Li, Lingjun

    2016-08-02

    Ion mobility-mass spectrometry (IM-MS) is often employed to look at the secondary, tertiary, and quaternary structures of naked peptides and proteins in the gas-phase. Recently, it has offered a unique glimpse into proline-containing peptides and their cis/trans Xxx-Pro isomers. An experimental "signature" has been identified wherein a proline-containing peptide has its Pro residues substituted with another amino acid and the presence or absence of conformations in the IM-MS spectra is observed. Despite the high probability that one could attribute these conformations to cis/trans isomers, it is also possible that cis/trans isomers are not the cause of the additional conformations in proline-containing peptides. However, the experimental evidence of such a system has not been demonstrated or reported. Herein, we present the IM-MS analysis of Neuropeptide Y's wild-type (WT) signal sequence and Leu7Pro (L7P) mutant. Although comparison of arrival times and collision cross-sections of [M + 4H](4+) ions yields the cis/trans "signature", molecular dynamics indicates that a cis-Pro7 is not very stable and that trans-Pro7 conformations of the same cross-section arise with equal frequency. We believe that this work further underscores the importance of theoretical calculations in IM-MS structural assignments.

  2. Local interactions in peptides. 1H-1H, 13C-H coupling constants and calculations for the conformational analysis of N-acetyl-N'-methylamides of aliphatic amino acids.

    PubMed

    Fermandjian, S; Sakarellos, C; Aumelas, A; Toma, F; Gresh, N

    1990-05-01

    We report the results of a joint NMR and theoretical investigation devoted to the conformational properties of N-acetyl-N'-methylamides of aliphatic amino acids with side chains of increasing bulkiness: Gly, Ala, Leu, Ile, and tert.Leu. In this series, determination of the coupling constants 3JHNC alpha H together with the coupling constants 3JC'NC alpha H (thanks to specific carbon-13 labeling of the N-acetyl carbonyl group) led to the derivation of alternative A, B, and C parameters in a Karplus-type relation expressing the dependence of 3JC'NC alpha H upon the phi dihedral angle. The value of the latter is found to increase regularly following the increase of the side-chain bulkiness. The theoretical conformational analysis is performed by applying the SIBFA procedure, which uses empirical formulas based on ab initio SCF computations. The conformational energy maps illustrate the progressive distortion of the backbone conformation incurred in the series Gly to tert.Leu. Theoretical values computed for 3JHNC alpha H and 3JC'NC alpha H are found to be in a good quantitative agreement with the experimental ones.

  3. Peptide conformation in gas phase probed by collision-induced dissociation and its correlation to conformation in condensed phases.

    PubMed

    Zhang, Zhongqi; Bordas-Nagy, Joseph

    2006-06-01

    A kinetic peptide fragmentation model for quantitative prediction of peptide CID spectra in an ion trap mass spectrometer has been reported recently. When applying the model to predict the CID spectra of large peptides, it was often found that the predicted spectra differed significantly from their experimental spectra, presumably due to noncovalent interactions in these large polypeptides, which are not considered in the fragmentation model. As a result, site-specific quantitative information correlated to the secondary/tertiary structure of an ionized peptide may be extracted from its CID spectrum. To extract this information, the kinetic peptide fragmentation model was modified by incorporating conformation-related parameters. These parameters are optimized for best fit between the predicted and the experimental spectrum. A conformational stability map is then generated from these conformation-related parameters. Analysis of a few bioactive alpha-helical peptides including melittin, glucagon and neuropeptide Y by this technique demonstrated that their stability maps in the gas phase correlate strongly to their secondary structures in the condensed phases.

  4. PEGylated nanoparticles bind to and alter amyloid-beta peptide conformation: toward engineering of functional nanomedicines for Alzheimer's disease.

    PubMed

    Brambilla, Davide; Verpillot, Romain; Le Droumaguet, Benjamin; Nicolas, Julien; Taverna, Myriam; Kóňa, Juraj; Lettiero, Barbara; Hashemi, S Hossein; De Kimpe, Line; Canovi, Mara; Gobbi, Marco; Nicolas, Valérie; Scheper, Wiep; Moghimi, S Moein; Tvaroška, Igor; Couvreur, Patrick; Andrieux, Karine

    2012-07-24

    We have demonstrated that the polyethylene glycol (PEG) corona of long-circulating polymeric nanoparticles (NPs) favors interaction with the amyloid-beta (Aβ(1-42)) peptide both in solution and in serum. The influence of PEGylation of poly(alkyl cyanoacrylate) and poly(lactic acid) NPs on the interaction with monomeric and soluble oligomeric forms of Aβ(1-42) peptide was demonstrated by capillary electrophoresis, surface plasmon resonance, thioflavin T assay, and confocal microscopy, where the binding affected peptide aggregation kinetics. The capture of peptide by NPs in serum was also evidenced by fluorescence spectroscopy and ELISA. Moreover, in silico and modeling experiments highlighted the mode of PEG interaction with the Aβ(1-42) peptide and its conformational changes at the nanoparticle surface. Finally, Aβ(1-42) peptide binding to NPs affected neither complement activation in serum nor apolipoprotein-E (Apo-E) adsorption from the serum. These observations have crucial implications in NP safety and clearance kinetics from the blood. Apo-E deposition is of prime importance since it can also interact with the Aβ(1-42) peptide and increase the affinity of NPs for the peptide in the blood. Collectively, our results suggest that these engineered long-circulating NPs may have the ability to capture the toxic forms of the Aβ(1-42) peptide from the systemic circulation and potentially improve Alzheimer's disease condition through the proposed "sink effect".

  5. The Inherent Conformational Preferences of Glutamine-Containing Peptides: the Role for Side-Chain Backbone Hydrogen Bonds

    NASA Astrophysics Data System (ADS)

    Walsh, Patrick S.; McBurney, Carl; Gellman, Samuel H.; Zwier, Timothy S.

    2015-06-01

    Glutamine is widely known to be found in critical regions of peptides which readily fold into amyloid fibrils, the structures commonly associated with Alzheimer's disease and glutamine repeat diseases such as Huntington's disease. Building on previous single-conformation data on Gln-containing peptides containing an aromatic cap on the N-terminus (Z-Gln-OH and Z-Gln-NHMe), we present here single-conformation UV and IR spectra of Ac-Gln-NHBn and Ac-Ala-Gln-NHBn, with its C-terminal benzyl cap. These results point towards side-chain to backbone hydrogen bonds dominating the structures observed in the cold, isolated environment of a molecular beam. We have identified and assigned three main conformers for Ac-Gln-NHBn all involving primary side-chain to backbone interactions. Ac-Ala-Gln-NHBn extends the peptide chain by one amino acid, but affords an improvement in the conformational flexibility. Despite this increase in the flexibility, only a single conformation is observed in the gas-phase: a structure which makes use of both side-chain-to-backbone and backbone-to-backbone hydrogen bonds.

  6. Hydrogen exchange-mass spectrometry measures stapled peptide conformational dynamics and predicts pharmacokinetic properties.

    PubMed

    Shi, Xiangguo Eric; Wales, Thomas E; Elkin, Carl; Kawahata, Noriyuki; Engen, John R; Annis, D Allen

    2013-12-03

    Peptide drugs have traditionally suffered from poor pharmacokinetic properties due to their conformational flexibility and the interaction of proteases with backbone amide bonds. "Stapled Peptides" are cyclized using an all-hydrocarbon cross-linking strategy to reinforce their α-helical conformation, yielding improved protease resistance and drug-like properties. Here we demonstrate that hydrogen exchange-mass spectrometry (HX-MS) effectively probes the conformational dynamics of Stapled Peptides derived from the survivin-borealin protein-protein interface and predicts their susceptibility to proteolytic degradation. In Stapled Peptides, amide exchange was reduced by over five orders-of-magnitude versus the native peptide sequence depending on staple placement. Furthermore, deuteration kinetics correlated directly with rates of proteolysis to reveal the optimal staple placement for improved drug properties.

  7. Importance of asparagine on the conformational stability and chemical reactivity of selected anti-inflammatory peptides

    NASA Astrophysics Data System (ADS)

    Soriano-Correa, Catalina; Barrientos-Salcedo, Carolina; Campos-Fernández, Linda; Alvarado-Salazar, Andres; Esquivel, Rodolfo O.

    2015-08-01

    Inflammatory response events are initiated by a complex series of molecular reactions that generate chemical intermediaries. The structure and properties of peptides and proteins are determined by the charge distribution of their side chains, which play an essential role in its electronic structure and physicochemical properties, hence on its biological functionality. The aim of this study was to analyze the effect of changing one central amino acid, such as substituting asparagine for aspartic acid, from Cys-Asn-Ser in aqueous solution, by assessing the conformational stability, physicochemical properties, chemical reactivity and their relationship with anti-inflammatory activity; employing quantum-chemical descriptors at the M06-2X/6-311+G(d,p) level. Our results suggest that asparagine plays a more critical role than aspartic acid in the structural stability, physicochemical features, and chemical reactivity of these tripeptides. Substituent groups in the side chain cause significant changes on the conformational stability and chemical reactivity, and consequently on their anti-inflammatory activity.

  8. Effect of graphene oxide on the conformational transitions of amyloid beta peptide: A molecular dynamics simulation study.

    PubMed

    Baweja, Lokesh; Balamurugan, Kanagasabai; Subramanian, Venkatesan; Dhawan, Alok

    2015-09-01

    The interactions between nanomaterials (NMs) and amyloid proteins are central to the nanotechnology-based diagnostics and therapy in neurodegenerative disorders such as Alzheimer's and Parkinson's. Graphene oxide (GO) and its derivatives have shown to modulate the aggregation pattern of disease causing amyloid beta (Aβ) peptide. However, the mechanism is still not well understood. Using molecular dynamics simulations, the effect of graphene oxide (GO) and reduced graphene oxide (rGO) having carbon:oxygen ratio of 4:1 and 10:1, respectively, on the conformational transitions (alpha-helix to beta-sheet) and the dynamics of the peptide was investigated. GO and rGO decreased the beta-strand propensity of amino acid residues in Aβ. The peptide displayed different modes of adsorption on GO and rGO. The adsorption on GO was dominated by electrostatic interactions, whereas on rGO, both van der Waals and electrostatic interactions contributed in the adsorption of the peptide. Our study revealed that the slight increase in the hydrophobic patches on rGO made it more effective inhibitor of conformational transitions in the peptide. Alpha helix-beta sheet transition in Aβ peptide could be one of the plausible mechanism by which graphene oxide may inhibit amyloid fibrillation.

  9. Tetrazole acetic acid: Tautomers, conformers, and isomerization

    SciTech Connect

    Araujo-Andrade, C.; Reva, I. Fausto, R.

    2014-02-14

    Monomers of (tetrazol-5-yl)-acetic acid (TAA) were obtained by sublimation of the crystalline compound and the resulting vapors were isolated in cryogenic nitrogen matrices at 13 K. The conformational and tautomeric composition of TAA in the matrix was characterized by infrared spectroscopy and vibrational calculations carried out at the B3LYP/6-311++G(d,p) level. TAA may adopt two tautomeric modifications, 1H- and 2H-, depending on the position of the annular hydrogen atom. Two-dimensional potential energy surfaces (PESs) of TAA were theoretically calculated at the MP2/6-311++G(d,p) level, for each tautomer. Four and six symmetry-unique minima were located on these PESs, for 1H- and 2H-TAA, respectively. The energetics of the detected minima was subsequently refined by calculations at the QCISD level. Two 1H- and three 2H-conformers fall within the 0–8 kJ mol{sup −1} energy range and should be appreciably populated at the sublimation temperature (∼330 K). Observation of only one conformer for each tautomer (1ccc and 2pcc) is explained in terms of calculated barriers to conformational rearrangements. All conformers with the cis O=COH moiety are separated by low barriers (less than 10 kJ mol{sup −1}) and collapse to the most stable 1ccc (1H-) and 2pcc (2H-) forms during deposition of the matrix. On the trans O=COH surfaces, the relative energies are very high (between 12 and 27 kJ mol{sup −1}). The trans forms are not thermally populated at the sublimation conditions and were not detected in matrices. One high-energy form in each tautomer, 1cct (1H-) and 2pct (2H-), was found to differ from the most stable form only by rotation of the OH group and separated from other forms by high barriers. This opened a perspective for their stabilization in a matrix. 1cct and 2pct were generated in the matrices selectively by means of narrow-band near-infrared (NIR) irradiations of the samples at 6920 and 6937 cm{sup −1}, where the first OH stretching overtone

  10. The RNA annealing mechanism of the HIV-1 Tat peptide: conversion of the RNA into an annealing-competent conformation

    PubMed Central

    Doetsch, Martina; Fürtig, Boris; Gstrein, Thomas; Stampfl, Sabine; Schroeder, Renée

    2011-01-01

    The annealing of nucleic acids to (partly) complementary RNA or DNA strands is involved in important cellular processes. A variety of proteins have been shown to accelerate RNA/RNA annealing but their mode of action is still mainly uncertain. In order to study the mechanism of protein-facilitated acceleration of annealing we selected a short peptide, HIV-1 Tat(44–61), which accelerates the reaction efficiently. The activity of the peptide is strongly regulated by mono- and divalent cations which hints at the importance of electrostatic interactions between RNA and peptide. Mutagenesis of the peptide illustrated the dominant role of positively charged amino acids in RNA annealing—both the overall charge of the molecule and a precise distribution of basic amino acids within the peptide are important. Additionally, we found that Tat(44–61) drives the RNA annealing reaction via entropic rather than enthalpic terms. One-dimensional-NMR data suggest that the peptide changes the population distribution of possible RNA structures to favor an annealing-prone RNA conformation, thereby increasing the fraction of colliding RNA molecules that successfully anneal. PMID:21297117

  11. Conformation Analysis of Peptides Derived from Laminin Alpha 1-2 Chain Using Molecular Dynamics Simulation

    NASA Astrophysics Data System (ADS)

    Yamada, Hironao; Fukuda, Masaki; Miyakawa, Takeshi; Morikawa, Ryota; Takasu, Masako

    Laminin is one of the components of the basement membrane and has diverse biological activities. Several functional peptides (EF1-EF5) are identified from LG4 modules of laminin alpha 1-5 chains. Thus, we perform conformation analysis of EF1 and EF2 using molecular dynamics simulations. In this study, we perform structure sampling with NPT ensemble (300 K, 1 bar). Our results show that EF1 peptide has β-sheet structure in water, and EF2 peptide does not have. Likewise, the EF2 peptide has unstable structure compared with the EF1 peptide in water.

  12. Conformational stability studies of a stapled hexa-β3-peptide library.

    PubMed

    Gopalan, Romila D; Del Borgo, Mark P; Bergman, Ylva E; Unabia, Sharon; Mulder, Roger J; Wilce, Matthew C J; Wilce, Jacqueline A; Aguilar, Marie-Isabel; Perlmutter, Patrick

    2012-03-07

    A library of 14-helical hexa β(3)-peptides was synthesized in order to determine the influence of sequence variation as well as staple size and location on conformational stability. From this study we show that appropriately stapled hexa-β(3)-peptides can allow for a number of variations without significant perturbation of the 14-helix.

  13. Structure and conformation of peptides at air/aqueous interface and their impact on interfacial water structure

    NASA Astrophysics Data System (ADS)

    Chandra Jena, Kailash; Tomar, Deepak

    Process of protein folding is very essential for the proper functioning of the protein molecules at membrane surface and other organelles. Understanding the process of protein folding at various biological relevant aqueous interfaces are very important to understand various complicated chemical and physical processes relevant to chemistry, physics, and medicine. The building blocks of proteins molecules are amino acids and the chemistry of each amino acid is very different; as a consequence their sequence plays an important role for various conformations upon adsorption for the protein molecules. In the present study, we have investigated the interfacial structure and conformation of two amino acids (L-Proline and L-Tyrosine) and peptide molecules formed from these two amino acids (L-Tyr-Pro). We have used sum frequency generation (SFG) vibrational spectroscopy to probe the air/aqueous interface. We have studied the impact of adsorption of the amino acids and the peptide molecules on the interfacial water structure by slowly varying concentration and ionic strength of the solutions. Our preliminary result shows a huge impact of the adsorption process of peptide molecules on the hydrogen bonding environment of interfacial structure of water. Indian Institute of Technology Ropar, Nangal Road, Rupnagar, Punjab-140001.

  14. Antibiotic gold: tethering of antimicrobial peptides to gold nanoparticles maintains conformational flexibility of peptides and improves trypsin susceptibility.

    PubMed

    Wadhwani, Parvesh; Heidenreich, Nico; Podeyn, Benjamin; Bürck, Jochen; Ulrich, Anne S

    2017-03-09

    Peptide-coated nanoparticles are valuable tools for diverse biological applications, such as drug delivery, molecular recognition, and antimicrobial action. The functionalization of pre-fabricated nanoparticles with free peptides in solution is inefficient either due to aggregation of the particles or due to the poor ligand exchange reaction. Here, we present a one-pot synthesis for preparing gold nanoparticles with a homogeneous distribution that are covered in situ with cationic peptides in a site-selective manner via Cys-residue at the N-terminus. Five representative peptides were selected, which are known to perturb cellular membranes and exert their antimicrobial and/or cell penetrating activity by folding into amphiphilic α-helical structures. When tethered to the nanoparticles at a single site, all peptides were found to switch their conformation from unordered state (in aqueous buffers) to their functionally relevant α-helical conformation in the presence of model membranes, as shown by circular dichroism spectroscopy. The conjugated peptides also maintained the same antibacterial activity as in the free form. Most importantly, when tethered to the gold nanoparticles the peptides showed an enormous increase in stability against trypsin digestion compared to the free forms, leading to a dramatic improvement of their lifetimes and activities. These findings suggest that site-selective surface tethering of peptides to gold nanoparticles has several advantages: (i) it does not prevent the peptides from folding into their biologically active conformation, (ii) such conjugation protects the peptides against protease digestion, and (iii) this way it is possible to prepare stable, water soluble antimicrobial nanoparticles as promising antibacterial agents.

  15. Facile synthesis of peptide nucleic acids and peptide nucleic acid-peptide conjugates on an automated peptide synthesizer.

    PubMed

    Joshi, Rajendra; Jha, Deepti; Su, Wu; Engelmann, Joern

    2011-01-01

    Peptide nucleic acids (PNAs) are DNA mimics with a neutral peptide backbone instead of the negatively charged sugar phosphates. PNAs exhibit several attractive features such as high chemical and thermal stability, resistance to enzymatic degradation, and stable binding to their RNA or DNA targets in a sequence-specific manner. Therefore, they are widely used in molecular diagnosis of antisense-targeted therapeutic drugs or probes and in pharmaceutical applications. However, the main hindrance to the effective use of PNAs is their poor uptake by cells as well as the difficult and laborious chemical synthesis. In order to achieve an efficient delivery of PNAs into cells, there are already many published reports of peptides being used for transport across the cell membrane. In this protocol, we describe the automated as well as cost-effective semi-automated synthesis of PNAs and PNA-peptide constructs on an automated peptide synthesizer. The facile synthesis of PNAs will be helpful in generating PNA libraries usable, e.g. for high-throughput screening in biomolecular studies. Efficient synthetic schemes, the automated procedure, the reduced consumption of costly reagents, and the high purity of the products are attractive features of the reported procedure. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd.

  16. Conformation-specific spectroscopy of peptide fragment ions in a low-temperature ion trap.

    PubMed

    Wassermann, Tobias N; Boyarkin, Oleg V; Paizs, Béla; Rizzo, Thomas R

    2012-06-01

    We have applied conformer-selective infrared-ultraviolet (IR-UV) double-resonance photofragment spectroscopy at low temperatures in an ion trap mass spectrometer for the spectroscopic characterization of peptide fragment ions. We investigate b- and a-type ions formed by collision-induced dissociation from protonated leucine-enkephalin. The vibrational analysis and assignment are supported by nitrogen-15 isotopic substitution of individual amino acid residues and assisted by density functional theory calculations. Under such conditions, b-type ions of different size are found to appear exclusively as linear oxazolone structures with protonation on the N-terminus, while a rearrangement reaction is confirmed for the a (4) ion in which the side chain of the C-terminal phenylalanine residue is transferred to the N-terminal side of the molecule. The vibrational spectra that we present here provide a particularly stringent test for theoretical approaches.

  17. Stereochemistry and conformation of skyllamycin, a non-ribosomally synthesized peptide from Streptomyces sp. Acta 2897.

    PubMed

    Schubert, Vivien; Di Meo, Florent; Saaidi, Pierre-Loïc; Bartoschek, Stefan; Fiedler, Hans-Peter; Trouillas, Patrick; Süssmuth, Roderich D

    2014-04-22

    Skyllamycin is a non-ribosomally synthesized cyclic depsipeptide from Streptomyces sp. Acta 2897 that inhibits PDGF-signaling. The peptide scaffold contains an N-terminal cinnamoyl moiety, a β-methylation of aspartic acid, three β-hydroxylated amino acids and one rarely occurring α-hydroxy glycine. With the exception of α-hydroxy glycine, the stereochemistry of the amino acids was assigned by comparison to synthetic reference amino acids applying chiral GC-MS and Marfey-HPLC analysis. The stereochemistry of α-hydroxy glycine, which is unstable under basic and acidic conditions, was determined by conformational analysis, employing a combination of data from NOESY-NMR spectroscopy, simulated annealing and free MD simulations. The simulation procedures were applied for both R- and S-configured α-hydroxy glycine of the skyllamycin structure and compared to the NOESY data. Both methods, simulated annealing and free MD simulations independently support S-configured α-hydroxy glycine thus enabling the assignment of all stereocenters in the structure of skyllamycin and devising the role of two-component flavin dependent monooxygenase (Sky39) as S-selective.

  18. Sequence-independent Control of Peptide Conformation in Liposomal Vaccines for Targeting Protein Misfolding Diseases*

    PubMed Central

    Hickman, David T.; López-Deber, María Pilar; Ndao, Dorin Mlaki; Silva, Alberto B.; Nand, Deepak; Pihlgren, Maria; Giriens, Valérie; Madani, Rime; St-Pierre, Annie; Karastaneva, Hristina; Nagel-Steger, Luitgard; Willbold, Dieter; Riesner, Detlev; Nicolau, Claude; Baldus, Marc; Pfeifer, Andrea; Muhs, Andreas

    2011-01-01

    Synthetic peptide immunogens that mimic the conformation of a target epitope of pathological relevance offer the possibility to precisely control the immune response specificity. Here, we performed conformational analyses using a panel of peptides in order to investigate the key parameters controlling their conformation upon integration into liposomal bilayers. These revealed that the peptide lipidation pattern, the lipid anchor chain length, and the liposome surface charge all significantly alter peptide conformation. Peptide aggregation could also be modulated post-liposome assembly by the addition of distinct small molecule β-sheet breakers. Immunization of both mice and monkeys with a model liposomal vaccine containing β-sheet aggregated lipopeptide (Palm1–15) induced polyclonal IgG antibodies that specifically recognized β-sheet multimers over monomer or non-pathological native protein. The rational design of liposome-bound peptide immunogens with defined conformation opens up the possibility to generate vaccines against a range of protein misfolding diseases, such as Alzheimer disease. PMID:21343310

  19. Crystal Structures of Polymorphic Prion Protein β1 Peptides Reveal Variable Steric Zipper Conformations.

    PubMed

    Yu, Lu; Lee, Seung-Joo; Yee, Vivien C

    2015-06-16

    The pathogenesis of prion diseases is associated with the conformational conversion of normal, predominantly α-helical prion protein (PrP(C)) into a pathogenic form that is enriched with β-sheets (PrP(Sc)). Several PrP(C) crystal structures have revealed β1-mediated intermolecular sheets, suggesting that the β1 strand may contribute to a possible initiation site for β-sheet-mediated PrP(Sc) propagation. This β1 strand contains the polymorphic residue 129 that influences disease susceptibility and phenotype. To investigate the effect of the residue 129 polymorphism on the conformation of amyloid-like continuous β-sheets formed by β1, crystal structures of β1 peptides containing each of the polymorphic residues were determined. To probe the conformational influence of the peptide construct design, four different lengths of β1 peptides were studied. From the 12 peptides studied, 11 yielded crystal structures ranging in resolution from 0.9 to 1.4 Å. This ensemble of β1 crystal structures reveals conformational differences that are influenced by both the nature of the polymorphic residue and the extent of the peptide construct, indicating that comprehensive studies in which peptide constructs vary are a more rigorous approach to surveying conformational possibilities.

  20. Conformation and Aggregation of LKα14 Peptide in Bulk Water and at the Air/Water Interface.

    PubMed

    Dalgicdir, Cahit; Sayar, Mehmet

    2015-12-10

    Historically, the protein folding problem has mainly been associated with understanding the relationship between amino acid sequence and structure. However, it is known that both the conformation of individual molecules and their aggregation strongly depend on the environmental conditions. Here, we study the aggregation behavior of the model peptide LKα14 (with amino acid sequence LKKLLKLLKKLLKL) in bulk water and at the air/water interface. We start by a quantitative analysis of the conformational space of a single LKα14 in bulk water. Next, in order to analyze the aggregation tendency of LKα14, by using the umbrella sampling technique we calculate the potential of mean force for pulling a single peptide from an n-molecule aggregate. In agreement with the experimental results, our calculations yield the optimal aggregate size as four. This equilibrium state is achieved by two opposing forces: Coulomb repulsion between the lysine side chains and the reduction of solvent accessible hydrophobic surface area upon aggregation. At the vacuum/water interface, however, even dimers of LKα14 become marginally stable, and any larger aggregate falls apart instantaneously. Our results indicate that even though the interface is highly influential in stabilizing the α-helix conformation for a single molecule, it significantly reduces the attraction between two LKα14 peptides, along with their aggregation tendency.

  1. The molecular basis of the temperature- and pH-induced conformational transitions in elastin-based peptides.

    PubMed

    Li, Bin; Daggett, Valerie

    2003-01-01

    Elastin undergoes an inverse temperature transition and collapses at high temperatures in both simulation and experiment. We investigated a pH-dependent modification of this transition by simulating a glutamic acid (Glu)-substituted elastin at varying pHs and temperatures. The Glu-substituted peptide collapsed at higher temperature than the unsubstituted elastin when Glu was charged. The charge effects could be reversed by neutralization of the Glu carboxyl groups at low pH, and in that case the peptide collapsed at a lower temperature. The collapse was accompanied by the formation of beta-turns and short distorted beta-sheets. Formation of contacts between hydrophobic side chains drives the collapse at high temperature, but interactions between water and polar groups (Glu and main chain) can attenuate this effect at high pH. The overall competition and balance of the polar and nonpolar groups determined the conformational states of the peptide. Water hydration contributed to the conformational transition, and the peptide and its hydration shell must be considered. Structurally, waters near polar residues mainly formed hydrogen bonds with the protein atoms, while waters around the hydrophobic side chains tended to be parallel to the peptide groups to maximize water-water interactions. Copyright 2002 Wiley Periodicals, Inc. Biopolymers 68: 121-129, 2003

  2. Tuning the conformation properties of a peptide by glycosylation and phosphorylation

    SciTech Connect

    Liang, F.-C.; Chen, Rita P.-Y.; Lin, C.-C.; Huang, K.-T.; Chan, S.I. . E-mail: SunneyChan@yahoo.com

    2006-04-07

    We have deployed the {alpha}-helical hairpin peptide ({alpha}-helix/turn/{alpha}-helix) and used it as a model system to explore how glycosylation and phosphorylation might affect the conformational properties of the peptide. The native conformations of the modified peptides in buffer solution have been compared with that of the wild-type peptide by nuclear magnetic resonance spectroscopy. Circular dichroism spectroscopy was used to probe the effects of an O-linked {beta}-GlcNAc and a phosphate group on the overall folding stability of the peptide. Finally, the rate of fibrillogenesis was used to infer the effects of these chemical modifications on the {alpha}-to-{beta} transition as well as the rate of nucleation of amyloidogenesis.

  3. Fluorescence study of conformational properties of melanotropins labeled with aminobenzoic acid.

    PubMed Central

    Ito, A S; Souza, E S; dos Reis Barbosa, S; Nakaie, C R

    2001-01-01

    The native hormone alpha-melanocyte-stimulating hormone (alpha-MSH) and its more potent analog [Nle(4),D-Phe(7)]alpha-MSH (NDP-alpha MSH), labeled at the amino terminal with the fluorescent aminobenzoic acid (Abz) isomers, were examined by fluorescence methods. We observed energy transfer between the tryptophan(9) residue acting as donor and Abz as acceptor, the transfer being more pronounced to the ortho-form of the acceptor. Within the hypothesis that different peptide conformations coexist in equilibrium during the fluorescence decay, we supposed that the intensity decay was modulated by an acceptor-donor distance distribution function f(r). From the time-resolved fluorescence experimental data, we recovered the distance distribution between Abz and Trp(9), using the CONTIN program, within the framework of the Förster resonance energy transfer model. The methodology proved to be useful to provide quantitative information about conformational dynamics of melanotropins and its dependency on the solvent. In aqueous medium, alpha-MSH has a broad Abz-Trp(9) distance distribution, reflecting the structural flexibility of the peptide. Three different distance populations could be identified in the labeled analog NDP-alpha MSH in water, indicating distinct conformational states for the synthetic peptide, compared with the native hormone. Measurements in trifluoroethanol resulted in the recovery of two Abz-Trp(9) distance populations, both for the native and the analog hormones, reflecting the decrease, induced by the solvent, of the conformational states available to the peptides. PMID:11463659

  4. Theoretical conformational analysis of the bovine adrenal medulla 12 residue peptide molecule

    NASA Astrophysics Data System (ADS)

    Akhmedov, N. A.; Tagiyev, Z. H.; Hasanov, E. M.; Akverdieva, G. A.

    2003-02-01

    The spatial structure and conformational properties of the bovine adrenal medulla 12 residue peptide Tyr1-Gly2-Gly3-Phe4-Met5-Arg6-Arg7-Val8-Gly9-Arg10-Pro11-Glu12 (BAM-12P) molecule were studied by theoretical conformational analysis. It is revealed that this molecule can exist in several stable states. The energy and geometrical parameters for the low-energy conformations are obtained. The conformationally rigid and labile segments of this molecule were revealed.

  5. Dependence of the AmII′p Proline Raman Band on Peptide Conformation

    PubMed Central

    Ahmed, Zeeshan; Myshakina, Nataliya S.; Asher, Sanford A.

    2009-01-01

    We utilized UV-resonance Raman (UVRR) measurements and density functional theory (DFT) calculations to relate that the AmII′p frequency to the ψ-angle. The AmII′p frequency shifts by ∼ 25 cm-1 as the ψ-angle is varied over allowed angles of the pro peptide bond. The AmII′p frequency does not show any significant dependence on the φ-dihedral angle. The conformation sensitivity of the AmII′p frequency derives from conformation-induced changes in the planarity of the Pro peptide bond; ψ angles changes push the amide nitrogen out of the peptide bond plane. We use this AmII′p frequency dependence on the ψ-angle to track temperature-induced conformation changes in a polyproline peptide. The temperature-induced 7 cm-1 downshift in the AmII′p frequency of the polyproline peptide results from a ∼45° rotation of the ψ dihedral angle from ψ = 145° (ideal PPII conformation) to ψ = 100° (collapsed PPII conformation). PMID:19627094

  6. Designed peptides with homochiral and heterochiral diproline templates as conformational constraints.

    PubMed

    Chatterjee, Bhaswati; Saha, Indranil; Raghothama, Srinivasarao; Aravinda, Subrayashastry; Rai, Rajkishor; Shamala, Narayanaswamy; Balaram, Padmanabhan

    2008-01-01

    Diproline segments have been advanced as templates for nucleation of folded structure in designed peptides. The conformational space available to homochiral and heterochiral diproline segments has been probed by crystallographic and NMR studies on model peptides containing L-Pro-L-Pro and D-Pro-L-Pro units. Four distinct classes of model peptides have been investigated: a) isolated D-Pro-L-Pro segments which form type II' beta-turn; b) D-Pro-L-Pro-L-Xxx sequences which form type II'-I (betaII'-I, consecutive beta-turns) turns; c) D-Pro-L-Pro-D-Xxx sequences; d) L-Pro-L-Pro-L-Xxx sequences. A total of 17 peptide crystal structures containing diproline segments are reported. Peptides of the type Piv-D-Pro-L-Pro-L-Xxx-NHMe are conformationally homogeneous, adopting consecutive beta-turn conformations. Peptides in the series Piv-D-Pro-L-Pro-D-Xxx-NHMe and Piv-L-Pro-L-Pro-L-Xxx-NHMe, display a heterogeneity of structures in crystals. A type VIa beta-turn conformation is characterized in Piv-L-Pro-L-Pro-L-Phe-OMe (18), while an example of a 5-->1 hydrogen bonded alpha-turn is observed in crystals of Piv-D-Pro-L-Pro-D-Ala-NHMe (11). An analysis of pyrrolidine conformations suggests a preferred proline puckering geometry is favored only in the case of heterochiral diproline segments. Solution NMR studies, reveal a strong conformational influence of the C-terminal Xxx residues on the structures of diproline segments. In L-Pro-L-Pro-L-Xxx sequences, the Xxx residues strongly determine the population of Pro-Pro cis conformers, with an overwhelming population of the trans form in L-Xxx=L-Ala (19).

  7. Multiple Simulated Annealing-Molecular Dynamics (MSA-MD) for Conformational Space Search of Peptide and Miniprotein.

    PubMed

    Hao, Ge-Fei; Xu, Wei-Fang; Yang, Sheng-Gang; Yang, Guang-Fu

    2015-10-23

    Protein and peptide structure predictions are of paramount importance for understanding their functions, as well as the interactions with other molecules. However, the use of molecular simulation techniques to directly predict the peptide structure from the primary amino acid sequence is always hindered by the rough topology of the conformational space and the limited simulation time scale. We developed here a new strategy, named Multiple Simulated Annealing-Molecular Dynamics (MSA-MD) to identify the native states of a peptide and miniprotein. A cluster of near native structures could be obtained by using the MSA-MD method, which turned out to be significantly more efficient in reaching the native structure compared to continuous MD and conventional SA-MD simulation.

  8. Peptide Conformations for a Microarray Surface-Tethered Epitope of the Tumor Suppressor p53

    SciTech Connect

    Feng, Jun; Wong, Ka-Yiu; Lynch, Gillian C.; Gao, Xiaolian; Pettitt, Bernard M.

    2007-12-13

    The research described in this product was performed in part in the Environmental Molecular Sciences Laboratory, a national scientific user facility sponsored by the Department of Energy's Office of Biological and Environmental Research and located at Pacific Northwest National Laboratory. Peptides or proteins near surfaces exhibit different structural properties from those present in a homogeneous solution, and these differences give rise to varied biological activity. Therefore, understanding the detailed molecular structure of these molecules tethered to a surface is important for interpreting the performance of the various microarrays based on the activities of the immobilized peptides or proteins. We performed molecular dynamics simulations of a pentapeptide, RHSVV, an epitope of the tumor suppressor protein p53, tethered via a spacer on a functionalized silica surface and free in solution, to study their structural and conformational differences. These calculations allowed analyses of the peptide-surface interactions, the sequence orientations, and the translational motions of the peptide on the surface to be performed. Conformational similarities are found among dominant structures of the tethered and free peptide. In the peptide microarray simulations, the peptide fluctuates between a parallel and tilted orientation driven in part by the hydrophobic interactions between the nonpolar peptide residues and the methyl-terminated silica surface. The perpendicular movement of the peptide relative to the surface is also restricted due to the hydrophobic nature of the microarray surface. With regard to structures available for recognition and binding, we find that similar conformations to those found in solution are available to the peptide tethered to the surface, but with a shifted equilibrium constant. Comparisons with experimental results show important implications of this for peptide microarray design and assays.

  9. Catalytic Activity of Peptide-Nanoparticle Conjugates Regulated by a Conformational Change.

    PubMed

    Mikolajczak, Dorian J; Heier, Jason L; Schade, Boris; Koksch, Beate

    2017-09-27

    Herein, we present the design and synthesis of a catalytically active peptide-nanoparticle conjugate whose activity is regulated by a defined conformational change in the self-assembled peptide monolayer. A catalytically active peptide, designed after the heterodimeric α-helical coiled-coil principle was immobilized onto gold nanoparticles, and kinetic studies were performed according to the Michaelis-Menten model. The formed peptide monolayer at the gold nanoparticle surface accelerated p-nitrophenylacetate (pNPA) hydrolysis by 1 order of magnitude compared to the soluble peptide while exhibiting no defined secondary structure as determined by infrared (IR) and circular dichroism (CD) spectroscopy. Addition of the complementary peptide-induced coiled-coil formation while significantly hindering the pNPA hydrolysis catalyzed by the peptide-nanoparticle conjugate. The heptad repeat sequence of a coiled-coil opens up the opportunity for regulation of conformation and thus catalytic activity of peptide-nanoparticle conjugates upon interaction with a complementary coiled-coil sequence. Strategies of regulation of catalytic activity by interaction with a complementary cofactor/ligand are well-established in nature and are introduced here into rationally designed peptide-nanoparticle conjugates.

  10. Retention of Conformational Entropy upon Calmodulin Binding to Target Peptides is Driven by Transient Salt Bridges

    SciTech Connect

    Smith, Dayle MA; Straatsma, TP; Squier, Thomas C.

    2012-10-03

    Calmodulin (CaM) is a highly flexible calcium-binding protein that mediates signal transduction through an ability to differentially bind to highly variable binding sequences in target proteins. To identify how binding affects CaM motions, and its relationship to conformational entropy and target peptide sequence, we have employed fully atomistic, explicit solvent molecular dynamics simulations of unbound CaM and CaM bound to five different target peptides. The calculated CaM conformational binding entropies correlate with experimentally derived conformational entropies with a correlation coefficient R2 of 0.95. Selected side-chain interactions with target peptides restrain interhelical loop motions, acting to tune the conformational entropy of the bound complex via widely distributed CaM motions. In the complex with the most conformational entropy retention (CaM in complex with the neuronal nitric oxide synthase binding sequence), Lys-148 at the C-terminus of CaM forms transient salt bridges alternating between Glu side chains in the N-domain, the central linker, and the binding target. Additional analyses of CaM structures, fluctuations, and CaM-target interactions illuminate the interplay between electrostatic, side chain, and backbone properties in the ability of CaM to recognize and discriminate against targets by tuning its conformational entropy, and suggest a need to consider conformational dynamics in optimizing binding affinities.

  11. Molecular conformation of a peptide fragment of transthyretin in an amyloid fibril

    PubMed Central

    Jaroniec, Christopher P.; MacPhee, Cait E.; Astrof, Nathan S.; Dobson, Christopher M.; Griffin, Robert G.

    2002-01-01

    The molecular conformation of peptide fragment 105–115 of transthyretin, TTR(105–115), previously shown to form amyloid fibrils in vitro, has been determined by magic-angle spinning solid-state NMR spectroscopy. 13C and 15N linewidth measurements indicate that TTR(105–115) forms a highly ordered structure with each amino acid in a unique environment. 2D 13C-13C and 15N-13C-13C chemical shift correlation experiments, performed on three fibril samples uniformly 13C,15N-labeled in consecutive stretches of 4 aa, allowed the complete sequence-specific backbone and side-chain 13C and 15N resonance assignments to be obtained for residues 105–114. Analysis of the 15N, 13CO, 13Cα, and 13Cβ chemical shifts allowed quantitative predictions to be made for the backbone torsion angles φ and ψ. Furthermore, four backbone 13C–15N distances were determined in two selectively 13C,15N-labeled fibril samples by using rotational-echo double-resonance NMR. The results show that TTR(105–115) adopts an extended β-strand conformation that is similar to that found in the native protein except for substantial differences in the vicinity of the proline residue. PMID:12481032

  12. Mapping the conformational free energy of aspartic acid in the gas phase and in aqueous solution.

    PubMed

    Comitani, Federico; Rossi, Kevin; Ceriotti, Michele; Sanz, M Eugenia; Molteni, Carla

    2017-04-14

    The conformational free energy landscape of aspartic acid, a proteogenic amino acid involved in a wide variety of biological functions, was investigated as an example of the complexity that multiple rotatable bonds produce even in relatively simple molecules. To efficiently explore such a landscape, this molecule was studied in the neutral and zwitterionic forms, in the gas phase and in water solution, by means of molecular dynamics and the enhanced sampling method metadynamics with classical force-fields. Multi-dimensional free energy landscapes were reduced to bi-dimensional maps through the non-linear dimensionality reduction algorithm sketch-map to identify the energetically stable conformers and their interconnection paths. Quantum chemical calculations were then performed on the minimum free energy structures. Our procedure returned the low energy conformations observed experimentally in the gas phase with rotational spectroscopy [M. E. Sanz et al., Phys. Chem. Chem. Phys. 12, 3573 (2010)]. Moreover, it provided information on higher energy conformers not accessible to experiments and on the conformers in water. The comparison between different force-fields and quantum chemical data highlighted the importance of the underlying potential energy surface to accurately capture energy rankings. The combination of force-field based metadynamics, sketch-map analysis, and quantum chemical calculations was able to produce an exhaustive conformational exploration in a range of significant free energies that complements the experimental data. Similar protocols can be applied to larger peptides with complex conformational landscapes and would greatly benefit from the next generation of accurate force-fields.

  13. Mapping the conformational free energy of aspartic acid in the gas phase and in aqueous solution

    NASA Astrophysics Data System (ADS)

    Comitani, Federico; Rossi, Kevin; Ceriotti, Michele; Sanz, M. Eugenia; Molteni, Carla

    2017-04-01

    The conformational free energy landscape of aspartic acid, a proteogenic amino acid involved in a wide variety of biological functions, was investigated as an example of the complexity that multiple rotatable bonds produce even in relatively simple molecules. To efficiently explore such a landscape, this molecule was studied in the neutral and zwitterionic forms, in the gas phase and in water solution, by means of molecular dynamics and the enhanced sampling method metadynamics with classical force-fields. Multi-dimensional free energy landscapes were reduced to bi-dimensional maps through the non-linear dimensionality reduction algorithm sketch-map to identify the energetically stable conformers and their interconnection paths. Quantum chemical calculations were then performed on the minimum free energy structures. Our procedure returned the low energy conformations observed experimentally in the gas phase with rotational spectroscopy [M. E. Sanz et al., Phys. Chem. Chem. Phys. 12, 3573 (2010)]. Moreover, it provided information on higher energy conformers not accessible to experiments and on the conformers in water. The comparison between different force-fields and quantum chemical data highlighted the importance of the underlying potential energy surface to accurately capture energy rankings. The combination of force-field based metadynamics, sketch-map analysis, and quantum chemical calculations was able to produce an exhaustive conformational exploration in a range of significant free energies that complements the experimental data. Similar protocols can be applied to larger peptides with complex conformational landscapes and would greatly benefit from the next generation of accurate force-fields.

  14. Peptide binding identifies an ERalpha conformation that generates selective activity in multiple in vitro assays.

    PubMed

    Larson, Christopher J; Osburn, Deborah L; Schmitz, Katherine; Giampa, Leslie; Mong, Shau-Ming; Marschke, Keith; Seidel, H Martin; Rosen, Jonathan; Negro-Vilar, Andrés

    2005-09-01

    Drugs such as tamoxifen, which act at the estrogen receptor (ER), have very different in vitro and in vivo effects from those of the native hormone. Previous research has established that different ligands induce distinct conformational changes in the ER, thus affecting the interactions of the receptor with cell-specific co-activating or co-repressing proteins (cofactors) and estrogen response elements (EREs), thus potentially driving differing biological effects. Affinity-selected peptides have been used to probe the conformational changes that occur within the ER upon binding various ligands. In this study, the authors characterize the ability of several peptides to be recruited to liganded ER under cellular conditions. Approximating ER conformation via recruitment of this peptide to the ER is concluded to be a better predictor of the agonist nature of an ER ligand under these different cellular contexts than is a canonical cotransfection transactivation assay.

  15. Understanding the connection between conformational changes of peptides and equilibrium thermal fluctuations.

    PubMed

    Soler, Miguel A; Zúñiga, José; Requena, Alberto; Bastida, Adolfo

    2017-02-01

    Despite the increasing evidence that conformational transitions in peptides and proteins are driven by specific vibrational energy pathways along the molecule, the current experimental techniques of analysis do as yet not allow to study these biophysical processes in terms of anisotropic energy flows. Computational methods offer a complementary approach to obtain a more detailed understanding of the vibrational and conformational dynamics of these systems. Accordingly, in this work we investigate jointly the vibrational energy distribution and the conformational dynamics of trialanine peptide in water solution at room temperature by applying the Instantaneous Normal Mode analysis to the results derived from equilibrium molecular dynamics simulations. It is shown that conformational changes in trialanine are triggered by the vibrational energy accumulated in the low-frequency modes of the molecule, and that excitation is caused exclusively by thermal fluctuations of the solute-solvent system, thus excluding the possibility of an intramolecular vibrational energy redistribution process.

  16. Single-Molecule Protein Folding: A Study of the Surface-Mediated Conformational Dynamics of a Model Amphipathic Peptide

    NASA Astrophysics Data System (ADS)

    Cunningham, Joy; English, Douglas

    2004-03-01

    Most surface-active polypeptides, composed of 10-50 amino acids, are devoid of well-defined tertiary structure. The conformation of these proteins is greatly dependent upon their environment and may assume totally different characteristics in an aqueous environment, in a detergent micelle, or in an organic solvent. Most antimicrobial peptides are helix-forming and are activated upon interaction with a membrane-mimicking environment. We are seeking to physically characterize the mechanism of membrane-peptide interaction through studying a simple model peptide, MT-1. MT-1 was designed as a nonhomologous analogue of melittin, the principle component in bee venom. We are using single molecule spectroscopy to examine the induction of secondary structure upon interaction of MT-1 with various membrane-mimicking interfaces. Specifically, we monitor coil-to-helix transition through single molecule fluorescence resonance energy transfer (sm-FRET) to determine conformational distributions of folded and unfolded peptides at an interface. Studies with MT-1 will focus upon the biologically relevant issues of orientation, aggregation, and folding at surfaces using both ensemble and single molecule experiments.

  17. Reversible photocontrol of peptide conformation with a rhodopsin-like photoswitch.

    PubMed

    Blanco-Lomas, Marina; Samanta, Subhas; Campos, Pedro J; Woolley, G Andrew; Sampedro, Diego

    2012-04-25

    Reversible photocontrol of biomolecules requires chromophores that can efficiently undergo large conformational changes upon exposure to wavelengths of light that are compatible with living systems. We designed a benzylidene-pyrroline chromophore that mimics the Schiff base of rhodopsin and can be used to introduce light-switchable intramolecular cross-links in peptides and proteins. This new class of photoswitch undergoes an ~10 Å change in end-to-end distance upon isomerization and can be used to control the conformation of a target peptide efficiently and reversibly using, alternately, violet (400 nm) and blue (446 nm) light.

  18. Single-Conformation IR and UV Spectroscopy of a Prototypical Heterogeneous α/β-PEPTIDE: is it a Mixed-Helix Former?

    NASA Astrophysics Data System (ADS)

    Blodgett, Karl N.; Walsh, Patrick S.; Zwier, Timothy S.

    2016-06-01

    Synthetic foldamers are non-natural polymers designed to fold into unique secondary structures that either mimic nature's preferred secondary structures, or expand their possibilities. Among the most studied synthetic foldamers are β-peptides, which lengthen the distance between amide groups from the single substituted carbon spacer in α-peptides by one additional carbon. We present data on a mixed α/β tri-peptide in which a single β-residue with a conformationally constrained cis-2-aminocyclohexanecarboxylic acid (cis-ACHC) substitution is inserted in an α-peptide backbone to form Ac-Ala-β-ACHC-Ala-NHBn. This αβα structure is known in longer sequences to prefer formation of a 9/11 mixed helix. Under isolated, jet cooled conditions, four unique conformers were observed in the expansion. The dominant conformer is configured in a tetramer cycle with every amide carbonyl and amine group involved in hydrogen bonding, giving rise to a tightly folded C12/C7/C8/C7 structure reminiscent of a β-turn. This talk will describe the conformation specific IR and UV spectroscopy methods used to study this mixed peptide, as well as its experimentally observed conformational preferences.

  19. Rational optimization of conformational effects induced by hydrocarbon staples in peptides and their binding interfaces.

    PubMed

    Lama, Dilraj; Quah, Soo T; Verma, Chandra S; Lakshminarayanan, Rajamani; Beuerman, Roger W; Lane, David P; Brown, Christopher J

    2013-12-13

    eIF4E is frequently over-expressed in different cancers and causes increased translation of oncogenic proteins via deregulated cap-dependent translation. Inhibitors of the eIF4E:eIF4G interactions represent an approach that would normalize cap-dependent translation. Stapled peptides represent an emerging class of therapeutics that can target protein: protein interactions. We present here molecular dynamics simulations for a set of rationally designed stapled peptides in solution and in complex with eIF4E, supported with biophysical and crystallographic data. Clustering of the simulated structures revealed the favoured conformational states of the stapled peptides in their bound or free forms in solution. Identifying these populations has allowed us to design peptides with improved affinities by introducing mutations into the peptide sequence to alter their conformational distributions. These studies emphasise the effects that engineered mutations have on the conformations of free and bound peptides, and illustrate that both states must be considered in efforts to attain high affinity binding.

  20. Rational Optimization of Conformational Effects Induced By Hydrocarbon Staples in Peptides and their Binding Interfaces

    NASA Astrophysics Data System (ADS)

    Lama, Dilraj; Quah, Soo T.; Verma, Chandra S.; Lakshminarayanan, Rajamani; Beuerman, Roger W.; Lane, David P.; Brown, Christopher J.

    2013-12-01

    eIF4E is frequently over-expressed in different cancers and causes increased translation of oncogenic proteins via deregulated cap-dependent translation. Inhibitors of the eIF4E:eIF4G interactions represent an approach that would normalize cap-dependent translation. Stapled peptides represent an emerging class of therapeutics that can target protein: protein interactions. We present here molecular dynamics simulations for a set of rationally designed stapled peptides in solution and in complex with eIF4E, supported with biophysical and crystallographic data. Clustering of the simulated structures revealed the favoured conformational states of the stapled peptides in their bound or free forms in solution. Identifying these populations has allowed us to design peptides with improved affinities by introducing mutations into the peptide sequence to alter their conformational distributions. These studies emphasise the effects that engineered mutations have on the conformations of free and bound peptides, and illustrate that both states must be considered in efforts to attain high affinity binding.

  1. Effect of urea on peptide conformation in water: molecular dynamics and experimental characterization.

    PubMed

    Caballero-Herrera, Ana; Nordstrand, Kerstin; Berndt, Kurt D; Nilsson, Lennart

    2005-08-01

    Molecular dynamics simulations of a ribonuclease A C-peptide analog and a sequence variant were performed in water at 277 and 300 K and in 8 M urea to clarify the molecular denaturation mechanism induced by urea and the early events in protein unfolding. Spectroscopic characterization of the peptides showed that the C-peptide analog had a high alpha-helical content, which was not the case for the variant. In the simulations, interdependent side-chain interactions were responsible for the high stability of the alpha-helical C-peptide analog in the different solvents. The other peptide displayed alpha-helical unwinding that propagated cooperatively toward the N-terminal. The conformations sampled by the peptides depended on their sequence and on the solvent. The ability of water molecules to form hydrogen bonds to the peptide as well as the hydrogen bond lifetimes increased in the presence of urea, whereas water mobility was reduced near the peptide. Urea accumulated in excess around the peptide, to which it formed long-lived hydrogen bonds. The unfolding mechanisms induced by thermal denaturation and by urea are of a different nature, with urea-aqueous solutions providing a better peptide solvation than pure water. Our results suggest that the effect of urea on the chemical denaturation process involves both the direct and indirect mechanisms.

  2. Interplay between Intrinsic Conformational Propensities and Intermolecular Interactions in the Self-Assembly of Short Surfactant-like Peptides Composed of Leucine/Isoleucine.

    PubMed

    Zhou, Peng; Deng, Li; Wang, Yanting; Lu, Jian R; Xu, Hai

    2016-05-10

    To study how the conformational propensities of individual amino acid residues, primary structures (i.e., adjacent residues and molecular lengths), and intermolecular interactions of peptides affect their self-assembly properties, we report the use of replica exchange molecular dynamics (REMD) to investigate the monomers, dimers, and trimers of a series of short surfactant-like peptides (I3K, L3K, L4K, and L5K). For four-residue peptides X3K (I3K and L3K), the results show that their different aggregation behaviors arise from the different intrinsic conformational propensities of isoleucine and leucine. For LmK peptides (L3K, L4K, and L5K), the molecular length is found to dictate their aggregation via primarily modulating intermolecular interactions. Increasing the number of hydrophobic amino acid residues of LmK peptides enhances their intermolecular H-bonding and promotes the formation of β-strands in dimer and trimer aggregates, overwhelming the intrinsic preference of Leu for helical structures. Thus, the interplay between the conformational propensities of individual amino acid residues for secondary structures and molecular interactions determines the self-assembly properties of the peptides, and the competition between intramolecular and intermolecular H-bonding interactions determines the probability of β-sheet alignment of peptide molecules. These results are validated by comparing simulated and experimental CD spectra of the peptides. This study will aid the design of short peptide amphiphiles and improve the mechanistic understanding of their self-assembly behavior.

  3. Signal enhancement for gene detection based on a redox reaction of [Fe(CN)(6)](4-) mediated by ferrocene at the terminal of a peptide nucleic acid as a probe with hybridization-amenable conformational flexibility.

    PubMed

    Aoki, Hiroshi; Tao, Hiroaki

    2008-07-01

    Electrochemically enhanced DNA detection was demonstrated by utilizing the couple of a synthesized ferrocene-terminated peptide nucleic acid (PNA) with a cysteine anchor and a sacrificial electron donor [Fe(CN)(6)](4-). DNA detection sensors were prepared by modifying a gold electrode surface with a mixed monolayer of the probe PNA and 11-hydroxy-1-undecanethiol (11-HUT), protecting [Fe(CN)(6)](4-) from any unexpected redox reaction. Before hybridization, the terminal ferrocene moiety of the probe was subject to a redox reaction due to the flexible probe structure and, in the presence of [Fe(CN)(6)](4-), the observed current was amplified based on regeneration of the ferrocene moiety. Hybridization decreased the redox current of the ferrocene. This occurred because hybridization rigidified the probe structure: the ferrocene moiety was then removed from the electrode surface, and the redox reaction of [Fe(CN)(6)](4-) was again prevented. The change in the anodic current before and after hybridization was enhanced 1.75-fold by using the electron donor [Fe(CN)(6)](4-). Sequence-specific detection of the complementary target DNA was also demonstrated.

  4. Effects of Trimethylamine-N -oxide on the Conformation of Peptides and its Implications for Proteins

    NASA Astrophysics Data System (ADS)

    Su, Zhaoqian; Mahmoudinobar, Farbod; Dias, Cristiano L.

    2017-09-01

    To provide insights into the stabilizing mechanisms of trimethylamine-N -oxide (TMAO) on protein structures, we perform all-atom molecular dynamics simulations of peptides and the Trp-cage miniprotein. The effects of TMAO on the backbone and charged residues of peptides are found to stabilize compact conformations, whereas effects of TMAO on nonpolar residues lead to peptide swelling. This suggests competing mechanisms of TMAO on proteins, which accounts for hydrophobic swelling, backbone collapse, and stabilization of charge-charge interactions. These mechanisms are observed in Trp cage.

  5. Conformational Entropy of Intrinsically Disordered Proteins from Amino Acid Triads

    PubMed Central

    Baruah, Anupaul; Rani, Pooja; Biswas, Parbati

    2015-01-01

    This work quantitatively characterizes intrinsic disorder in proteins in terms of sequence composition and backbone conformational entropy. Analysis of the normalized relative composition of the amino acid triads highlights a distinct boundary between globular and disordered proteins. The conformational entropy is calculated from the dihedral angles of the middle amino acid in the amino acid triad for the conformational ensemble of the globular, partially and completely disordered proteins relative to the non-redundant database. Both Monte Carlo (MC) and Molecular Dynamics (MD) simulations are used to characterize the conformational ensemble of the representative proteins of each group. The results show that the globular proteins span approximately half of the allowed conformational states in the Ramachandran space, while the amino acid triads in disordered proteins sample the entire range of the allowed dihedral angle space following Flory’s isolated-pair hypothesis. Therefore, only the sequence information in terms of the relative amino acid triad composition may be sufficient to predict protein disorder and the backbone conformational entropy, even in the absence of well-defined structure. The predicted entropies are found to agree with those calculated using mutual information expansion and the histogram method. PMID:26138206

  6. Conformational equilibria of terminally blocked single amino acids at the water-hexane interface. A molecular dynamics study.

    PubMed

    Chipot, C; Pohorille, A

    1998-01-01

    The conformational equilibria of the acetyl and methyl amide terminally blocked L-alanine, L-leucine and L-glutamine amino acids are examined in vacuum, in bulk water, and at the water-hexane interface, using multi-nanosecond molecular dynamics simulations. The two-dimensional probability distribution functions of finding the peptides at different dihedral angles of the backbone, phi and psi, are calculated, and free energy differences between different conformational states are determined. All three peptides are interfacially active, i.e. tend to accumulate at the interface even though they are not amphiphilic. Conformational states stable in both gas phase and water are also stable in the interfacial environment. Their populations, however, cannot be simply predicted from the knowledge of conformational equilibria in the bulk phases, indicating that the interface exerts a unique effect on the peptides. Conformational preferences in the interfacial environment arise from the interplay between electrostatic and hydrophobic effects. As in an aqueous solution, electrostatic solute-solvent interactions lead to the stabilization of more polar peptide conformations. The hydrophobic effect is manifested at the interface by a tendency to segregate polar and nonpolar moieties of the solute into the aqueous and the hexane phases, respectively. For the terminally blocked glutamine, this favors conformations for which such a segregation is compatible with the formation of strong, backbone-side chain intramolecular hydrogen bonds on the hexane side of the interface. The influence of the hydrophobic effect can be also noted in the orientational preferences of the peptides at the interface. The terminally blocked leucine is oriented such that its nonpolar side chain is buried in hexane, whereas the polar side chain of glutamine is immersed in water. The free energies of rotating the peptides along the axis parallel to the interface by more than 90 degrees are substantial. This

  7. Structural and conformational similarity between synthetic peptides of curaremimetic neurotoxins and rabies virus glycoprotein.

    PubMed

    Donnelly-Roberts, D L; Lentz, T L

    1991-09-01

    Antibodies were raised in rabbits against synthetic peptides corresponding to loop 2, the 'toxic' loop reacting with the acetylcholine-binding site on the nicotinic acetylcholine receptor, of curaremimetic neurotoxins and the structurally similar segment of the rabies virus glycoprotein. Some of the antibodies cross-reacted with the corresponding peptides confirming the structural similarity between the neurotoxin and glycoprotein peptides. A polyclonal antibody raised against a 29 residue glycoprotein peptide (175-203) in the presence of 0.1% sodium dodecyl sulfate reacted with native alpha-bungarotoxin and rabies virus. Circular dichroism spectroscopy of the 29 residue glycoprotein peptide and a 20 residue king cobra loop 2 peptide (25-44) revealed these peptides to be conformationally similar and composed predominantly of beta sheet structure. These results show the rabies glycoprotein segment is structurally and conformationally similar to neurotoxin loop 2. This similarity may confer on the glycoprotein the capability of interacting with the neurotoxin-binding site on the acetylcholine receptor.

  8. Structure and dynamics of water in crowded environments slows down peptide conformational changes

    SciTech Connect

    Lu, Cheng; Prada-Gracia, Diego; Rao, Francesco

    2014-07-28

    The concentration of macromolecules inside the cell is high with respect to conventional in vitro experiments or simulations. In an effort to characterize the effects of crowding on the thermodynamics and kinetics of disordered peptides, molecular dynamics simulations were run at different concentrations by varying the number of identical weakly interacting peptides inside the simulation box. We found that the presence of crowding does not influence very much the overall thermodynamics. On the other hand, peptide conformational dynamics was found to be strongly affected, resulting in a dramatic slowing down at larger concentrations. The observation of long lived water bridges between peptides at higher concentrations points to a nontrivial role of the solvent in the altered peptide kinetics. Our results reinforce the idea for an active role of water in molecular crowding, an effect that is expected to be relevant for problems influenced by large solvent exposure areas like in intrinsically disordered proteins.

  9. NCAD, a database integrating the intrinsic conformational preferences of non-coded amino acids

    PubMed Central

    Revilla-López, Guillem; Torras, Juan; Curcó, David; Casanovas, Jordi; Calaza, M. Isabel; Zanuy, David; Jiménez, Ana I.; Cativiela, Carlos; Nussinov, Ruth; Grodzinski, Piotr; Alemán, Carlos

    2010-01-01

    Peptides and proteins find an ever-increasing number of applications in the biomedical and materials engineering fields. The use of non-proteinogenic amino acids endowed with diverse physicochemical and structural features opens the possibility to design proteins and peptides with novel properties and functions. Moreover, non-proteinogenic residues are particularly useful to control the three-dimensional arrangement of peptidic chains, which is a crucial issue for most applications. However, information regarding such amino acids –also called non-coded, non-canonical or non-standard– is usually scattered among publications specialized in quite diverse fields as well as in patents. Making all these data useful to the scientific community requires new tools and a framework for their assembly and coherent organization. We have successfully compiled, organized and built a database (NCAD, Non-Coded Amino acids Database) containing information about the intrinsic conformational preferences of non-proteinogenic residues determined by quantum mechanical calculations, as well as bibliographic information about their synthesis, physical and spectroscopic characterization, conformational propensities established experimentally, and applications. The architecture of the database is presented in this work together with the first family of non-coded residues included, namely, α-tetrasubstituted α-amino acids. Furthermore, the NCAD usefulness is demonstrated through a test-case application example. PMID:20455555

  10. Pyrazole amino acids: hydrogen bonding directed conformations of 3-amino-1H-pyrazole-5-carboxylic acid residue.

    PubMed

    Kusakiewicz-Dawid, Anna; Porada, Monika; Ochędzan-Siodłak, Wioletta; Broda, Małgorzata A; Bujak, Maciej; Siodłak, Dawid

    2017-09-01

    A series of model compounds containing 3-amino-1H-pyrazole-5-carboxylic acid residue with N-terminal amide/urethane and C-terminal amide/hydrazide/ester groups were investigated by using NMR, Fourier transform infrared, and single-crystal X-ray diffraction methods, additionally supported by theoretical calculations. The studies demonstrate that the most preferred is the extended conformation with torsion angles ϕ and ψ close to ±180°. The studied 1H-pyrazole with N-terminal amide/urethane and C-terminal amide/hydrazide groups solely adopts this energetically favored conformation confirming rigidity of that structural motif. However, when the C-terminal ester group is present, the second conformation with torsion angles ϕ and ψ close to ±180° and 0°, respectively, is accessible. The conformational equilibrium is observed in NMR and Fourier transform infrared studies in solution in polar environment as well as in the crystal structures of other related compounds. The observed conformational preferences are clearly related to the presence of intramolecular interactions formed within the studied residue. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.

  11. Conformational preferences of 3,4-dihydroxyphenylacetic acid (DOPAC)

    NASA Astrophysics Data System (ADS)

    Lopes Jesus, A. J.; Jarmelo, S.; Fausto, R.; Reva, I.

    2015-04-01

    The conformational space of 3,4-dihydroxyphenylacetic acid (DOPAC), an important dopamine metabolite, has been investigated by quantum chemical methods (B3LYP and MP2, with the 6-311++G(d,p) basis set) and matrix-isolation infrared spectroscopy. Detailed analysis of the calculated potential energy surfaces of the molecule led to identification of thirteen unique conformers, all of them showing the acetic acid side chain out of the aromatic ring plane by 60-95°. According to the calculated Gibbs energies, the five lowest energy conformers make up 99.7% of the conformational mixture at 298.15 K, exhibiting individual populations falling between 16% and 24%. The main conformational trends of this molecule were interpreted on the grounds of a thorough analysis of the structural parameters and by the application of the Natural Bond Orbital theory. The role of the intramolecular interactions on the relative stability and structure of the conformers was also investigated. The infrared spectrum of DOPAC was registered after isolation of its monomers in argon and xenon matrices. Only one of DOPAC forms populated in the gas phase could be trapped in both matrix gases. This result is in agreement with the predicted low energy barriers for conformational isomerization and is also supported by annealing experiments. The spectra of matrix-isolated model compounds, phenylacetic acid and catechol, were studied under the same experimental conditions. These data were used as references and assisted in the interpretation of the results obtained for DOPAC.

  12. The intrinsic conformational features of amino acids from a protein coil library and their applications in force field development.

    PubMed

    Jiang, Fan; Han, Wei; Wu, Yun-Dong

    2013-03-14

    The local conformational (φ, ψ, χ) preferences of amino acid residues remain an active research area, which are important for the development of protein force fields. In this perspective article, we first summarize spectroscopic studies of alanine-based short peptides in aqueous solution. While most studies indicate a preference for the P(II) conformation in the unfolded state over α and β conformations, significant variations are also observed. A statistical analysis from various coil libraries of high-resolution protein structures is then summarized, which gives a more coherent view of the local conformational features. The φ, ψ, χ distributions of the 20 amino acids have been obtained from a protein coil library, considering both backbone and side-chain conformational preferences. The intrinsic side-chain χ(1) rotamer preference and χ(1)-dependent Ramachandran plot can be generally understood by combining the interaction of the side-chain Cγ/Oγ atom with two neighboring backbone peptide groups. Current all-atom force fields such as AMBER ff99sb-ILDN, ff03 and OPLS-AA/L do not reproduce these distributions well. A method has been developed by combining the φ, ψ plot of alanine with the influence of side-chain χ(1) rotamers to derive the local conformational features of various amino acids. It has been further applied to improve the OPLS-AA force field. The modified force field (OPLS-AA/C) reproduces experimental (3)J coupling constants for various short peptides quite well. It also better reproduces the temperature-dependence of the helix-coil transition for alanine-based peptides. The new force field can fold a series of peptides and proteins with various secondary structures to their experimental structures. MD simulations of several globular proteins using the improved force field give significantly less deviation (RMSD) to experimental structures. The results indicate that the local conformational features from coil libraries are valuable for

  13. Single-conformation and diastereomer specific ultraviolet and infrared spectroscopy of model synthetic foldamers: alpha/beta-peptides.

    PubMed

    James, William H; Baquero, Esteban E; Shubert, V Alvin; Choi, Soo Hyuk; Gellman, Samuel H; Zwier, Timothy S

    2009-05-13

    Resonant two-photon ionization (R2PI), UV hole-burning (UVHB), and resonant ion-dip infrared (RIDIR) spectroscopies have been used to record single-conformation infrared and ultraviolet spectra of three model synthetic foldamers with heterogeneous backbones, alpha/beta-peptides Ac-beta(3)-hAla-L-Phe-NHMe (betaalphaL), Ac-beta(3)-hAla-D-Phe-NHMe (betaalphaD), and Ac-L-Phe-beta(3)-hAla-NHMe (alphabetaL), isolated and cooled in a supersonic expansion. BetaalphaL and betaalphaD are diastereomers, differing only in the configuration of the alpha-amino acid residue; betaalphaL and alphabetaL contain the same residues, but differ in residue order. In all three alpha/beta-peptides the beta(3)-residue has S absolute configuration. UVHB spectroscopy is used to determine that there are six conformers of each molecule and to locate and characterize their S(0)-S(1) transitions in the origin region. RIDIR spectra in the amide NH stretch region reflect the number and strength of intramolecular H-bonds present. Comparison of the RIDIR spectra with scaled, harmonic vibrational frequencies and infrared intensities leads to definite assignments for the conformational families involved. C8/C7(eq) double-ring structures are responsible for three conformers of betaalphaL and four of betaalphaD, including those with the most intense transitions in the R2PI spectra. This preference for C8/C7(eq) double rings appears to be dictated by the C7(eq) ring of the alpha-peptide subunit. Three of the conformers of betaalphaL and betaalphaD form diastereomeric pairs (A/A', C/C', and G/G') that have nearly identical S(0)-S(1) origin positions in the UV and belong to the same conformational family, indicating no significant change associated with the change in chirality of the alpha-peptide subunit. However, betaalphaL favors formation of a C6/C5 conformer over C11, while the reverse preference holds in betaalphaD. Calculations indicate that the selective stabilization of the lowest-energy C11(g

  14. The conformational properties of α,β-dehydroamino acids with a C-terminal ester group.

    PubMed

    Siodłak, Dawid; Grondys, Justyna; Broda, Małgorzata A

    2011-10-01

    α,β-Dehydroamino acid esters occur in nature. To investigate their conformational properties, a systematic theoretical analysis was performed on the model molecules Ac-ΔXaa-OMe [ΔXaa = ΔAla, (E)-ΔAbu, (Z)-ΔAbu, ΔVal] at the B3LYP/6-311+ + G(d,p) level in the gas phase as well as in chloroform and water solutions with the self-consistent reaction field-polarisable continuum model method. The Fourier transform IR spectra in CCl(4) and CHCl(3) have been analysed as well as the analogous solid state conformations drawn from The Cambridge Structural Database. The ΔAla residue has a considerable tendency to adopt planar conformations C5 (ϕ, ψ ≈ - 180°, 180°) and β2 (ϕ, ψ ≈ - 180°, 0°), regardless of the environment. The ΔVal residue prefers the conformation β2 (ϕ, ψ ≈ - 120°, 0°) in a low polar environment, but the conformations α (ϕ, ψ ≈ - 55°, 35°) and β (ϕ, ψ ≈ - 55°, 145°) when the polarity increases. The ΔAbu residues reveal intermediate properties, but their conformational dispositions depend on configuration of the side chain of residue: (E)-ΔAbu is similar to ΔAla, whereas (Z)-ΔAbu to ΔVal. Results indicate that the low-energy conformation β2 is the characteristic feature of dehydroamino acid esters. The studied molecules constitute conformational patterns for dehydroamino acid esters with various side chain substituents in either or both Z and E positions. Copyright © 2011 European Peptide Society and John Wiley & Sons, Ltd.

  15. Characterizing the Conformational Landscape of Flavivirus Fusion Peptides via Simulation and Experiment.

    PubMed

    Marzinek, Jan K; Lakshminarayanan, Rajamani; Goh, Eunice; Huber, Roland G; Panzade, Sadhana; Verma, Chandra; Bond, Peter J

    2016-01-20

    Conformational changes in the envelope proteins of flaviviruses help to expose the highly conserved fusion peptide (FP), a region which is critical to membrane fusion and host cell infection, and which represents a significant target for antiviral drugs and antibodies. In principle, extended timescale atomic-resolution simulations may be used to characterize the dynamics of such peptides. However, the resultant accuracy is critically dependent upon both the underlying force field and sufficient conformational sampling. In the present study, we report a comprehensive comparison of three simulation methods and four force fields comprising a total of more than 40 μs of sampling. Additionally, we describe the conformational landscape of the FP fold across all flavivirus family members. All investigated methods sampled conformations close to available X-ray structures, but exhibited differently populated ensembles. The best force field / sampling combination was sufficiently accurate to predict that the solvated peptide fold is less ordered than in the crystallographic state, which was subsequently confirmed via circular dichroism and spectrofluorometric measurements. Finally, the conformational landscape of a mutant incapable of membrane fusion was significantly shallower than wild-type variants, suggesting that dynamics should be considered when therapeutically targeting FP epitopes.

  16. Characterizing the Conformational Landscape of Flavivirus Fusion Peptides via Simulation and Experiment

    PubMed Central

    Marzinek, Jan K.; Lakshminarayanan, Rajamani; Goh, Eunice; Huber, Roland G.; Panzade, Sadhana; Verma, Chandra; Bond, Peter J.

    2016-01-01

    Conformational changes in the envelope proteins of flaviviruses help to expose the highly conserved fusion peptide (FP), a region which is critical to membrane fusion and host cell infection, and which represents a significant target for antiviral drugs and antibodies. In principle, extended timescale atomic-resolution simulations may be used to characterize the dynamics of such peptides. However, the resultant accuracy is critically dependent upon both the underlying force field and sufficient conformational sampling. In the present study, we report a comprehensive comparison of three simulation methods and four force fields comprising a total of more than 40 μs of sampling. Additionally, we describe the conformational landscape of the FP fold across all flavivirus family members. All investigated methods sampled conformations close to available X-ray structures, but exhibited differently populated ensembles. The best force field / sampling combination was sufficiently accurate to predict that the solvated peptide fold is less ordered than in the crystallographic state, which was subsequently confirmed via circular dichroism and spectrofluorometric measurements. Finally, the conformational landscape of a mutant incapable of membrane fusion was significantly shallower than wild-type variants, suggesting that dynamics should be considered when therapeutically targeting FP epitopes. PMID:26785994

  17. Characterizing the Conformational Landscape of Flavivirus Fusion Peptides via Simulation and Experiment

    NASA Astrophysics Data System (ADS)

    Marzinek, Jan K.; Lakshminarayanan, Rajamani; Goh, Eunice; Huber, Roland G.; Panzade, Sadhana; Verma, Chandra; Bond, Peter J.

    2016-01-01

    Conformational changes in the envelope proteins of flaviviruses help to expose the highly conserved fusion peptide (FP), a region which is critical to membrane fusion and host cell infection, and which represents a significant target for antiviral drugs and antibodies. In principle, extended timescale atomic-resolution simulations may be used to characterize the dynamics of such peptides. However, the resultant accuracy is critically dependent upon both the underlying force field and sufficient conformational sampling. In the present study, we report a comprehensive comparison of three simulation methods and four force fields comprising a total of more than 40 μs of sampling. Additionally, we describe the conformational landscape of the FP fold across all flavivirus family members. All investigated methods sampled conformations close to available X-ray structures, but exhibited differently populated ensembles. The best force field / sampling combination was sufficiently accurate to predict that the solvated peptide fold is less ordered than in the crystallographic state, which was subsequently confirmed via circular dichroism and spectrofluorometric measurements. Finally, the conformational landscape of a mutant incapable of membrane fusion was significantly shallower than wild-type variants, suggesting that dynamics should be considered when therapeutically targeting FP epitopes.

  18. Structural similarity between β(3)-peptides synthesized from β(3)-homo-amino acids and aspartic acid monomers.

    PubMed

    Ahmed, Sahar; Sprules, Tara; Kaur, Kamaljit

    2014-07-01

    Formation of stable secondary structures by oligomers that mimic natural peptides is a key asset for enhanced biological response. Here we show that oligomeric β(3)-hexapeptides synthesized from L-aspartic acid monomers (β(3)-peptides 1, 5a, and 6) or homologated β(3)-amino acids (β(3)-peptide 2), fold into similar stable 14-helical secondary structures in solution, except that the former form right-handed 14-helix and the later form left-handed 14-helix. β(3)-Peptides from L-Asp monomers contain an additional amide bond in the side chains that provides opportunities for more hydrogen bonding. However, based on the NMR solution structures, we found that β(3)-peptide from L-Asp monomers (1) and from homologated amino acids (2) form similar structures with no additional side-chain interactions. These results suggest that the β(3)-peptides derived from L-Asp are promising peptide-mimetics that can be readily synthesized using L-Asp monomers as well as the right-handed 14-helical conformation of these β(3)-peptides (such as 1 and 6) may prove beneficial in the design of mimics for right-handed α-helix of α-peptides. © 2014 Wiley Periodicals, Inc.

  19. Conformational study of melectin and antapin antimicrobial peptides in model membrane environments

    NASA Astrophysics Data System (ADS)

    Kocourková, Lucie; Novotná, Pavlína; Čujová, Sabína; Čeřovský, Václav; Urbanová, Marie; Setnička, Vladimír

    2017-01-01

    Antimicrobial peptides have long been considered as promising compounds against drug-resistant pathogens. In this work, we studied the secondary structure of antimicrobial peptides melectin and antapin using electronic (ECD) and vibrational circular dichroism (VCD) spectroscopies that are sensitive to peptide secondary structures. The results from quantitative ECD spectral evaluation by Dichroweb and CDNN program and from the qualitative evaluation of the VCD spectra were compared. The antimicrobial activity of the selected peptides depends on their ability to adopt an amphipathic α-helical conformation on the surface of the bacterial membrane. Hence, solutions of different zwitterionic and negatively charged liposomes and micelles were used to mimic the eukaryotic and bacterial biological membranes. The results show a significant content of α-helical conformation in the solutions of negatively charged liposomes mimicking the bacterial membrane, thus correlating with the antimicrobial activity of the studied peptides. On the other hand in the solutions of zwitterionic liposomes used as models of the eukaryotic membranes, the fraction of α-helical conformation was lower, which corresponds with their moderate hemolytic activity.

  20. De novo design of conformationally flexible transmembrane peptides driving membrane fusion

    PubMed Central

    Hofmann, Mathias W.; Weise, Katrin; Ollesch, Julian; Agrawal, Prashant; Stalz, Holger; Stelzer, Walter; Hulsbergen, Frans; de Groot, Huub; Gerwert, Klaus; Reed, Jennifer; Langosch, Dieter

    2004-01-01

    Fusion of biological membranes is mediated by distinct integral membrane proteins, e.g., soluble N-ethylmaleimide-sensitive factor attachment protein receptors and viral fusion proteins. Previous work has indicated that the transmembrane segments (TMSs) of such integral membrane proteins play an important role in fusion. Furthermore, peptide mimics of the transmembrane part can drive the fusion of liposomes, and evidence had been obtained that fusogenicity depends on their conformational flexibility. To test this hypothesis, we present a series of unnatural TMSs that were designed de novo based on the structural properties of hydrophobic residues. We find that the fusogenicity of these peptides depends on the ratio of α-helix-promoting Leu and β-sheet-promoting Val residues and is enhanced by helix-destabilizing Pro and Gly residues within their hydrophobic cores. The ability of these peptides to refold from an α-helical state to a β-sheet conformation and backwards was determined under different conditions. Membrane fusogenic peptides with mixed Leu/Val sequences tend to switch more readily between different conformations than a nonfusogenic peptide with an oligo-Leu core. We propose that structural flexibility of these TMSs is a prerequisite of fusogenicity. PMID:15456911

  1. Nanotechnology for delivery of peptide nucleic acids (PNAs).

    PubMed

    Gupta, Anisha; Bahal, Raman; Gupta, Meera; Glazer, Peter M; Saltzman, W Mark

    2016-10-28

    Over the past three decades, peptide nucleic acids have been employed in numerous chemical and biological applications. Peptide nucleic acids possess enormous potential because of their superior biophysical properties, compared to other oligonucleotide chemistries. However, for therapeutic applications, intracellular delivery of peptide nucleic acids remains a challenge. In this review, we summarize the progress that has been made in delivering peptide nucleic acids to intracellular targets. In addition, we emphasize recent nanoparticle-based strategies for efficient delivery of conventional and chemically-modified peptides nucleic acids.

  2. Arginine changes the conformation of the arginine attenuator peptide relative to the ribosome tunnel

    PubMed Central

    Wu, Cheng; Wei, Jiajie; Lin, Pen-Jen; Tu, Liwei; Deutsch, Carol; Johnson, Arthur E.; Sachs, Matthew S.

    2012-01-01

    The fungal arginine attenuator peptide (AAP) is a regulatory peptide that controls ribosome function. As a nascent peptide within the ribosome exit tunnel, it acts to stall ribosomes in response to arginine (Arg). We used three approaches to probe the molecular basis for stalling. First, PEGylation assays revealed that the AAP did not undergo overall compaction in the tunnel in response to Arg. Second, site-specific photocrosslinking showed that Arg altered the conformation of the wild-type AAP, but not nonfunctional mutants, with respect to the tunnel. Third, using time-resolved spectral measurements with a fluorescent probe placed in the nascent AAP, we detected sequence-specific changes in the disposition of the AAP near the peptidyltransferase center in response to Arg. These data provide evidence that an Arg-induced change in AAP conformation and/or environment in the ribosome tunnel is important for stalling. PMID:22244852

  3. Arginine changes the conformation of the arginine attenuator peptide relative to the ribosome tunnel.

    PubMed

    Wu, Cheng; Wei, Jiajie; Lin, Pen-Jen; Tu, Liwei; Deutsch, Carol; Johnson, Arthur E; Sachs, Matthew S

    2012-03-02

    The fungal arginine attenuator peptide (AAP) is a regulatory peptide that controls ribosome function. As a nascent peptide within the ribosome exit tunnel, it acts to stall ribosomes in response to arginine (Arg). We used three approaches to probe the molecular basis for stalling. First, PEGylation assays revealed that the AAP did not undergo overall compaction in the tunnel in response to Arg. Second, site-specific photocross-linking showed that Arg altered the conformation of the wild-type AAP, but not of nonfunctional mutants, with respect to the tunnel. Third, using time-resolved spectral measurements with a fluorescent probe placed in the nascent AAP, we detected sequence-specific changes in the disposition of the AAP near the peptidyltransferase center in response to Arg. These data provide evidence that an Arg-induced change in AAP conformation and/or environment in the ribosome tunnel is important for stalling. Copyright © 2012 Elsevier Ltd. All rights reserved.

  4. Amyloid β peptide conformational changes in the presence of a lipid membrane system.

    PubMed

    Accardo, Angelo; Shalabaeva, Victoria; Cotte, Marine; Burghammer, Manfred; Krahne, Roman; Riekel, Christian; Dante, Silvia

    2014-03-25

    Here we are presenting a comparative analysis of conformational changes of two amyloid β peptides, Aβ(25-35) and Aβ(1-42), in the presence and absence of a phospholipid system, namely, POPC/POPS (1-palmitoyl-2-oleoylphospatidylcholine/palmitoyl-2-oleoylphospatidylserine), through Raman spectroscopy, synchrotron radiation micro Fourier-transform infrared spectroscopy, and micro X-ray diffraction. Ringlike samples were obtained from the evaporation of pure and mixed solutions of the proteins together with the POPC/POPS system on highly hydrophilic substrates. The results confirm the presence of a α-helical to β-sheet transition from the internal rim of the ringlike samples to the external one in the pure Aβ(25-35) residual, probably due to the convective flow inside the droplets sitting on highly hydrophilic substrates enhancing the local concentration of the peptide at the external edge of the dried drop. In contrast, the presence of POPC/POPS lipids in the peptide does not result in α-helical structures and introduces the presence of antiparallel β-sheet material together with parallel β-sheet structures and possible β-turns. As a control, Aβ(1-42) peptide was also tested and shows β-sheet conformations independently from the presence of the lipid system. The μXRD analysis further confirmed these conclusions, showing how the absence of the phospholipid system induces in the Aβ(25-35) a probable composite α/β material while its coexistence with the peptide leads to a not oriented β-sheet conformation. These results open interesting scenarios on the study of conformational changes of Aβ peptides and could help, with further investigations, to better clarify the role of enzymes and alternative lipid systems involved in the amyloidosis process of Aβ fragments.

  5. Molecular Dynamics Simulation of Tau Peptides for the Investigation of Conformational Changes Induced by Specific Phosphorylation Patterns.

    PubMed

    Gandhi, Neha S; Kukic, Predrag; Lippens, Guy; Mancera, Ricardo L

    2017-01-01

    The Tau protein plays an important role due to its biomolecular interactions in neurodegenerative diseases. The lack of stable structure and various posttranslational modifications such as phosphorylation at various sites in the Tau protein pose a challenge for many experimental methods that are traditionally used to study protein folding and aggregation. Atomistic molecular dynamics (MD) simulations can help around deciphering relationship between phosphorylation and various intermediate and stable conformations of the Tau protein which occur on longer timescales. This chapter outlines protocols for the preparation, execution, and analysis of all-atom MD simulations of a 21-amino acid-long phosphorylated Tau peptide with the aim of generating biologically relevant structural and dynamic information. The simulations are done in explicit solvent and starting from nearly extended configurations of the peptide. The scaled MD method implemented in AMBER14 was chosen to achieve enhanced conformational sampling in addition to a conventional MD approach, thereby allowing the characterization of folding for such an intrinsically disordered peptide at 293 K. Emphasis is placed on the analysis of the simulation trajectories to establish correlations with NMR data (i.e., chemical shifts and NOEs). Finally, in-depth discussions are provided for commonly encountered problems.

  6. Lipid Tail Protrusion in Simulations Predicts Fusogenic Activity of Influenza Fusion Peptide Mutants and Conformational Models

    PubMed Central

    Larsson, Per; Kasson, Peter M.

    2013-01-01

    Fusion peptides from influenza hemagglutinin act on membranes to promote membrane fusion, but the mechanism by which they do so remains unknown. Recent theoretical work has suggested that contact of protruding lipid tails may be an important feature of the transition state for membrane fusion. If this is so, then influenza fusion peptides would be expected to promote tail protrusion in proportion to the ability of the corresponding full-length hemagglutinin to drive lipid mixing in fusion assays. We have performed molecular dynamics simulations of influenza fusion peptides in lipid bilayers, comparing the X-31 influenza strain against a series of N-terminal mutants. As hypothesized, the probability of lipid tail protrusion correlates well with the lipid mixing rate induced by each mutant. This supports the conclusion that tail protrusion is important to the transition state for fusion. Furthermore, it suggests that tail protrusion can be used to examine how fusion peptides might interact with membranes to promote fusion. Previous models for native influenza fusion peptide structure in membranes include a kinked helix, a straight helix, and a helical hairpin. Our simulations visit each of these conformations. Thus, the free energy differences between each are likely low enough that specifics of the membrane environment and peptide construct may be sufficient to modulate the equilibrium between them. However, the kinked helix promotes lipid tail protrusion in our simulations much more strongly than the other two structures. We therefore predict that the kinked helix is the most fusogenic of these three conformations. PMID:23505359

  7. Lipid tail protrusion in simulations predicts fusogenic activity of influenza fusion peptide mutants and conformational models.

    PubMed

    Larsson, Per; Kasson, Peter M

    2013-01-01

    Fusion peptides from influenza hemagglutinin act on membranes to promote membrane fusion, but the mechanism by which they do so remains unknown. Recent theoretical work has suggested that contact of protruding lipid tails may be an important feature of the transition state for membrane fusion. If this is so, then influenza fusion peptides would be expected to promote tail protrusion in proportion to the ability of the corresponding full-length hemagglutinin to drive lipid mixing in fusion assays. We have performed molecular dynamics simulations of influenza fusion peptides in lipid bilayers, comparing the X-31 influenza strain against a series of N-terminal mutants. As hypothesized, the probability of lipid tail protrusion correlates well with the lipid mixing rate induced by each mutant. This supports the conclusion that tail protrusion is important to the transition state for fusion. Furthermore, it suggests that tail protrusion can be used to examine how fusion peptides might interact with membranes to promote fusion. Previous models for native influenza fusion peptide structure in membranes include a kinked helix, a straight helix, and a helical hairpin. Our simulations visit each of these conformations. Thus, the free energy differences between each are likely low enough that specifics of the membrane environment and peptide construct may be sufficient to modulate the equilibrium between them. However, the kinked helix promotes lipid tail protrusion in our simulations much more strongly than the other two structures. We therefore predict that the kinked helix is the most fusogenic of these three conformations.

  8. Experimental conformational energy maps of proteins and peptides.

    PubMed

    Balaji, Govardhan A; Nagendra, H G; Balaji, Vitukudi N; Rao, Shashidhar N

    2017-02-07

    We have presented an extensive analysis of the peptide backbone dihedral angles in the PDB structures and computed experimental Ramachandran plots for their distributions seen under a various constraints on X-ray resolution, representativeness at different sequence identity percentages, and hydrogen bonding distances. These experimental distributions have been converted into isoenergy contour plots using the approach employed previously by F. M. Pohl. This has led to the identification of energetically favored minima in the Ramachandran (ϕ, ψ) plots in which global minima are predominantly observed either in the right-handed α-helical or the polyproline II regions. Further, we have identified low energy pathways for transitions between various minima in the (ϕ,ψ) plots. We have compared and presented the experimental plots with published theoretical plots obtained from both molecular mechanics and quantum mechanical approaches. In addition, we have developed and employed a root mean square deviation (RMSD) metric for isoenergy contours in various ranges, as a measure (in kcal.mol(-1) ) to compare any two plots and determine the extent of correlation and similarity between their isoenergy contours. In general, we observe a greater degree of compatibility with experimental plots for energy maps obtained from molecular mechanics methods compared to most quantum mechanical methods. The experimental energy plots we have investigated could be helpful in refining protein structures obtained from X-ray, NMR, and electron microscopy and in refining force field parameters to enable simulations of peptide and protein structures that have higher degree of consistency with experiments. Proteins 2017. © 2017 Wiley Periodicals, Inc.

  9. Conformation of poly(γ-glutamic acid) in aqueous solution.

    PubMed

    Muroga, Yoshio; Nakaya, Asami; Inoue, Atsuki; Itoh, Daiki; Abiru, Masaya; Wada, Kaori; Takada, Masako; Ikake, Hiroki; Shimizu, Shigeru

    2016-04-01

    Local conformation and overall conformation of poly(γ-DL-glutamic acid) (PγDLGA) and poly(γ-L-glutamic acid) (PγLGA) in aqueous solution was studied as a function of degree of ionization ε by (1) H-NMR, circular dichroism, and potentiometric titration. It was clarified that their local conformation is represented by random coil over an entire ε range and their overall conformation is represented by expanded random-coil in a range of ε > ε(*) , where ε(*) is about 0.3, 0.35, 0.45, and 0.5 for added-salt concentration of 0.02M, 0.05M, 0.1M, and 0.2M, respectively. In a range of ε < ε(*) , however, ε dependence of their overall conformation is significantly differentiated from each other. PγDLGA tends to aggregate intramolecularly and/or intermolecularly with decreasing ε, but PγLGA still behaves as expanded random-coil. It is speculated that spatial arrangement of adjacent carboxyl groups along the backbone chain essentially affects the overall conformation of PγGA in acidic media.

  10. The Periplasmic Bacterial Molecular Chaperone SurA Adapts Its Structure to Bind Peptides in Different Conformations to Assert a Sequence Preference for Aromatic Residues

    SciTech Connect

    Xu, X.; Wang, S.; Hu, Y.-X.; McKay, D.B.

    2009-06-04

    The periplasmic molecular chaperone protein SurA facilitates correct folding and maturation of outer membrane proteins in Gram-negative bacteria. It preferentially binds peptides that have a high fraction of aromatic amino acids. Phage display selections, isothermal titration calorimetry and crystallographic structure determination have been used to elucidate the basis of the binding specificity. The peptide recognition is imparted by the first peptidyl-prolyl isomerase (PPIase) domain of SurA. Crystal structures of complexes between peptides of sequence WEYIPNV and NFTLKFWDIFRK with the first PPIase domain of the Escherichia coli SurA protein at 1.3 A resolution, and of a complex between the dodecapeptide and a SurA fragment lacking the second PPIase domain at 3.4 A resolution, have been solved. SurA binds as a monomer to the heptapeptide in an extended conformation. It binds as a dimer to the dodecapeptide in an alpha-helical conformation, predicated on a substantial structural rearrangement of the SurA protein. In both cases, side-chains of aromatic residues of the peptides contribute a large fraction of the binding interactions. SurA therefore asserts a recognition preference for aromatic amino acids in a variety of sequence configurations by adopting alternative tertiary and quaternary structures to bind peptides in different conformations.

  11. Comparison of the solution conformations of a cell-adhesive peptide LBE and its reverse sequence EBL.

    PubMed

    Jois, S D; Hughes, R; Siahaan, T J

    1999-12-01

    T-cell adhesion is mediated by an ICAM-1/LFA-1 interaction; this interaction plays a crucial role in T-cell activation during immune response. LBE peptide, which is derived from the beta-subunit of LFA-1, has been shown to inhibit ICAM-1/LFA-1-mediated T-cell adhesion. In this work, we studied the solution conformations of LBE peptide and its reverse sequence (EBL) by NMR, CD and molecular dynamics simulations. Reverse peptides have been used as controls in biological studies. The effect of reversing the sequence of LBE to EBL peptides on their respective conformations is important in understanding their biological properties in vitro or in vivo. The NMR studies for these peptides were carried out in water and in TFE/water solvent systems. In 40% TFE/water, both peptides exhibited helical conformation. CD studies suggested that the LBE exhibits 30% helical conformation, while the EBL exhibits 20% helical conformation. From the NMR and MD simulation studies, it was evident that the peptides exhibited a stable helical conformation; a stable helical structure was found at Leu6 to Leu15 for LBE and at Gly9 to Leu17 for EBL. The helical conformations of LBE and EBL may be in equilibrium with other possible conformers; the other conformers contain loop and turn structures. Both peptides bind to divalent cations because the LBE is derived from the cation-binding region of the LFA-1. This study shows that reversing the peptide sequence did not alter the secondary structure of the corresponding sequence. Hence, caution must be exercised when using reverse peptides as controls in biological studies. This report will improve our ability to design a better inhibitor of ICAM-1/LFA-1 interaction.

  12. The key position: influence of staple location on constrained peptide conformation and binding.

    PubMed

    Keeling, Kelly L; Cho, Okki; Scanlon, Denis B; Booker, Grant W; Abell, Andrew D; Wegener, Kate L

    2016-10-18

    Constrained α-helical peptides are showing potential as biological probes and therapeutic agents that target protein-protein interactions. However, the factors that determine the optimal constraint locations are still largely unknown. Using the β-integrin/talin protein interaction as a model system, we examine the effect of constraint location on helical conformation, as well as binding affinity, using circular dichroism and NMR spectroscopy. Stapling increased the overall helical content of each integrin-based peptide tested. However, NMR analysis revealed that different regions within the peptide are stabilised, depending on constraint location, and that these differences correlate with the changes observed in talin binding mode and affinity. In addition, we show that examination of the atomic structure of the parent peptide provides insight into the appropriate placement of helical constraints.

  13. Cross-Correlation Effects on NMR Lineshapes and Peptide Conformation

    NASA Astrophysics Data System (ADS)

    Čuperlović, Miroslava; Palke, William E.; Gerig, J. T.; Gray, G. A.

    1996-01-01

    Information about molecular structure and dynamics can potentially be obtained by studying dipole-dipole and chemical-shift anisotropy (CSA) auto-correlation and dipole-CSA cross-correlation effects in high-resolution NMR spectra. Equations for the lineshapes of the HNmultiplet in the fragment-15NH-CH- as a function of NH-CH dihedral angle are derived by including these effects within the framework of the Redfield treatment of relaxation. To test the utility of the theoretical results,1H{15N} HSQC proton lineshape data for a variant of the enzyme staphylococcal nuclease in which all valine residues are labeled with15N have been analyzed to obtain the conformational angle (φ) between the N-H and adjacent C-H bonds. The results are generally in good agreement with values of φ obtained from crystal structure data. Considerations in the further development of the analysis of the lineshape of the HNmultiplet for experimental determinations of φ are discussed.

  14. Effects of glycosylation on peptide conformation: a synergistic experimental and computational study.

    PubMed

    Bosques, Carlos J; Tschampel, Sarah M; Woods, Robert J; Imperiali, Barbara

    2004-07-14

    Asparagine-linked glycosylation, the co-translational covalent attachment of carbohydrates to asparagine side chains, has a major effect on the folding, stability, and function of many proteins. The carbohydrate composition in mature glycoproteins is heterogeneous due to modification of the initial oligosaccharide by glycosidases and glycosyltransferases during the glycoprotein passage through the endoplasmic reticulum and Golgi apparatus. Despite the diversity of carbohydrate structures, the core beta-D-(GlcNAc)(2) remains conserved in all N-linked glycoproteins. Previously, results from our laboratory showed that the molecular composition of the core disaccharide has a critical and unique conformational effect on the peptide backbone. Herein, we employ a synergistic experimental and computational approach to study the effect of the stereochemistry of the carbohydrate--peptide linkage on glycopeptide structure. A glycopeptide derived from a hemagglutinin protein fragment was synthesized, with the carbohydrate attached to the peptide with an alpha-linked stereochemistry. Computational and biophysical analyses reveal that the conformations of the peptide and alpha- and beta-linked glycopeptides are uniquely influenced by the attached saccharide. The value of computational approaches for probing the influence of attached saccharides on polypeptide conformation is highlighted.

  15. Effects of Glycosylation on Peptide Conformation: A Synergistic Experimental and Computational Study

    PubMed Central

    Bosques, Carlos J.; Tschampel, Sarah M.; Woods, Robert J.; Imperiali, Barbara

    2006-01-01

    Asparagine-linked glycosylation, the co-translational covalent attachment of carbohydrates to asparagine side chains, has a major effect on the folding, stability, and function of many proteins. The carbohydrate composition in mature glycoproteins is heterogeneous due to modification of the initial oligosaccharide by glycosidases and glycosyltransferases during the glycoprotein passage through the endoplasmic reticulum and Golgi apparatus. Despite the diversity of carbohydrate structures, the core β-d-(GlcNAc)2 remains conserved in all N-linked glycoproteins. Previously, results from our laboratory showed that the molecular composition of the core disaccharide has a critical and unique conformational effect on the peptide backbone. Herein, we employ a synergistic experimental and computational approach to study the effect of the stereochemistry of the carbohydrate–peptide linkage on glycopeptide structure. A glycopeptide derived from a hemagglutinin protein fragment was synthesized, with the carbohydrate attached to the peptide with an α-linked stereochemistry. Computational and biophysical analyses reveal that the conformations of the peptide and α- and β-linked glycopeptides are uniquely influenced by the attached saccharide. The value of computational approaches for probing the influence of attached saccharides on polypeptide conformation is highlighted. PMID:15237998

  16. Predictable conformational diversity in foldamers of sugar amino acids.

    PubMed

    Menyhard, Dora K; Hudaky, Ilona; Jákli, Imre; Juhász, György; Perczel, András

    2017-03-27

    Systematic conformational search was carried out for monomers and homohexamers of furanoid β-amino acids: cis-(S,R) and trans-(S,S) stereoisomers of aminocyclopentane carboxylic acid (ACPC), two different aminofuranuronic-acids (AFU(α) and AFU(β)), their isopropylidene derivatives (AFU(ip)) as well as the key intermediate β-aminotetrahydrofurancarboxylic acid (ATFC). Stereochemistry of the building blocks was chosen to match with that of natural sugar amino acid (xylose and ribose) precursors. Results show that hexamers of cis furanoid β-amino acids show great variability: while hydrophobic cyclopentane (cis(ACPC)6), and hydrophilic (cisXylAFU(α/β))6 foldamers favor two different zigzagged conformation as hexamers, the backbone fold turns into a helix in case of (cisATFC)6 (10-helix) and (cisAFU(ip))6 (14-helix). Trans stereochemistry resulted in hexamers exclusively of right-handed helix conformation, (H12(P))6, regardless of their polarity. We found that the preferred oligomeric structure of cis/(S,R)AFU(α/β) is conformationally compatible with β-pleated sheets, while that of the trans/(S,S) units match with α-helices of α-proteins.

  17. The Unexpected Advantages of Using D-Amino Acids for Peptide Self-Assembly into Nanostructured Hydrogels for Medicine

    PubMed Central

    Melchionna, Michele; Styan, Katie E.; Marchesan, Silvia

    2016-01-01

    Self-assembled peptide hydrogels have brought innovation to the medicinal field, not only as responsive biomaterials but also as nanostructured therapeutic agents or as smart drug delivery systems. D-amino acids are typically introduced to increase the peptide enzymatic stability. However, there are several reports of unexpected effects on peptide conformation, self-assembly behavior, cytotoxicity and even therapeutic activity. This mini-review discusses all the surprising twists of heterochiral self-assembled peptide hydrogels, and delineates emerging key findings to exploit all the benefits of D-amino acids in this novel medicinal area. PMID:26876522

  18. 4-Fluoroproline derivative peptides: effect on PPII conformation and SH3 affinity.

    PubMed

    Ruzza, Paolo; Siligardi, Giuliano; Donella-Deana, Arianna; Calderan, Andrea; Hussain, Rohanah; Rubini, Chiara; Cesaro, Luca; Osler, Alessio; Guiotto, Andrea; Pinna, Lorenzo A; Borin, Gianfranco

    2006-07-01

    Eukaryotic signal transduction involves the assembly of transient protein-protein complexes mediated by modular interaction domains. Specific Pro-rich sequences with the consensus core motif PxxP adopt the PPII helix conformation upon binding to SH3 domains. For short Pro-rich peptides, little or no ordered secondary structure is usually observed before binding interactions. The association of a Pro-rich peptide with the SH3 domain involves unfavorable binding entropy due to the loss of rotational freedom on forming the PPII helix. With the aim of stabilizing the PPII helix conformation in the Pro-rich HPK1 decapeptide PPPLPPKPKF (P2), a series of P2 analogues was prepared, in which specific Pro positions were alternatively occupied by 4(S)- or 4(R)-4-fluoro-L-proline. The interactions of these peptides with the SH3 domain of the HPK1-binding partner HS1 were quantitatively analyzed by the NILIA-CD approach. A CD thermal analysis of the P2 analogues was performed to assess their propensity to adopt the PPII helix conformation. Contrary to our expectations, the K(d) values of the analogues were lower than that of the parent peptide P2. These results clearly show that the induction of a stable PPII helix conformation in short Pro-rich peptides is not sufficient to increase their affinity toward the SH3 domain and that the effect of 4-fluoroproline strongly depends on the position of this residue in the sequence and the chirality of the substituent in the pyrrolidine ring.

  19. Solution conformation of peptides by the intramolecular nuclear Overhauser effect experiment. Study of valinomycin-K+.

    PubMed Central

    Krishna, N R; Agresti, D G; Glickson, J D; Walter, R

    1978-01-01

    This study demonstrates how the intramolecular nuclear Overhauser effect (NOE) experiment can be employed quantitatively to select from a set of possible conformations for a peptide or a protein the particular conformation (or a group of conformations) most consistent with the data. This procedure is demonstrated on a model depsipeptide system--valinomycin K+ in CDCl3--for which the solution conformation has been inferred by other methods. The NOE enhancements are very sensitive to the conformations assumed by this antibiotic. It is shown that the set of conformations, collectively labeled as A2 (including the X-ray crystallographic structure) gives a very good description of the NOE enhancements. The structure proposed by Bystrov et al. (1977. Eur. J. Biochem. 78:63) for the uncomplexed valinomycin in nonpolar solvents is also consistent with the experimental data on the potassium complex. Using statistical hypothesis testing involving the Hamilton R-factor ratio criterion, all the other models have been rejected as inconsistent with the experimental data. A general formalism is presented for describing the NOE effects in isotropically reorienting molecules. The formalism is not restricted to the extreme narrowing limit of the rotational correlation times and hence applies to both small and large molecules. Some of the factors that can influence the NOE measurements, viz. anisotropic rotational diffusion, conformational averaging, and nuclear spin diffusion, have been considered in this study. PMID:737287

  20. Mapping the Conformational Dynamics and Pathways of Spontaneous Steric Zipper Peptide Oligomerization

    PubMed Central

    Matthes, Dirk; Gapsys, Vytautas; Daebel, Venita; de Groot, Bert L.

    2011-01-01

    The process of protein misfolding and self-assembly into various, polymorphic aggregates is associated with a number of important neurodegenerative diseases. Only recently, crystal structures of several short peptides have provided detailed structural insights into -sheet rich aggregates, known as amyloid fibrils. Knowledge about early events of the formation and interconversion of small oligomeric states, an inevitable step in the cascade of peptide self-assembly, however, remains still limited. We employ molecular dynamics simulations in explicit solvent to study the spontaneous aggregation process of steric zipper peptide segments from the tau protein and insulin in atomistic detail. Starting from separated chains with random conformations, we find a rapid formation of structurally heterogeneous, -sheet rich oligomers, emerging from multiple bimolecular association steps and diverse assembly pathways. Furthermore, our study provides evidence that aggregate intermediates as small as dimers can be kinetically trapped and thus affect the structural evolution of larger oligomers. Alternative aggregate structures are found for both peptide sequences in the different independent simulations, some of which feature characteristics of the known steric zipper conformation (e.g., -sheet bilayers with a dry interface). The final aggregates interconvert with topologically distinct oligomeric states exclusively via internal rearrangements. The peptide oligomerization was analyzed through the perspective of a minimal oligomer, i.e., the dimer. Thereby all observed multimeric aggregates can be consistently mapped onto a space of reduced dimensionality. This novel method of conformational mapping reveals heterogeneous association and reorganization dynamics that are governed by the characteristics of peptide sequence and oligomer size. PMID:21559277

  1. Interactions of a designed peptide with lipopolysaccharide: Bound conformation and anti-endotoxic activity

    SciTech Connect

    Bhunia, Anirban; Chua, Geok Lin; Domadia, Prerna N.; Warshakoon, Hemamali; Cromer, Jens R.; David, Sunil A.; Bhattacharjya, Surajit

    2008-05-09

    Designed peptides that would selectively interact with lipopolysaccharide (LPS) or endotoxin and fold into specific conformations could serve as important scaffolds toward the development of antisepsis compounds. Here, we describe solution structure of a designed amphipathic peptide, H{sub 2}N-YVKLWRMIKFIR-CONH{sub 2} (YW12D) in complex with endotoxin as determined by transferred nuclear Overhauser effect spectroscopy. The conformation of the isolated peptide is highly flexible, but undergoes a dramatic structural stabilization in the presence of LPS. Structure calculations reveal that the peptide presents two amphipathic surfaces in its bound state to LPS whereby each surface is characterized by two positive charges and a number of aromatic and/or aliphatic residues. ITC data suggests that peptide interacts with two molecules of lipid A. In activity assays, YW12D exhibits neutralization of LPS toxicity with very little hemolysis of red blood cells. Structural and functional properties of YW12D would be applicable in designing low molecular weight non-toxic antisepsis molecules.

  2. Mapping the bound conformation and protein interactions of microtubule destabilizing peptides by STD-NMR spectroscopy.

    PubMed

    Milton, Mark J; Thomas Williamson, R; Koehn, Frank E

    2006-08-15

    Using the hemiasterlin analogs taltobulin (I, HTI-286), II, and III as model compounds, we demonstrate that relaxation-compensated STD-NMR can be used as an effective tool to efficiently provide a qualitative epitope map for microtubule destabilizing peptides. Due to the disparate relaxation behavior of the protons in these model compounds, it was essential to collect STD with very short saturation times to render an accurate picture of the binding interaction. The conformation of HTI-286 (I) in complex with the protein was determined from TRNOESY/ROESY experiments and is similar to the X-ray crystal structure conformation observed for hemiasterlin methyl ester in the absence of protein.

  3. Side-chain conformational space analysis (SCSA): a multi conformation-based QSAR approach for modeling and prediction of protein-peptide binding affinities.

    PubMed

    Zhou, Peng; Chen, Xiang; Shang, Zhicai

    2009-03-01

    In this article, the concept of multi conformation-based quantitative structure-activity relationship (MCB-QSAR) is proposed, and based upon that, we describe a new approach called the side-chain conformational space analysis (SCSA) to model and predict protein-peptide binding affinities. In SCSA, multi-conformations (rather than traditional single-conformation) have received much attention, and the statistical average information on multi-conformations of side chains is determined using self-consistent mean field theory based upon side chain rotamer library. Thereby, enthalpy contributions (including electrostatic, steric, hydrophobic interaction and hydrogen bond) and conformational entropy effects to the binding are investigated in terms of occurrence probability of residue rotamers. Then, SCSA was applied into the dataset of 419 HLA-A 0201 binding peptides, and nonbonding contributions of each position in peptide ligands are well determined. For the peptides, the hydrogen bond and electrostatic interactions of the two ends are essential to the binding specificity, van der Waals and hydrophobic interactions of all the positions ensure strong binding affinity, and the loss of conformational entropy at anchor positions partially counteracts other favorable nonbonding effects.

  4. Conformational preferences of 3,4-dihydroxyphenylacetic acid (DOPAC).

    PubMed

    Lopes Jesus, A J; Jarmelo, S; Fausto, R; Reva, I

    2015-04-05

    The conformational space of 3,4-dihydroxyphenylacetic acid (DOPAC), an important dopamine metabolite, has been investigated by quantum chemical methods (B3LYP and MP2, with the 6-311++G(d,p) basis set) and matrix-isolation infrared spectroscopy. Detailed analysis of the calculated potential energy surfaces of the molecule led to identification of thirteen unique conformers, all of them showing the acetic acid side chain out of the aromatic ring plane by 60-95°. According to the calculated Gibbs energies, the five lowest energy conformers make up 99.7% of the conformational mixture at 298.15K, exhibiting individual populations falling between 16% and 24%. The main conformational trends of this molecule were interpreted on the grounds of a thorough analysis of the structural parameters and by the application of the Natural Bond Orbital theory. The role of the intramolecular interactions on the relative stability and structure of the conformers was also investigated. The infrared spectrum of DOPAC was registered after isolation of its monomers in argon and xenon matrices. Only one of DOPAC forms populated in the gas phase could be trapped in both matrix gases. This result is in agreement with the predicted low energy barriers for conformational isomerization and is also supported by annealing experiments. The spectra of matrix-isolated model compounds, phenylacetic acid and catechol, were studied under the same experimental conditions. These data were used as references and assisted in the interpretation of the results obtained for DOPAC. Copyright © 2014 Elsevier B.V. All rights reserved.

  5. Conformational studies of the glycopeptide Ac-Tyr-[Man5GlcNAc-beta-(1-->4)GlcNAc-beta-(1-->Ndelta)]-Asn-Leu-Thr-Se r-OBz and the constituent peptide and oligosaccharide.

    PubMed

    Bailey, D; Renouf, D V; Large, D G; Warren, C D; Hounsell, E F

    2000-03-10

    Glycopeptides of desired structure can be conveniently prepared by the coupling of reducing oligosaccharides to aspartic acid of peptides via their glycosylamines formed in the presence of saturated aqueous ammonium hydrogen carbonate. The resulting oligosaccharide chains are N-linked to asparagine as in natural glycoproteins, allowing different peptide oligosaccharide combinations to be analysed for conformational effects. In the present paper, a pentapeptide of ovalbumin was coupled to Man5GlcNAc2 oligosaccharide and the glycopeptide and the two parent compounds compared by NMR ROESY experiments and molecular dynamics simulations. Despite the small size of the peptide, conformational effects were observed suggestive of the oligosaccharide stabilising the peptide in solution and of the peptide influencing oligosaccharide conformation. These effects are relevant to the function of glycosylation and the enzymic processing of oligosaccharide chains.

  6. Cu nanocrystal growth on peptide nanotubes by biomineralization: Size control of Cu nanocrystals by tuning peptide conformation

    NASA Astrophysics Data System (ADS)

    Banerjee, Ipsita A.; Yu, Lingtao; Matsui, Hiroshi

    2003-12-01

    With recent interest in seeking new biologically inspired device-fabrication methods in nanotechnology, a new biological approach was examined to fabricate Cu nanotubes by using sequenced histidine-rich peptide nanotubes as templates. The sequenced histidine-rich peptide molecules were assembled as nanotubes, and the biological recognition of the specific sequence toward Cu lead to efficient Cu coating on the nanotubes. Cu nanocrystals were uniformly coated on the histidine-incorporated nanotubes with high packing density. In addition, the diameter of Cu nanocrystal was controlled between 10 and 30 nm on the nanotube by controlling the conformation of histidine-rich peptide by means of pH changes. Those nanotubes showed significant change in electronic structure by varying the nanocrystal diameter; therefore, this system may be developed to a conductivity-tunable building block for microelectronics and biological sensors. This simple biomineralization method can be applied to fabricate various metallic and semiconductor nanotubes with peptides whose sequences are known to mineralize specific ions.

  7. The Presence of Two Cyclase Thioesterases Expands the Conformational Freedom of the Cyclic Peptide Occidiofungin

    PubMed Central

    Ravichandran, Akshaya; Gu, Ganyu; Escano, Jerome; Lu, Shi-En; Smith, Leif

    2014-01-01

    Occidiofungin is a cyclic nonribosomally synthesized antifungal peptide with submicromolar activity produced by Gram-negative bacterium Burkholderia contaminans. The biosynthetic gene cluster was confirmed to contain two cyclase thioesterases. NMR analysis revealed that the presence of both thioesterases is used to increase the conformational repertoire of the cyclic peptide. The loss of the OcfN cyclic thioesterase by mutagenesis results in a reduction of conformational variants and an appreciable decrease in bioactivity against Candida species. Presumably, the presence of both asparagine and β-hydroxyasparagine variants coordinate the enzymatic function of both of the cyclase thioesterases. OcfN has presumably evolved to be part of the biosynthetic gene cluster due to its ability to produce structural variants that enhance antifungal activity against some fungi. The enhancement of the antifungal activity from the incorporation of an additional cyclase thioesterase into the biosynthetic gene cluster of occidiofungin supports the need to explore new conformational variants of other therapeutic or potentially therapeutic cyclic peptides. PMID:23394257

  8. The presence of two cyclase thioesterases expands the conformational freedom of the cyclic Peptide occidiofungin.

    PubMed

    Ravichandran, Akshaya; Gu, Ganyu; Escano, Jerome; Lu, Shi-En; Smith, Leif

    2013-02-22

    Occidiofungin is a cyclic nonribosomally synthesized antifungal peptide with submicromolar activity produced by the Gram-negative bacterium Burkholderia contaminans. The biosynthetic gene cluster was confirmed to contain two cyclase thioesterases. NMR analysis revealed that the presence of both thioesterases is used to increase the conformational repertoire of the cyclic peptide. The loss of the OcfN cyclic thioesterase by mutagenesis results in a reduction of conformational variants and an appreciable decrease in bioactivity against Candida species. Presumably, the presence of both asparagine and β-hydroxyasparagine variants coordinates the enzymatic function of both of the cyclase thioesterases. OcfN has presumably evolved to be part of the biosynthetic gene cluster due to its ability to produce structural variants that enhance antifungal activity against some fungi. The enhancement of the antifungal activity from the incorporation of an additional cyclase thioesterase into the biosynthetic gene cluster of occidiofungin supports the need to explore new conformational variants of other therapeutic or potentially therapeutic cyclic peptides.

  9. Vibrational infrared conformational studies of model peptides representing the semicrystalline domains of Bombyx mori silk fibroin.

    PubMed

    Taddei, Paola; Monti, Patrizia

    2005-08-05

    The structural organization of Bombyx mori silk fibroin was investigated by infrared (IR) spectroscopy. To this aim, (AG)15 and other model peptides of varying chain length, containing tyrosine (Y), valine (V), and serine (S) in the basic (AG)n sequence were synthesized by the solid phase method and their spectroscopic properties were determined. Both the position and the relative content of Y, V, and S residues in the (AG)n model system appeared critical in determining the preferred conformation, i.e., silk I, silk II, and unordered structures. Curve fitting analysis in the amide I range showed that the model peptides with prevailing silk II structure displayed different beta-sheet content, which was dependent on the degree of interruption of the (AG)n sequence. In this regard, the bands at about 1000 and 980 cm(-1), specifically assigned to the AG sequence of the B. mori silk fibroin chain, were identified as marker of the degree of interruption of the (AG)n sequence.A stable silk I structure was observed only when the Y residue was located near the chain terminus, while a silk I --> silk II conformational transition occurred when it was positioned in the central region of the peptide. Analysis of the second-derivative spectra in the amide I range allowed us to identify a band at 1639 cm(-1) (4 --> 1 hydrogen-bonded type II beta-turns), which is characteristic of the silk I conformation.

  10. Glutamic Acid Selective Chemical Cleavage of Peptide Bonds.

    PubMed

    Nalbone, Joseph M; Lahankar, Neelam; Buissereth, Lyssa; Raj, Monika

    2016-03-04

    Site-specific hydrolysis of peptide bonds at glutamic acid under neutral aqueous conditions is reported. The method relies on the activation of the backbone amide chain at glutamic acid by the formation of a pyroglutamyl (pGlu) imide moiety. This activation increases the susceptibility of a peptide bond toward hydrolysis. The method is highly specific and demonstrates broad substrate scope including cleavage of various bioactive peptides with unnatural amino acid residues, which are unsuitable substrates for enzymatic hydrolysis.

  11. Conformation of protonated glutamic acid at room and cryogenic temperatures.

    PubMed

    Bouchet, Aude; Klyne, Johanna; Ishiuchi, Shun-Ichi; Fujii, Masaaki; Dopfer, Otto

    2017-05-03

    Recognition properties of biologically relevant molecules depend on their conformation. Herein, the conformation of protonated glutamic acid (H(+)Glu) isolated in quadruple ion traps is characterized by vibrational spectroscopy at room and cryogenic temperatures and dispersion-corrected density functional theory calculations at the B3LYP-D3/aug-cc-pVTZ level. The infrared multiple photon dissociation (IRMPD) spectrum recorded in the fingerprint range at room temperature using an IR free electron laser is attributed to the two most stable and nearly isoenergetic conformations (1-cc and 2-cc) with roughly equal population (ΔG298 = 0.0 kJ mol(-1)). Both have bridging C[double bond, length as m-dash]O(HNH)(+)O[double bond, length as m-dash]C ionic H-bonds of rather different strengths but cannot be distinguished by their similar IRMPD spectra. In contrast, the higher-resolution single-photon IRPD spectrum of H2-tagged H(+)Glu recorded in the conformation-sensitive X-H stretch range in a trap held at 10 K distinguishes both conformers. At low temperature, 1-cc is roughly twice more abundant than 2-cc, in line with its slightly lower calculated energy (ΔE0 = 0.5 kJ mol(-1)). This example illustrates the importance of cryogenic cooling, single-photon absorption conditions, and the consideration of the X-H stretch range for the identification of biomolecular conformations involving hydrogen bonds.

  12. Subtle conformational changes induced in major histocompatibility complex class II molecules by binding peptides.

    PubMed

    Chervonsky, A V; Medzhitov, R M; Denzin, L K; Barlow, A K; Rudensky, A Y; Janeway, C A

    1998-08-18

    Intracellular trafficking of major histocompatibility complex (MHC) class II molecules is characterized by passage through specialized endocytic compartment(s) where antigenic peptides replace invariant chain fragments in the presence of the DM protein. These changes are accompanied by structural transitions of the MHC molecules that can be visualized by formation of compact SDS-resistant dimers, by changes in binding of mAbs, and by changes in T cell responses. We have observed that a mAb (25-9-17) that is capable of staining I-Ab on the surface of normal B cells failed to interact with I-Ab complexes with a peptide derived from the Ealpha chain of the I-E molecule but bound a similar covalent complex of I-Ab with the class II binding fragment (class II-associated invariant chain peptides) of the invariant chain. Moreover, 25-9-17 blocked activation of several I-Ab-reactive T cell hybridomas but failed to block others, suggesting that numerous I-Ab-peptide complexes acquire the 25-9-17(+) or 25-9-17(-) conformation. Alloreactive T cells were also able to discriminate peptide-dependent variants of MHC class II molecules. Thus, peptides impose subtle structural transitions upon MHC class II molecules that affect T cell recognition and may thus be critical for T cell selection and autiommunity.

  13. Conformational photoswitching of a synthetic peptide foldamer bound within a phospholipid bilayer.

    PubMed

    De Poli, Matteo; Zawodny, Wojciech; Quinonero, Ophélie; Lorch, Mark; Webb, Simon J; Clayden, Jonathan

    2016-04-29

    The dynamic properties of foldamers, synthetic molecules that mimic folded biomolecules, have mainly been explored in free solution. We report on the design, synthesis, and conformational behavior of photoresponsive foldamers bound in a phospholipid bilayer akin to a biological membrane phase. These molecules contain a chromophore, which can be switched between two configurations by different wavelengths of light, attached to a helical synthetic peptide that both promotes membrane insertion and communicates conformational change along its length. Light-induced structural changes in the chromophore are translated into global conformational changes, which are detected by monitoring the solid-state (19)F nuclear magnetic resonance signals of a remote fluorine-containing residue located 1 to 2 nanometers away. The behavior of the foldamers in the membrane phase is similar to that of analogous compounds in organic solvents. Copyright © 2016, American Association for the Advancement of Science.

  14. How accurately do current force fields predict experimental peptide conformations? An adiabatic free energy dynamics study.

    PubMed

    Tzanov, Alexandar T; Cuendet, Michel A; Tuckerman, Mark E

    2014-06-19

    The quality of classical biomolecular simulations is inevitably limited by two problems: the accuracy of the force field used and the comprehensiveness of configuration space sampling. In this work we tackle the sampling problem by carrying out driven adiabatic free energy dynamics to obtain converged free energy surfaces of dipeptides in the gas phase and in solution using selected dihedral angles as collective variables. To calculate populations of conformational macrostates observed in experiment, we introduce a fuzzy clustering algorithm in collective-variable space, which delineates macrostates without prior definition of arbitrary boundaries. With this approach, we calculate the conformational preferences of small peptides with six biomolecular force fields chosen from among the most recent and widely used. We assess the accuracy of each force field against recently published Raman or IR-UV spectroscopy measurements of conformer populations for the dipeptides in solution or in the gas phase.

  15. Conformational analysis of glutamic acid: a density functional approach using implicit continuum solvent model.

    PubMed

    Turan, Başak; Selçuki, Cenk

    2014-09-01

    Amino acids are constituents of proteins and enzymes which take part almost in all metabolic reactions. Glutamic acid, with an ability to form a negatively charged side chain, plays a major role in intra and intermolecular interactions of proteins, peptides, and enzymes. An exhaustive conformational analysis has been performed for all eight possible forms at B3LYP/cc-pVTZ level. All possible neutral, zwitterionic, protonated, and deprotonated forms of glutamic acid structures have been investigated in solution by using polarizable continuum model mimicking water as the solvent. Nine families based on the dihedral angles have been classified for eight glutamic acid forms. The electrostatic effects included in the solvent model usually stabilize the charged forms more. However, the stability of the zwitterionic form has been underestimated due to the lack of hydrogen bonding between the solute and solvent; therefore, it is observed that compact neutral glutamic acid structures are more stable in solution than they are in vacuum. Our calculations have shown that among all eight possible forms, some are not stable in solution and are immediately converted to other more stable forms. Comparison of isoelectronic glutamic acid forms indicated that one of the structures among possible zwitterionic and anionic forms may dominate over the other possible forms. Additional investigations using explicit solvent models are necessary to determine the stability of charged forms of glutamic acid in solution as our results clearly indicate that hydrogen bonding and its type have a major role in the structure and energy of conformers.

  16. Exploring the Alzheimer amyloid-β peptide conformational ensemble: A review of molecular dynamics approaches.

    PubMed

    Tran, Linh; Ha-Duong, Tâp

    2015-07-01

    Alzheimer's disease is one of the most common dementia among elderly worldwide. There is no therapeutic drugs until now to treat effectively this disease. One main reason is due to the poorly understood mechanism of Aβ peptide aggregation, which plays a crucial role in the development of Alzheimer's disease. It remains challenging to experimentally or theoretically characterize the secondary and tertiary structures of the Aβ monomer because of its high flexibility and aggregation propensity, and its conformations that lead to the aggregation are not fully identified. In this review, we highlight various structural ensembles of Aβ peptide revealed and characterized by computational approaches in order to find converging structures of Aβ monomer. Understanding how Aβ peptide forms transiently stable structures prior to aggregation will contribute to the design of new therapeutic molecules against the Alzheimer's disease. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Conformational preferences of 1-amino-2-phenylcyclohexanecarboxylic acid, a phenylalanine cyclohexane analogue

    PubMed Central

    Alemán, Carlos; Jiménez, Ana I.; Cativiela, Carlos; Nussinov, Ruth; Casanovas, Jordi

    2009-01-01

    The intrinsic conformational preferences of the restricted phenylalanine analogue generated by including the α and β carbon atoms into a cyclohexane ring (1-amino-2-phenylcyclohexanecarboxylic acid, c6Phe) have been determined using quantum mechanical calculations. Specifically, the conformational profile of the N-acetyl-N’-methylamide derivative of the c6Phe stereoisomers exhibiting either a cis or a trans relative orientation between the amino and phenyl substituents has been analyzed in different environments (gas phase, chloroform and aqueous solutions). Calculations were performed using B3LYP, MP2 and HF methods combined with the 6-31+G(d,p) and 6-311++G(d,p) basis sets, and a self-consistent reaction-field (SCRF) method was applied to analyze the influence of the solvent. The amino acids investigated can be viewed as constrained phenylalanine analogues with a rigidly oriented aromatic side chain that may interact with the peptide backbone not only sterically but also electronically through the aromatic π orbitals. Their conformational propensities have been found to be strongly influenced by the specific orientation of the aromatic substituent in each stereoisomer and the conformation adopted by the cyclohexane ring, as well as by the environment. PMID:19772338

  18. Is the Conformational Ensemble of Alzheimer's Aβ10-40 Peptide Force Field Dependent?

    PubMed

    Siwy, Christopher M; Lockhart, Christopher; Klimov, Dmitri K

    2017-01-01

    By applying REMD simulations we have performed comparative analysis of the conformational ensembles of amino-truncated Aβ10-40 peptide produced with five force fields, which combine four protein parameterizations (CHARMM36, CHARMM22*, CHARMM22/cmap, and OPLS-AA) and two water models (standard and modified TIP3P). Aβ10-40 conformations were analyzed by computing secondary structure, backbone fluctuations, tertiary interactions, and radius of gyration. We have also calculated Aβ10-40 3JHNHα-coupling and RDC constants and compared them with their experimental counterparts obtained for the full-length Aβ1-40 peptide. Our study led us to several conclusions. First, all force fields predict that Aβ adopts unfolded structure dominated by turn and random coil conformations. Second, specific TIP3P water model does not dramatically affect secondary or tertiary Aβ10-40 structure, albeit standard TIP3P model favors slightly more compact states. Third, although the secondary structures observed in CHARMM36 and CHARMM22/cmap simulations are qualitatively similar, their tertiary interactions show little consistency. Fourth, two force fields, OPLS-AA and CHARMM22* have unique features setting them apart from CHARMM36 or CHARMM22/cmap. OPLS-AA reveals moderate β-structure propensity coupled with extensive, but weak long-range tertiary interactions leading to Aβ collapsed conformations. CHARMM22* exhibits moderate helix propensity and generates multiple exceptionally stable long- and short-range interactions. Our investigation suggests that among all force fields CHARMM22* differs the most from CHARMM36. Fifth, the analysis of 3JHNHα-coupling and RDC constants based on CHARMM36 force field with standard TIP3P model led us to an unexpected finding that in silico Aβ10-40 and experimental Aβ1-40 constants are generally in better agreement than these quantities computed and measured for identical peptides, such as Aβ1-40 or Aβ1-42. This observation suggests that the

  19. Is the Conformational Ensemble of Alzheimer’s Aβ10-40 Peptide Force Field Dependent?

    PubMed Central

    Siwy, Christopher M.

    2017-01-01

    By applying REMD simulations we have performed comparative analysis of the conformational ensembles of amino-truncated Aβ10-40 peptide produced with five force fields, which combine four protein parameterizations (CHARMM36, CHARMM22*, CHARMM22/cmap, and OPLS-AA) and two water models (standard and modified TIP3P). Aβ10-40 conformations were analyzed by computing secondary structure, backbone fluctuations, tertiary interactions, and radius of gyration. We have also calculated Aβ10-40 3JHNHα-coupling and RDC constants and compared them with their experimental counterparts obtained for the full-length Aβ1-40 peptide. Our study led us to several conclusions. First, all force fields predict that Aβ adopts unfolded structure dominated by turn and random coil conformations. Second, specific TIP3P water model does not dramatically affect secondary or tertiary Aβ10-40 structure, albeit standard TIP3P model favors slightly more compact states. Third, although the secondary structures observed in CHARMM36 and CHARMM22/cmap simulations are qualitatively similar, their tertiary interactions show little consistency. Fourth, two force fields, OPLS-AA and CHARMM22* have unique features setting them apart from CHARMM36 or CHARMM22/cmap. OPLS-AA reveals moderate β-structure propensity coupled with extensive, but weak long-range tertiary interactions leading to Aβ collapsed conformations. CHARMM22* exhibits moderate helix propensity and generates multiple exceptionally stable long- and short-range interactions. Our investigation suggests that among all force fields CHARMM22* differs the most from CHARMM36. Fifth, the analysis of 3JHNHα-coupling and RDC constants based on CHARMM36 force field with standard TIP3P model led us to an unexpected finding that in silico Aβ10-40 and experimental Aβ1-40 constants are generally in better agreement than these quantities computed and measured for identical peptides, such as Aβ1-40 or Aβ1-42. This observation suggests that the

  20. Syntaxin1a variants lacking an N-peptide or bearing the LE mutation bind to Munc18a in a closed conformation

    SciTech Connect

    Colbert, Karen N.; Hattendorf, Douglas A.; Weiss, Thomas M.; Burkhardt, Pawel; Fasshauer, Dirk; Weis, William I.

    2013-07-15

    In neurons, soluble N-ethylmaleimide–sensitive factor attachment receptor (SNARE) proteins drive the fusion of synaptic vesicles to the plasma membrane through the formation of a four-helix SNARE complex. Members of the Sec1/Munc18 protein family regulate membrane fusion through interactions with the syntaxin family of SNARE proteins. The neuronal protein Munc18a interacts with a closed conformation of the SNARE protein syntaxin1a (Syx1a) and with an assembled SNARE complex containing Syx1a in an open conformation. The N-peptide of Syx1a (amino acids 1–24) has been implicated in the transition of Munc18a-bound Syx1a to Munc18a-bound SNARE complex, but the underlying mechanism is not understood. In addition, we report the X-ray crystal structures of Munc18a bound to Syx1a with and without its native N-peptide (Syx1aΔN), along with small-angle X-ray scattering (SAXS) data for Munc18a bound to Syx1a, Syx1aΔN, and Syx1a L165A/E166A (LE), a mutation thought to render Syx1a in a constitutively open conformation. We show that all three complexes adopt the same global structure, in which Munc18a binds a closed conformation of Syx1a. We also identify a possible structural connection between the Syx1a N-peptide and SNARE domain that might be important for the transition of closed-to-open Syx1a in SNARE complex assembly. Although the role of the N-peptide in Munc18a-mediated SNARE complex assembly remains unclear, our results demonstrate that the N-peptide and LE mutation have no effect on the global conformation of the Munc18a–Syx1a complex.

  1. Syntaxin1a variants lacking an N-peptide or bearing the LE mutation bind to Munc18a in a closed conformation

    DOE PAGES

    Colbert, Karen N.; Hattendorf, Douglas A.; Weiss, Thomas M.; ...

    2013-07-15

    In neurons, soluble N-ethylmaleimide–sensitive factor attachment receptor (SNARE) proteins drive the fusion of synaptic vesicles to the plasma membrane through the formation of a four-helix SNARE complex. Members of the Sec1/Munc18 protein family regulate membrane fusion through interactions with the syntaxin family of SNARE proteins. The neuronal protein Munc18a interacts with a closed conformation of the SNARE protein syntaxin1a (Syx1a) and with an assembled SNARE complex containing Syx1a in an open conformation. The N-peptide of Syx1a (amino acids 1–24) has been implicated in the transition of Munc18a-bound Syx1a to Munc18a-bound SNARE complex, but the underlying mechanism is not understood. Inmore » addition, we report the X-ray crystal structures of Munc18a bound to Syx1a with and without its native N-peptide (Syx1aΔN), along with small-angle X-ray scattering (SAXS) data for Munc18a bound to Syx1a, Syx1aΔN, and Syx1a L165A/E166A (LE), a mutation thought to render Syx1a in a constitutively open conformation. We show that all three complexes adopt the same global structure, in which Munc18a binds a closed conformation of Syx1a. We also identify a possible structural connection between the Syx1a N-peptide and SNARE domain that might be important for the transition of closed-to-open Syx1a in SNARE complex assembly. Although the role of the N-peptide in Munc18a-mediated SNARE complex assembly remains unclear, our results demonstrate that the N-peptide and LE mutation have no effect on the global conformation of the Munc18a–Syx1a complex.« less

  2. Fmoc/Trt-amino acids: comparison to Fmoc/tBu-amino acids in peptide synthesis.

    PubMed

    Barlos, K; Gatos, D; Koutsogianni, S

    1998-03-01

    Model peptides containing the nucleophilic amino acids Trp and Met have been synthesized with the application of Fmoc/Trt- and Fmoc/tBu-amino acids, for comparison. The deprotection of the peptides synthesized using Fmoc/Trt-amino acids in all cases leads to crude peptides of higher purity than that of the same peptides synthesized using Fmoc/tBu-amino acids.

  3. Mimicking the membrane-mediated conformation of dynorphin A-(1-13)-peptide: Circular dichroism and nuclear magnetic resonance studies in methanolic solution

    SciTech Connect

    Lancaster, C.R.D.; Hughes, D.W.; Epand, R.M. ); Mishra, P.K.; Bothner-By, A.A. ); St.Pierre, S.A. )

    1991-05-14

    The structural requirements for the binding of dynorphin to the {kappa}-opioid receptor are of profound clinical interest in the search for a powerful nonaddictive analgesic. These requirements are thought to be met by the membrane-mediated conformation of the opioid peptide dynorphin A-(1-13)-peptide, Tyr{sup 1}-Gly{sup 2}-Gly{sup 3}-Phe{sup 4}-Leu{sup 5}-Arg{sup 6}-Arg{sup 7}-Ile{sup 8}-Arg{sup 9}-Pro{sup 10}-Lys{sup 11}-Leu{sup 12}-Lys{sup 13}. Schwyzer has proposed an essentially {alpha}-helical membrane-mediated conformation of the 13 amino acid peptide. In the present study, circular dichroism (CD) studies on dynorphin A-(1-13)-peptide bound to an anionic phospholipid signified negligible helical content of the peptide. CD studies also demonstrated that the aqueous-membraneous interphase may be mimicked by methanol. The 500- and 620-MHz {sup 1}H nuclear magnetic resonance (NMR) spectra of dynorphin A-(1-13)-peptide in methanolic solution were sequence-specifically assigned with the aid of correlated spectroscopy (COSY), double-quantum filtered phase-sensitive COSY (DQF-COSY), relayed COSY (RELAY), and nuclear Overhauser enhancement spectroscopy (NOESY). 2-D CAMELSPIN/ROESY experiments indicated that at least the part of the molecule from Arg{sup 7} to Arg{sup 9} was in an extended or {beta}-strand conformation, which agreed with deuterium-exchange and temperature-dependence studies of the amide protons and analysis of the vicinal spin-spin coupling constants {sup 3}J{sub HN{alpha}}. The results clearly demonstrated the absence of extensive {alpha}-helix formation. {chi}{sub 1} rotamer analysis of the {sup 3}J{sub {alpha}{beta}} demonstrated no preferred side-chain conformations.

  4. Conformational Flexibility and pH Effects on Anisotropic Growth of Sheet-Like Assembly of Amphiphilic Peptides.

    PubMed

    Huang, Hongzhou; Ganguly, Debabani; Chen, Jianhan; Sun, Xiuzhi S

    2015-06-01

    Peptide-based biomaterials have many potential applications in tissue engineering, drug delivery, surface engineering, and other areas. In this study, we exploited a series of amphiphilic diblock model peptides (L5K10, L5GSIIK10, and L5P(D)PK10) to understand how the supramolecular assembly morphology may be modulated by the physical properties of the peptide monomer and experimental conditions. A combination of experimentation and simulation revealed that although all three peptides lack stable structures as monomers, their levels of conformational heterogeneity differ significantly. Importantly, such differences appear to be correlated with the peptides' ability to form sheet-like assemblies. In particular, substantial conformational heterogeneity appears to be required for anisotropic growth of sheet-like materials, likely by reducing the peptide assembly kinetics. To test this hypothesis, we increased the pH to neutralize the lysine residues and promote peptide aggregation, and the resulting faster assembly rate hindered the growth of the sheet morphology as predicted. In addition, we designed and investigated the assembly morphologies of a series of diblock peptides with various lengths of polyglycine inserts, L5GxK10, x = 1, 2, 3, 4. The results further supported the importance of peptide conformational flexibility and pH in modulation of the peptide supramolecular assembly morphology.

  5. Conformational studies of a peptide corresponding to a region of the C-terminus of ribonuclease A: implications as a potential chain-folding initiation site.

    PubMed

    Beals, J M; Haas, E; Krausz, S; Scheraga, H A

    1991-08-06

    Conformational properties of the OT-16 peptide, the C-terminal 20 amino acids of RNase A, were examined by nonradiative energy transfer. A modified OT-16 peptide was prepared by solid-phase synthesis with the inclusion of diaminobutyric acid (DABA) at the C-terminus. The OT-16-DABA peptide was labeled with a fluorescent 1,5-dimethylaminonaphthalene sulfonyl (dansyl, DNS) acceptor at the N-terminal amine and a fluorescent naphthoxyacetic acid (NAA) donor at the gamma-amine of the DABA located at the C-terminus of the peptide by using an orthogonal protection scheme. Energy transfer was monitored in DNS-OT-16-DABA-NAA by using both fluorescence intensity (sensitized emission) and lifetime (donor quenching) experiments. The lifetime data indicate that the peptide system is a dynamic, flexible one. A detailed analysis, based on a dynamic model that includes a skewed Gaussian function to model the equilibrium distribution of interprobe distances and a mutual diffusion coefficient between the two probes to model conformational dynamics in the peptide [Beechem & Haas (1989) Biophys. J. 55, 1225.], identified the existence of a partially ordered structure (relatively narrow distribution of interprobe distances) at temperatures greater than or equal to 20 degrees C in the absence of denaturant. The width and the position of the average of the distributions decrease with increasing temperature, in this range; this suggests that the structure is stabilized by hydrophobic interactions. In addition, the peptide undergoes cold denaturation at around 1.5 degrees C as indicated by broadening of the distance distribution. The addition of 6 M guanidine hydrochloride (Gdn-HCl) also broadens the distance distribution significantly, presumably by eliminating the hydrophobic interactions and unfolding the peptide. The results of the analysis of the distance distribution demonstrate that (1) nonradiative energy transfer can be used to study the conformational dynamics of peptides on the

  6. Understanding the influence of guest-host interactions on the conformation of short peptides in a hydrophobic cavity: a computational study.

    PubMed

    Hua, Weijie; Xu, Lina; Luo, Yi; Li, Shuhua

    2011-05-09

    We performed a computational investigation to understand the conformational preferences of four short peptides in a self-assembled cage based on the experimental work by Y. Hatakeyama et al. (Angew. Chem. Int. Ed.2009, 48, 8695). For this purpose, we combined molecular dynamics simulations, Monte Carlo simulations, and quantum mechanical calculations to obtain energies and structures for several low-lying conformers of four peptides and the corresponding peptide-cage inclusion complexes. Our calculations at both B3LYP and MP2 levels show that for each peptide, the corresponding conformation within the host (as revealed by the crystal structure) does not represent the lowest-energy conformation of this peptide in vacuum. By comparing some low-lying conformers in vacuum and in the cavity (for the same peptide), we found that the cage has a significant influence on the conformational propensities of peptides. First, one carbonyl oxygen of each peptide tends to bind to one Zn(II) atom of the cage, forming a Zn-O bond. The formation of this bond leads to significant charge transfer from the cage to the peptide. Second, this Zn-O bond causes the peptide to go through some local conformational changes. For larger peptides, such as penta- and hexapeptides, our calculations also show that some of their conformers must undergo significant structural changes, due to the confinement of the host. This computational study reveals the noticeable influence of the guest-host interaction on the conformational preferences of short peptides.

  7. Conformation and intermolecular interactions of SA2 peptides self-assembled into vesicles.

    PubMed

    van Hell, Albert J; Klymchenko, Andrey; Burgers, Pepijn P; Moret, Ed E; Jiskoot, Wim; Hennink, Wim E; Crommelin, Daan J A; Mastrobattista, Enrico

    2010-09-02

    Previously we have shown that the recombinantly produced SA2 amphiphilic oligopeptide (Ac-Ala-Ala-Val-Val-Leu-Leu-Leu-Trp-Glu-Glu-COOH) self-assembles into nanovesicles (van Hell et al. 2007). In this study, the intermolecular interactions that contribute to the formation of such peptide vesicles are examined. First, analysis of a 3-hydroxyflavone fluorescent probe inserted into the peptide assemblies demonstrated that the peptide self-assembly is based on hydrophobic clustering. The polarity of this hydrophobic microenvironment was comparable to that of negatively charged lipid bilayers. A substantial level of hydration at the hydrophilic-hydrophobic interface was detected, as was further confirmed by tryptophan fluorescence analysis. However, organic solvents such as acetonitrile, tetrahydrofuran, or ethanol could not disrupt SA2 oligopeptide vesicles, whereas these solvents fully disintegrated lipid vesicles. Instead, the SA2 assembly immediately disintegrated in hydrogen breaking solvents such dimethylsulfoxide and dimethylformamide, suggesting the involvement of additional intermolecular interactions via hydrogen bonding. Circular dichroism and Fourier transform infrared spectroscopy excluded well-defined patterns of intramolecular hydrogen bonding and indicated the polyproline type II as the dominant SA2 peptide conformation, which enables intermolecular hydrogen bonding. All-atom computational simulations were used to confirm the presence of such intermolecular hydrogen bonds and degrees of hydration. On the basis of the experimental and computational data presented, we propose a model of an interdigitated peptide assembly that involves intermolecular hydrogen bonding in addition to hydrophobic interactions that stabilize SA2 oligopeptide vesicles.

  8. Disaggregation of Amylin Aggregate by Novel Conformationally Restricted Aminobenzoic Acid containing α/β and α/γ Hybrid Peptidomimetics

    NASA Astrophysics Data System (ADS)

    Paul, Ashim; Kalita, Sourav; Kalita, Sujan; Sukumar, Piruthivi; Mandal, Bhubaneswar

    2017-01-01

    Diabetes has emerged as a threat to the current world. More than ninety five per cent of all the diabetic population has type 2 diabetes mellitus (T2DM). Aggregates of Amylin hormone, which is co-secreted with insulin from the pancreatic β-cells, inhibit the activities of insulin and glucagon and cause T2DM. Importance of the conformationally restricted peptides for drug design against T2DM has been invigorated by recent FDA approval of Symlin, which is a large conformationally restricted peptide. However, Symlin still has some issues including solubility, oral bioavailability and cost of preparation. Herein, we introduced a novel strategy for conformationally restricted peptide design adopting a minimalistic approach for cost reduction. We have demonstrated efficient inhibition of amyloid formation of Amylin and its disruption by a novel class of conformationally restricted β-sheet breaker hybrid peptidomimetics (BSBHps). We have inserted β, γ and δ -aminobenzoic acid separately into an amyloidogenic peptide sequence, synthesized α/β, α/γ and α/δ hybrid peptidomimetics, respectively. Interestingly, we observed the aggregation inhibitory efficacy of α/β and α/γ BSBHps, but not of α/δ analogues. They also disrupt existing amyloids into non-toxic forms. Results may be useful for newer drug design against T2DM as well as other amyloidoses and understanding amyloidogenesis.

  9. Disaggregation of Amylin Aggregate by Novel Conformationally Restricted Aminobenzoic Acid containing α/β and α/γ Hybrid Peptidomimetics

    PubMed Central

    Paul, Ashim; Kalita, Sourav; Kalita, Sujan; Sukumar, Piruthivi; Mandal, Bhubaneswar

    2017-01-01

    Diabetes has emerged as a threat to the current world. More than ninety five per cent of all the diabetic population has type 2 diabetes mellitus (T2DM). Aggregates of Amylin hormone, which is co-secreted with insulin from the pancreatic β-cells, inhibit the activities of insulin and glucagon and cause T2DM. Importance of the conformationally restricted peptides for drug design against T2DM has been invigorated by recent FDA approval of Symlin, which is a large conformationally restricted peptide. However, Symlin still has some issues including solubility, oral bioavailability and cost of preparation. Herein, we introduced a novel strategy for conformationally restricted peptide design adopting a minimalistic approach for cost reduction. We have demonstrated efficient inhibition of amyloid formation of Amylin and its disruption by a novel class of conformationally restricted β-sheet breaker hybrid peptidomimetics (BSBHps). We have inserted β, γ and δ -aminobenzoic acid separately into an amyloidogenic peptide sequence, synthesized α/β, α/γ and α/δ hybrid peptidomimetics, respectively. Interestingly, we observed the aggregation inhibitory efficacy of α/β and α/γ BSBHps, but not of α/δ analogues. They also disrupt existing amyloids into non-toxic forms. Results may be useful for newer drug design against T2DM as well as other amyloidoses and understanding amyloidogenesis. PMID:28054630

  10. Conformational Fine-Tuning of Pore-Forming Peptide Potency and Selectivity

    PubMed Central

    2015-01-01

    To better understand the sequence–structure–function relationships that control the activity and selectivity of membrane-permeabilizing peptides, we screened a peptide library, based on the archetypal pore-former melittin, for loss-of-function variants. This was accomplished by assaying library members for failure to cause leakage of entrapped contents from synthetic lipid vesicles at a peptide-to-lipid ratio of 1:20, 10-fold higher than the concentration at which melittin efficiently permeabilizes the same vesicles. Surprisingly, about one-third of the library members are inactive under these conditions. In the negative peptides, two changes of hydrophobic residues to glycine were especially abundant. We show that loss-of-function activity can be completely recapitulated by a single-residue change of the leucine at position 16 to glycine. Unlike the potently cytolytic melittin, the loss-of-function peptides, including the single-site variant, are essentially inactive against phosphatidylcholine vesicles and multiple types of eukaryotic cells. Loss of function is shown to result from a shift in the binding–folding equilibrium away from the active, bound, α-helical state toward the inactive, unbound, random-coil state. Accordingly, the addition of anionic lipids to synthetic lipid vesicles restored binding, α-helical secondary structure, and potent activity of the “negative” peptides. While nontoxic to mammalian cells, the single-site variant has potent bactericidal activity, consistent with the anionic nature of bacterial membranes. The results show that conformational fine-tuning of helical pore-forming peptides is a powerful way to modulate their activity and selectivity. PMID:26632653

  11. Assessment of density functionals with long-range and/or empirical dispersion corrections for conformational energy calculations of peptides.

    PubMed

    Kang, Young Kee; Byun, Byung Jin

    2010-12-01

    Density functionals with long-range and/or empirical dispersion corrections, including LC-ωPBE, B97-D, ωB97X-D, M06-2X, B2PLYP-D, and mPW2PLYP-D functionals, are assessed for their ability to describe the conformational preferences of Ac-Ala-NHMe (the alanine dipeptide) and Ac-Pro-NHMe (the proline dipeptide) in the gas phase and in water, which have been used as prototypes for amino acid residues of peptides. For both dipeptides, the mean absolute deviation (MAD) is estimated to be 0.22-0.40 kcal/mol in conformational energy and 2.0-3.2° in torsion angles φ and ψ using these functionals with the 6-311++G(d,p) basis set against the reference values calculated at the MP2/aug-cc-pVTZ//MP2/aug-cc-pVDZ level of theory in the gas phase. The overall performance is obtained in the order B2PLYP-D ≈ mPW2PLYP-D > ωB97X-D ≈ M06-2X > MP2 > LC-ωPBE > B3LYP with the 6-311++G(d,p) basis set. The SMD model at the M06-2X/6-31+G(d) level of theory well reproduced experimental hydration free energies of the model compounds for backbone and side chains of peptides with MADs of 0.47 and 4.3 kcal/mol for 20 neutral and 5 charged molecules, respectively. The B2PLYP-D/6-311++G(d,p)//SMD M06-2X/6-31+G(d) level of theory provides the populations of backbone and/or prolyl peptide bond for the alanine and proline dipeptides in water that are consistent with the observed values.

  12. Synthesis and conformational analysis of hybrid α/β-dipeptides incorporating S-glycosyl-β(2,2)-amino acids.

    PubMed

    García-González, Iván; Mata, Lara; Corzana, Francisco; Jiménez-Osés, Gonzalo; Avenoza, Alberto; Busto, Jesús H; Peregrina, Jesús M

    2015-01-12

    We synthesized and carried out the conformational analysis of several hybrid dipeptides consisting of an α-amino acid attached to a quaternary glyco-β-amino acid. In particular, we combined a S-glycosylated β(2,2)-amino acid and two different types of α-amino acid, namely, aliphatic (alanine) and aromatic (phenylalanine and tryptophan) in the sequence of hybrid α/β-dipeptides. The key step in the synthesis involved the ring-opening reaction of a chiral cyclic sulfamidate, inserted in the peptidic sequence, with a sulfur-containing nucleophile by using 1-thio-β-D-glucopyranose derivatives. This reaction of glycosylation occurred with inversion of configuration at the quaternary center. The conformational behavior in aqueous solution of the peptide backbone and the glycosidic linkage for all synthesized hybrid glycopeptides was analyzed by using a protocol that combined NMR experiments and molecular dynamics with time-averaged restraints (MD-tar). Interestingly, the presence of the sulfur heteroatom at the quaternary center of the β-amino acid induced θ torsional angles close to 180° (anti). Notably, this value changed to 60° (gauche) when the peptidic sequence displayed aromatic α-amino acids due to the presence of CH-π interactions between the phenyl or indole ring and the methyl groups of the β-amino acid unit. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Effect of osmolytes on the conformation and aggregation of some amyloid peptides: CD spectroscopic data.

    PubMed

    Inayathullah, Mohammed; Rajadas, Jayakumar

    2016-06-01

    Protein misfolding and aggregation are responsible for a large number of diseases called protein conformational diseases or disorders that include Alzheimer׳s disease, Huntington׳s diseases, Prion related encephalopathies and type-II diabetes (http://dx.doi.org/10.1038/35041139) (Kopito and Ron, 2000) [1]. A variety of studies have shown that some small organic molecules, known as osmolytes have the ability to stabilize native conformation of proteins and prevent misfolding and aggregation (http://www.la-press.com/article.php?article_id=447) (Zhao et al., 2008) [2]. It has been shown that certain short segment or fragment of respective proteins can also form amyloids, and the segments also promote the aggregation in the full-length protein (http://dx.doi.org/10.2174/0929867023369187) (Gazit, 2002) [3]. This article presents circular dichroism spectroscopic data on conformational analysis and effect of osmolytes on Aβ peptide fragments, different lengths of polyglutamine peptide and the amyloidogenic segment of islet amyloid polypeptide.

  14. Peptide and amino acid separation with nanofiltration membranes

    SciTech Connect

    Tsuru, Toshinori; Shutou, Takatoshi; Nakao, Shin-Ichi; Kimura, Shoji )

    1994-05-01

    Several nanofiltration membranes [UTC-20, 60 (Toray Industries), NF-40 (Film-Tech Corporation), Desal-5, G-20 (Desalination Systems), and NTR-7450 (Nitto Electric Industrial Co.)] were applied to separate amino acids and peptides on the basis of charge interaction with the membranes since most of them contain charged functional groups. Nanofiltration membranes having a molecular weight cutoff (MWCO) below 300 (UTC-20, 60, NF-40 and Desal-5) were not suitable for separation of amino acids. On the other hand, separation of amino acids and peptides with nanofiltration membranes having a MWCO around 2000-3000 (NTR-7450 and G-20) was satisfactory based on a charge effect mechanism; charged amino acids and peptides were rejected while neutral amino acids and peptides permeated through the membranes. Separation of peptides having different isoelectric points with nanofiltration membranes was possible by adjusting the pH. 15 refs., 11 figs., 4 tabs.

  15. Beta-hairpin conformation of fibrillogenic peptides: structure and alpha-beta transition mechanism revealed by molecular dynamics simulations.

    PubMed

    Daidone, Isabella; Simona, Fabio; Roccatano, Danilo; Broglia, Ricardo A; Tiana, Guido; Colombo, Giorgio; Di Nola, Alfredo

    2004-10-01

    Understanding the conformational transitions that trigger the aggregation and amyloidogenesis of otherwise soluble peptides at atomic resolution is of fundamental relevance for the design of effective therapeutic agents against amyloid-related disorders. In the present study the transition from ideal alpha-helical to beta-hairpin conformations is revealed by long timescale molecular dynamics simulations in explicit water solvent, for two well-known amyloidogenic peptides: the H1 peptide from prion protein and the Abeta(12-28) fragment from the Abeta(1-42) peptide responsible for Alzheimer's disease. The simulations highlight the unfolding of alpha-helices, followed by the formation of bent conformations and a final convergence to ordered in register beta-hairpin conformations. The beta-hairpins observed, despite different sequences, exhibit a common dynamic behavior and the presence of a peculiar pattern of the hydrophobic side-chains, in particular in the region of the turns. These observations hint at a possible common aggregation mechanism for the onset of different amyloid diseases and a common mechanism in the transition to the beta-hairpin structures. Furthermore the simulations presented herein evidence the stabilization of the alpha-helical conformations induced by the presence of an organic fluorinated cosolvent. The results of MD simulation in 2,2,2-trifluoroethanol (TFE)/water mixture provide further evidence that the peptide coating effect of TFE molecules is responsible for the stabilization of the soluble helical conformation.

  16. The interaction with gold suppresses fiber-like conformations of the amyloid β (16-22) peptide

    NASA Astrophysics Data System (ADS)

    Bellucci, Luca; Ardèvol, Albert; Parrinello, Michele; Lutz, Helmut; Lu, Hao; Weidner, Tobias; Corni, Stefano

    2016-04-01

    Inorganic surfaces and nanoparticles can accelerate or inhibit the fibrillation process of proteins and peptides, including the biomedically relevant amyloid β peptide. However, the microscopic mechanisms that determine such an effect are still poorly understood. By means of large-scale, state-of-the-art enhanced sampling molecular dynamics simulations, here we identify an interaction mechanism between the segments 16-22 of the amyloid β peptide, known to be fibrillogenic by itself, and the Au(111) surface in water that leads to the suppression of fiber-like conformations from the peptide conformational ensemble. Moreover, thanks to advanced simulation analysis techniques, we characterize the conformational selection vs. induced fit nature of the gold effect. Our results disclose an inhibition mechanism that is rooted in the details of the microscopic peptide-surface interaction rather than in general phenomena such as peptide sequestration from the solution.Inorganic surfaces and nanoparticles can accelerate or inhibit the fibrillation process of proteins and peptides, including the biomedically relevant amyloid β peptide. However, the microscopic mechanisms that determine such an effect are still poorly understood. By means of large-scale, state-of-the-art enhanced sampling molecular dynamics simulations, here we identify an interaction mechanism between the segments 16-22 of the amyloid β peptide, known to be fibrillogenic by itself, and the Au(111) surface in water that leads to the suppression of fiber-like conformations from the peptide conformational ensemble. Moreover, thanks to advanced simulation analysis techniques, we characterize the conformational selection vs. induced fit nature of the gold effect. Our results disclose an inhibition mechanism that is rooted in the details of the microscopic peptide-surface interaction rather than in general phenomena such as peptide sequestration from the solution. Electronic supplementary information (ESI

  17. BIOACTIVE PROTEINS, PEPTIDES, AND AMINO ACIDS FROM MACROALGAE(1).

    PubMed

    Harnedy, Pádraigín A; FitzGerald, Richard J

    2011-04-01

    Macroalgae are a diverse group of marine organisms that have developed complex and unique metabolic pathways to ensure survival in highly competitive marine environments. As a result, these organisms have been targeted for mining of natural biologically active components. The exploration of marine organisms has revealed numerous bioactive compounds that are proteinaceous in nature. These include proteins, linear peptides, cyclic peptides and depsipeptides, peptide derivatives, amino acids, and amino acid-like components. Furthermore, some species of macroalgae have been shown to contain significant levels of protein. While some protein-derived bioactive peptides have been characterized from macroalgae, macroalgal proteins currently still represent good candidate raw materials for biofunctional peptide mining. This review will provide an overview of the important bioactive amino-acid-containing compounds that have been identified in macroalgae. Moreover, the potential of macroalgal proteins as substrates for the generation of biofunctional peptides for utilization as functional foods to provide specific health benefits will be discussed.

  18. UV Resonance Raman Elucidation of the Terminal and Internal Peptide Bond Conformations of Crystalline and Solution Oligoglycines

    PubMed Central

    Bykov, Sergei V.; Asher, Sanford A.

    2010-01-01

    Spectroscopic investigations of macromolecules generally attempt to interpret the measured spectra in terms of the summed contributions of the different molecular fragments. This is the basis of the local mode approximation in vibrational spectroscopy. In the case of resonance Raman spectroscopy independent contributions of molecular fragments require both a local mode-like behavior and the uncoupled electronic transitions. Here we show that the deep UV resonance Raman spectra of aqueous solution phase oligoglycines show independent peptide bond molecular fragment contributions indicating that peptide bonds electronic transitions and vibrational modes are uncoupled. We utilize this result to separately determine the conformational distributions of the internal and penultimate peptide bonds of oligoglycines. Our data indicate that in aqueous solution the oligoglycine terminal residues populate conformations similar to those found in crystals (31-helices and β-strands), but with a broader distribution, while the internal peptide bond conformations are centered around the 31-helix Ramachandran angles. PMID:20657703

  19. An exhaustive survey of regular peptide conformations using a new metric for backbone handedness ( h )

    DOE PAGES

    Mannige, Ranjan V.

    2017-05-16

    The Ramachandran plot is important to structural biology as it describes a peptide backbone in the context of its dominant degrees of freedom—the backbone dihedral anglesφandψ(Ramachandran, Ramakrishnan & Sasisekharan, 1963). Since its introduction, the Ramachandran plot has been a crucial tool to characterize protein backbone features. However, the conformation or twist of a backbone as a function ofφandψhas not been completely described for bothcisandtransbackbones. Additionally, little intuitive understanding is available about a peptide’s conformation simply from knowing theφandψvalues of a peptide (e.g., is the regular peptide defined byφ = ψ =  - 100°  left-handed or right-handed?). This report provides a new metric for backbone handednessmore » (h) based on interpreting a peptide backbone as a helix with axial displacementdand angular displacementθ, both of which are derived from a peptide backbone’s internal coordinates, especially dihedral anglesφ,ψandω. In particular,hequals sin(θ)d/d|, with range [-1, 1] and negative (or positive) values indicating left(or right)-handedness. The metrichis used to characterize the handedness of every region of the Ramachandran plot for bothcis(ω = 0°) and trans (ω = 180°) backbones, which provides the first exhaustive survey of twist handedness in Ramachandran (φ,ψ) space. These maps fill in the ‘dead space’ within the Ramachandran plot, which are regions that are not commonly accessed by structured proteins, but which may be accessible to intrinsically disordered proteins, short peptide fragments, and protein mimics such as peptoids. Finally, building on the work of (Zacharias & Knapp, 2013), this report presents a new plot based ondandθthat serves as a universal and intuitive alternative to the Ramachandran plot. The universality arises from the fact that the co-inhabitants of such a plot include every possible peptide backbone includingcisandtransbackbones. The intuitiveness

  20. An exhaustive survey of regular peptide conformations using a new metric for backbone handedness (h)

    PubMed Central

    2017-01-01

    The Ramachandran plot is important to structural biology as it describes a peptide backbone in the context of its dominant degrees of freedom—the backbone dihedral angles φ and ψ (Ramachandran, Ramakrishnan & Sasisekharan, 1963). Since its introduction, the Ramachandran plot has been a crucial tool to characterize protein backbone features. However, the conformation or twist of a backbone as a function of φ and ψ has not been completely described for both cis and trans backbones. Additionally, little intuitive understanding is available about a peptide’s conformation simply from knowing the φ and ψ values of a peptide (e.g., is the regular peptide defined by φ = ψ =  − 100°  left-handed or right-handed?). This report provides a new metric for backbone handedness (h) based on interpreting a peptide backbone as a helix with axial displacement d and angular displacement θ, both of which are derived from a peptide backbone’s internal coordinates, especially dihedral angles φ, ψ and ω. In particular, h equals sin(θ)d∕|d|, with range [−1, 1] and negative (or positive) values indicating left(or right)-handedness. The metric h is used to characterize the handedness of every region of the Ramachandran plot for both cis (ω = 0°) and trans (ω = 180°) backbones, which provides the first exhaustive survey of twist handedness in Ramachandran (φ, ψ) space. These maps fill in the ‘dead space’ within the Ramachandran plot, which are regions that are not commonly accessed by structured proteins, but which may be accessible to intrinsically disordered proteins, short peptide fragments, and protein mimics such as peptoids. Finally, building on the work of (Zacharias & Knapp, 2013), this report presents a new plot based on d and θ that serves as a universal and intuitive alternative to the Ramachandran plot. The universality arises from the fact that the co-inhabitants of such a plot include every possible peptide backbone including cis

  1. Conformational properties of oxazole-amino acids: effect of the intramolecular N-H···N hydrogen bond.

    PubMed

    Siodłak, Dawid; Staś, Monika; Broda, Małgorzata A; Bujak, Maciej; Lis, Tadeusz

    2014-03-06

    Oxazole ring occurs in numerous natural peptides, but conformational properties of the amino acid residue containing the oxazole ring in place of the C-terminal amide bond are poorly recognized. A series of model compounds constituted by the oxazole-amino acids occurring in nature, that is, oxazole-alanine (L-Ala-Ozl), oxazole-dehydroalanine (ΔAla-Ozl), and oxazole-dehydrobutyrine ((Z)-ΔAbu-Ozl), was investigated using theoretical calculations supported by FTIR and NMR spectra and single-crystal X-ray diffraction. It was found that the main feature of the studied oxazole-amino acids is the stable conformation β2 with the torsion angles φ and ψ of -150°, -10° for L-Ala-Ozl, -180°, 0° for ΔAla-Ozl, and -120°, 0° for (Z)-ΔAbu-Ozl, respectively. The conformation β2 is stabilized by the intramolecular N-H···N hydrogen bond and predominates in the low polar environment. In the case of the oxazole-dehydroamino acids, the π-electron conjugation that is spread on the oxazole ring and C(α)═C(β) double bond is an additional stabilizing interaction. The tendency to adopt the conformation β2 clearly decreases with increasing the polarity of environment, but still the oxazole-dehydroamino acids are considered to be more rigid and resistant to conformational changes.

  2. Charge, Color, and Conformation: Spectroscopy on Isomer-Selected Peptide Ions.

    PubMed

    Choi, Chang Min; Simon, Anne-Laure; Chirot, Fabien; Kulesza, Alexander; Knight, Geoffrey; Daly, Steven; MacAleese, Luke; Antoine, Rodolphe; Dugourd, Philippe

    2016-02-04

    Monitoring the chromism induced by intramolecular hydrogen and charge transfers within proteins as well as the isomerization of both protein and cofactor is essential not only to understand photoactive signaling pathways but also to design targeted opto-switchable proteins. We used a dual-ion mobility drift tube coupled to a tunable picosecond laser to explore the optical and structural properties of a peptide chain bound to a chromophore-a prototype system allowing for a proton transfer coupled to conformational change. With the support of molecular dynamics and DFT calculations, we show how proton transfer between the peptide and its cofactor can dramatically modify the optical properties of the system and demonstrate that these changes can be triggered by collisional activation in the gas phase.

  3. Peptide Conformational Preferences in Osmolyte Solutions: Transfer Free Energies of Decaalanine

    SciTech Connect

    Kokubo, Hironori; Hu, Char Y.; Pettitt, Bernard M.

    2011-02-16

    The nature in which the protecting osmolyte trimethylamine N-oxide (TMAO) and the denaturing osmolyte urea affect protein stability is investigated, simulating a decaalanine peptide model in multiple conformations of the denatured ensemble. Binary solutions of both osmolytes and mixed osmolyte solutions at physiologically relevant concentrations of 2:1 (urea:TMAO) are studied using standard molecular dynamics simulations and solvation free energy calculations. Component analysis reveals the differences in the importance of the van der Waals (vdW) and electrostatic interactions for protecting and denaturing osmolytes. We find that urea denaturation governed by transfer free energy differences is dominated by vdW attractions, whereas TMAO exerts its effect by causing unfavorable electrostatic interactions both in the binary solution and mixed osmolyte solution. Analysis of the results showed no evidence in the ternary solution of disruption of the correlations among the peptide and osmolytes, nor of significant changes in the strength of the water hydrogen bond network.

  4. Charge, Color, and Conformation: Spectroscopy on Isomer-Selected Peptide Ions

    PubMed Central

    2016-01-01

    Monitoring the chromism induced by intramolecular hydrogen and charge transfers within proteins as well as the isomerization of both protein and cofactor is essential not only to understand photoactive signaling pathways but also to design targeted opto-switchable proteins. We used a dual-ion mobility drift tube coupled to a tunable picosecond laser to explore the optical and structural properties of a peptide chain bound to a chromophore—a prototype system allowing for a proton transfer coupled to conformational change. With the support of molecular dynamics and DFT calculations, we show how proton transfer between the peptide and its cofactor can dramatically modify the optical properties of the system and demonstrate that these changes can be triggered by collisional activation in the gas phase. PMID:26756462

  5. FTIR Studies of Collagen Model Peptides: Complementary Experimental and Simulation Approaches to Conformation and Unfolding

    PubMed Central

    Bryan, Michael A.; Brauner, Joseph W.; Anderle, Gloria; Flach, Carol R.; Brodsky, Barbara; Mendelsohn, Richard

    2008-01-01

    X-ray crystallography of collagen model peptides has provided high resolution structures of the basic triple-helical conformation and its water-mediated hydration network. Vibrational spectroscopy provides a useful bridge for transferring the structural information from x-ray diffraction to collagen in its native environment. The vibrational mode most useful for this purpose is the Amide I mode (mostly peptide bond C=O stretch) near 1650 cm−1. The current study refines and extends the range of utility of a novel simulation method that accurately predicts the IR Amide I spectral contour from the three dimensional structure of a protein or peptide. The approach is demonstrated through accurate simulation of the experimental Amide I contour in solution for both a standard triple-helix, (Pro-Pro-Gly)10, and a second peptide with a Gly → Ala substitution in the middle of the chain that models the effect of a mutation in the native collagen sequence. Monitoring the major Amide I peak as a function of temperature gives sharp thermal transitions for both peptides, similar to those obtained by circular dichroism spectroscopy, and the FTIR spectra of the unfolded states were compared with polyproline II. The simulation studies were extended to model early stages of thermal denaturation of (Pro-Pro-Gly)10. Dihedral angle changes suggested by molecular dynamics simulations were made in a stepwise fashion to generate peptide unwinding from each end, which emulates the effect of increasing temperature. Simulated bands from these new structures were then compared to the experimental bands obtained as temperature was increased. The similarity between the simulated and experimental IR spectra lends credence to the simulation method, and paves the way for a variety of applications. PMID:17550251

  6. Conformational dynamics and aggregation behavior of piezoelectric diphenylalanine peptides in an external electric field.

    PubMed

    Kelly, Catherine M; Northey, Thomas; Ryan, Kate; Brooks, Bernard R; Kholkin, Andrei L; Rodriguez, Brian J; Buchete, Nicolae-Viorel

    2015-01-01

    Aromatic peptides including diphenylalanine (FF) have the capacity to self-assemble into ordered, biocompatible nanostructures with piezoelectric properties relevant to a variety of biomedical applications. Electric fields are commonly applied to align FF nanotubes, yet little is known about the effect of the electric field on the assembly process. Using all-atom molecular dynamics with explicit water molecules, we examine the response of FF monomers to the application of a constant external electric field over a range of intensities. We probe the aggregation mechanism of FF peptides, and find that the presence of even relatively weak fields can accelerate ordered aggregation, primarily by facilitating the alignment of individual molecular dipole moments. This is modulated by the conformational response of individual FF peptides (e.g., backbone stretching) and by the cooperative alignment of neighboring FF and water molecules. These observations may facilitate future studies on the controlled formation of nanostructured aggregates of piezoelectric peptides and the understanding of their electro-mechanical properties. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. The role of metals in protein conformational disorders - The case of prion protein and Aβ -peptide

    NASA Astrophysics Data System (ADS)

    De Santis, E.; Minicozzi, V.; Morante, S.; Rossi, G. C.; Stellato, F.

    2016-02-01

    Protein conformational disorders are members of a vast class of pathologies in which endogenous proteins or peptides undergo a misfolding process by switching from the physiological soluble configuration to a pathological fibrillar insoluble state. An important, but not yet fully elucidated, role in the process appears to be played by transition metal ions, mainly copper and zinc. X-ray absorption spectroscopy is one of the most suitable techniques for the structural characterization of biological molecules in complex with metal. Owing to its chemical selectivity and sensitivity to the local atomic geometry around the absorber, it can be successfully used to study the environment of metal ions in complex with proteins and peptides in physiological conditions. In this paper we present X-ray absorption spectroscopy studies of the metal ions coordination modes in systems where metals are complexed with specific amyloidogenic proteins and peptides. In particular, we show results concerning the Amyloid β peptide, that is involved in Alzheimer's disease, and the Prion protein, that is responsible for the Transmissible Spongiform Encephalopathy. Our findings suggest that the copper and zinc ions may play a crucial role in the aggregation and fibril formation process of these two biomolecules. Elucidating this kind of interaction could be a key preliminary step before any viable therapy can be conceived or designed.

  8. Site-Specific Characterization of d-Amino Acid Containing Peptide Epimers by Ion Mobility Spectrometry

    PubMed Central

    2013-01-01

    Traditionally, the d-amino acid containing peptide (DAACP) candidate can be discovered by observing the differences of biological activity and chromatographic retention time between the synthetic peptides and naturally occurring peptides. However, it is difficult to determine the exact position of d-amino acid in the DAACP candidates. Herein, we developed a novel site-specific strategy to rapidly and precisely localize d-amino acids in peptides by ion mobility spectrometry (IMS) analysis of mass spectrometry (MS)-generated epimeric fragment ions. Briefly, the d/l-peptide epimers were separated by online reversed-phase liquid chromatography and fragmented by collision-induced dissociation (CID), followed by IMS analysis. The epimeric fragment ions resulting from d/l-peptide epimers exhibit conformational differences, thus showing different mobilities in IMS. The arrival time shift between the epimeric fragment ions was used as criteria to localize the d-amino acid substitution. The utility of this strategy was demonstrated by analysis of peptide epimers with different molecular sizes, [d-Trp]-melanocyte-stimulating hormone, [d-Ala]-deltorphin, [d-Phe]-achatin-I, and their counterparts that contain all-l amino acids. Furthermore, the crustacean hyperglycemia hormones (CHHs, 8.5 kDa) were isolated from the American lobster Homarus americanus and identified by integration of MS-based bottom-up and top-down sequencing approaches. The IMS data acquired using our novel site-specific strategy localized the site of isomerization of l- to d-Phe at the third residue of the CHHs from the N-terminus. Collectively, this study demonstrates a new method for discovery of DAACPs using IMS technique with the ability to localize d-amino acid residues. PMID:24328107

  9. On the conformational memory in the photodissociation of formic acid.

    PubMed

    Martínez-Núñez, E; Vazquez, S A; Borges, I; Rocha, A B; Estévez, C M; Castillo, J F; Aoiz, F J

    2005-03-31

    The photodissociation of formic acid at 248 and 193 nm was investigated by classical trajectory and RRKM calculations using an interpolated potential energy surface, iteratively constructed using the B3LYP/aug-cc-pVDZ level of calculation. Several sampling schemes in the ground electronic state were employed to explore the possibility of conformational memory in formic acid. The CO/CO2 branching ratios obtained from trajectories initiated at the cis and at the trans conformers are almost identical to each other and in very good accordance with the RRKM results. In addition, when a specific initial excitation that simulates more rigorously the internal conversion process is used, the calculated branching ratio does not vary with respect to those obtained from cis and trans initializations. This result is at odds with the idea of conformational memory in the ground state proposed recently for the interpretation of the experimental results. It was also found that the calculated CO vibrational distributions after dissociation of the parent molecule at 248 nm are in agreement with the experimental available data.

  10. Fatty acid conjugation enhances the activities of antimicrobial peptides.

    PubMed

    Li, Zhining; Yuan, Penghui; Xing, Meng; He, Zhumei; Dong, Chuanfu; Cao, Yongchang; Liu, Qiuyun

    2013-04-01

    Antimicrobial peptides are small molecules that play a crucial role in innate immunity in multi-cellular organisms, and usually expressed and secreted constantly at basal levels to prevent infection, but local production can be augmented upon an infection. The clock is ticking as rising antibiotic abuse has led to the emergence of many drug resistance bacteria. Due to their broad spectrum antibiotic and antifungal activities as well as anti-viral and anti-tumor activities, efforts are being made to develop antimicrobial peptides into future microbial agents. This article describes some of the recent patents on antimicrobial peptides with fatty acid conjugation. Potency and selectivity of antimicrobial peptide can be modulated with fatty acid tails of variable length. Interaction between membranes and antimicrobial peptides was affected by fatty acid conjugation. At concentrations above the critical miscelle concentration (CMC), propensity of solution selfassembly hampered binding of the peptide to cell membranes. Overall, fatty acid conjugation has enhanced the activities of antimicrobial peptides, and occasionally it rendered inactive antimicrobial peptides to be bioactive. Antimicrobial peptides can not only be used as medicine but also as food additives.

  11. Contribution of the empirical dispersion correction on the conformation of short alanine peptides obtained by gas-phase QM calculations.

    PubMed

    Fadda, Elisa; Woods, Robert J

    2013-09-01

    In this work we analyze the effect of the inclusion of an empirical dispersion term to standard DFT (DFT-D) in the prediction of the conformational energy of the alanine dipeptide (Ala2) and in assessing the relative stabilities of short polyala-nine peptides in helical conformations, i.e., α and 310 helices, from Ala4 to Ala16. The Ala2 conformational energies obtained with the dispersion-corrected GGA functional B97-D are compared to previously published high level MP2 data. Meanwhile, the B97-D performance on larger polyalanine peptides is compared to MP2, B3LYP and RHF calculations obtained at a lower level of theory. Our results show that electron correlation affects the conformational energies of short peptides with a weight that increases with the peptide length. Indeed, while the contribution of vdW forces is significant for larger peptides, in the case of Ala2 it is negligible when compared to solvent effects. Even for short peptides, the inclusion of an empirical dispersion term greatly improves accuracy of DFT methods, providing results that correlate very well with the MP2 reference at no additional computational cost.

  12. Contribution of the empirical dispersion correction on the conformation of short alanine peptides obtained by gas-phase QM calculations

    PubMed Central

    Fadda, Elisa; Woods, Robert J.

    2014-01-01

    In this work we analyze the effect of the inclusion of an empirical dispersion term to standard DFT (DFT-D) in the prediction of the conformational energy of the alanine dipeptide (Ala2) and in assessing the relative stabilities of short polyala-nine peptides in helical conformations, i.e., α and 310 helices, from Ala4 to Ala16. The Ala2 conformational energies obtained with the dispersion-corrected GGA functional B97-D are compared to previously published high level MP2 data. Meanwhile, the B97-D performance on larger polyalanine peptides is compared to MP2, B3LYP and RHF calculations obtained at a lower level of theory. Our results show that electron correlation affects the conformational energies of short peptides with a weight that increases with the peptide length. Indeed, while the contribution of vdW forces is significant for larger peptides, in the case of Ala2 it is negligible when compared to solvent effects. Even for short peptides, the inclusion of an empirical dispersion term greatly improves accuracy of DFT methods, providing results that correlate very well with the MP2 reference at no additional computational cost. PMID:25418993

  13. Conformational Flexibility and Peptide Interaction of the Translocation ATPase SecA

    SciTech Connect

    Zimmer, Jochen; Rapoport, Tom A.; Harvard-Med

    2010-09-21

    The SecA ATPase forms a functional complex with the protein-conducting SecY channel to translocate polypeptides across the bacterial cell membrane. SecA recognizes the translocation substrate and catalyzes its unidirectional movement through the SecY channel. The recent crystal structure of the Thermotoga maritima SecA-SecYEG complex shows the ATPase in a conformation where the nucleotide-binding domains (NBDs) have closed around a bound ADP-BeFx complex and SecA's polypeptide-binding clamp is shut. Here, we present the crystal structure of T. maritima SecA in isolation, determined in its ADP-bound form at 3.1 {angstrom} resolution. SecA alone has a drastically different conformation in which the nucleotide-binding pocket between NBD1 and NBD2 is open and the preprotein cross-linking domain has rotated away from both NBDs, thereby opening the polypeptide-binding clamp. To investigate how this clamp binds polypeptide substrates, we also determined a structure of Bacillus subtilis SecA in complex with a peptide at 2.5 {angstrom} resolution. This structure shows that the peptide augments the highly conserved {beta}-sheet at the back of the clamp. Taken together, these structures suggest a mechanism by which ATP hydrolysis can lead to polypeptide translocation.

  14. Relationship between conformational dynamics and electron transfer in a desolvated peptide. Part I. Structures.

    PubMed

    Semrouni, David; Clavaguéra, Carine; Ohanessian, Gilles; Parks, Joel H

    2013-02-14

    The structures, dynamics and energetics of the protonated, derivatized peptide DyeX-(Pro)(4)-Arg(+)-Trp, where "Dye" stands for the BODIPY analogue of tetramethylrhodamine and X is a (CH(2))(5) linker, have been investigated using a combination of modeling approaches in order to provide a numerical framework to the interpretation of fluorescence quenching data in the gas phase. Molecular dynamics (MD) calculations using the new generation AMOEBA force field were carried out using a representative set of conformations, at eight temperatures ranging from 150 to 500 K. Force field parameters were derived from ab initio calculations for the Dye. Strong electrostatic, polarization and dispersion interactions combine to shape this charged peptide. These effects arise in particular from the electric field generated by the charge of the protonated arginine and from several hydrogen bonds that can be established between the Dye linker and the terminal Trp. This conclusion is based on both the analysis of all structures generated in the MD simulations and on an energy decomposition analysis at classical and quantum mechanical levels. Structural analysis of the simulations at the different temperatures reveals that the relatively rigid polyproline segment allows for the Dye and Trp indole side chain to adopt stacking conformations favorable to electron transfer, yielding support to a model in which it is electron transfer from tryptophan to the dye that drives fluorescence quenching.

  15. Zeta Inhibitory Peptide Disrupts Electrostatic Interactions That Maintain Atypical Protein Kinase C in Its Active Conformation on the Scaffold p62.

    PubMed

    Tsai, Li-Chun Lisa; Xie, Lei; Dore, Kim; Xie, Li; Del Rio, Jason C; King, Charles C; Martinez-Ariza, Guillermo; Hulme, Christopher; Malinow, Roberto; Bourne, Philip E; Newton, Alexandra C

    2015-09-04

    Atypical protein kinase C (aPKC) enzymes signal on protein scaffolds, yet how they are maintained in an active conformation on scaffolds is unclear. A myristoylated peptide based on the autoinhibitory pseudosubstrate fragment of the atypical PKCζ, zeta inhibitory peptide (ZIP), has been extensively used to inhibit aPKC activity; however, we have previously shown that ZIP does not inhibit the catalytic activity of aPKC isozymes in cells (Wu-Zhang, A. X., Schramm, C. L., Nabavi, S., Malinow, R., and Newton, A. C. (2012) J. Biol. Chem. 287, 12879-12885). Here we sought to identify a bona fide target of ZIP and, in so doing, unveiled a novel mechanism by which aPKCs are maintained in an active conformation on a protein scaffold. Specifically, we used protein-protein interaction network analysis, structural modeling, and protein-protein docking to predict that ZIP binds an acidic surface on the Phox and Bem1 (PB1) domain of p62, an interaction validated by peptide array analysis. Using a genetically encoded reporter for PKC activity fused to the p62 scaffold, we show that ZIP inhibits the activity of wild-type aPKC, but not a construct lacking the pseudosubstrate. These data support a model in which the pseudosubstrate of aPKCs is tethered to the acidic surface on p62, locking aPKC in an open, signaling-competent conformation. ZIP competes for binding to the acidic surface, resulting in displacement of the pseudosubstrate of aPKC and re-engagement in the substrate-binding cavity. This study not only identifies a cellular target for ZIP, but also unveils a novel mechanism by which scaffolded aPKC is maintained in an active conformation. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  16. Conformational analysis of opioid peptides in the solid states and the membrane environments by NMR spectroscopy.

    PubMed

    Naito, Aira; Nishimura, Katsuyuki

    2004-01-01

    Determination of conformations and structures of opioid peptides in the membrane environments is an essential step to understand the action of the peptide to the specialized receptors. This information not only gains insight into the structure-function relationship of opioid peptide but also gives proper guidelines to design a new drug to have same neuroendocrine functions. This review provides the structural studies of three types of opioid peptide families such as enkephalin, beta-endorphin and dynorphin in the solid states and the membrane environments. The structures of enkephalins show that they take beta-bend, extended and double beta-bend structures in the crystals. Moreover, enkephalin molecules take a variety of structures in the crystals and are easily converted to the other structures with slightly different torsion angles. On the other hand, beta-bend structures are mostly seen in the membrane environments. Membrane bound structure of dynorphin shows that the N-terminus forms alpha-helical structure and is inserted into the membrane with the helical axis almost perpendicular to the membrane surface. It is discussed that the helical region of the extracellular loop II of the kappa-opioid receptor may interact with the helical region of dynorphin with a high affinity in the membrane environments. beta-endorphin takes alpha-helical structure at N-terminus and the central regions and the rest of regions take unordered structure when the bind to the membrane. Since the membrane bound structures of opioid peptides differ from those of the solution states, membrane association is an important process for exerting the affinity and the selectivity to the specific opioid receptors.

  17. Design of peptides with α,β-dehydro-residues: syntheses, crystal structures and molecular conformations of two ΔPhe-Trp containing peptides

    NASA Astrophysics Data System (ADS)

    Vijayaraghavan, R.; Makker, J.; Kumar, P.; Dey, S.; Singh, T. P.

    2003-06-01

    The ΔPhe-Trp is a newly designed moiety that was found inducing a unique conformation in peptides. The peptides Boc-L-Val-ΔPhe-L-Trp-OCH 3 (I) and Boc-L-Leu-ΔPhe-L-Trp-OCH 3 (II) were synthesized by azlactone method in solution phase. The peptide (I) was crystallized from its solution in ethanol-water mixture in orthorhombic space group P2 12 12 1 with a=10.663(3) Å, b=11.204(3) Å, c=26.516(10) Å and peptide (II) was crystallized from its solution in acetone in a monoclinic space group P2 1 with a=9.354(1)Å, b=11.218(4)Å, c=15.633(1)Å and β=101.83(1)°. The structures were determined by direct methods. Peptide (I) was refined to an R value of 0.059 for 1554 observed reflections [ I≥2 σ (I)] and peptide (II) was refined to an R value of 0.043 for 2920 observed reflections [ I≥2 σ (I)]. The structures of peptides (I) and (II) were found to be identical. They formed an unusual type VIa β-turn conformation which is observed for the first time with a ΔPhe residue at ( i+2) position indicating a unique influence of ΔPhe-Trp moiety in inducing a reproducible new structure in peptides.

  18. Ribosomal Synthesis of Macrocyclic Peptides in Vitro and in Vivo Mediated by Genetically Encoded Amino-Thiol Unnatural Amino Acids

    PubMed Central

    Frost, John R.; Jacob, Nicholas T.; Papa, Louis J.; Owens, Andrew E.

    2015-01-01

    A versatile method for orchestrating the formation of side-chain-to-tail cyclic peptides from ribosomally derived polypeptide precursors is reported. Upon ribosomal incorporation into intein-containing precursor proteins, designer unnatural amino acids bearing side-chain 1,3- or 1,2-aminothiol functionalities are able to promote the cyclization of a downstream target peptide sequence via a C-terminal ligation/ring contraction mechanism. Using this approach, peptide macrocycles of variable size and composition could be generated in a pH-triggered manner in vitro, or directly in living bacterial cells. This methodology furnishes a new platform for the creation and screening of genetically encoded libraries of conformationally constrained peptides. This strategy was applied to identify and isolate a low micromolar streptavidin binder (KD = 1.1 µM) from a library of cyclic peptides produced in E. coli, thereby illustrating its potential toward aiding the discovery of functional peptide macrocycles. PMID:25933125

  19. Force field-based conformational searches: efficiency and performance for peptide receptor complexes

    NASA Astrophysics Data System (ADS)

    Grebner, Christoph; Niebling, Stephan; Schmuck, Carsten; Schlücker, Sebastian; Engels, Bernd

    2013-09-01

    Conformational search using force field methods on complex biomolecular systems is a key factor in understanding molecular and structural properties. The reliability of such investigations strongly depends on the efficiency of the conformational search algorithm as well as the accuracy of the employed force field. In the present work we compared the performance of two different approaches: the Monte-Carlo multiple minimum/low mode sampling (MCMM/LM), in combination with the OPLS2005 (MCMM/LM//OPLS2005), and Tabu-Search combined with Basin Hopping (TS/BH), employing the original OPLS-AA implementation proposed by Jorgensen (TS/BH//OPLS-AA). We investigated their performance in locating energetically low-lying structures and the efficiency in scanning the conformational phase space of non-covalently bonded complexes. As test systems we employed complexes of the artificial peptide receptor CBS-KKF with four different tetrapeptide ligands. The reliability and the accuracy of both approaches were examined by re-optimising all low-energy structures employing density functional theory with empirical dispersion correction in combination with triple zeta basis sets. Solvent effects were mimicked by a continuum solvent model. In all the four-test systems, the TS/BH//OPLS-AA approach yielded structures that are much lower in energy after the DFT optimisation. Additionally, it provided many low-lying structures that were not identified by the MCMM/LM//OPLS2005 approach.

  20. Pulsed hydrogen–deuterium exchange mass spectrometry probes conformational changes in amyloid beta (Aβ) peptide aggregation

    PubMed Central

    Zhang, Ying; Rempel, Don L.; Zhang, Jun; Sharma, Anuj K.; Mirica, Liviu M.; Gross, Michael L.

    2013-01-01

    Probing the conformational changes of amyloid beta (Aβ) peptide aggregation is challenging owing to the vast heterogeneity of the resulting soluble aggregates. To investigate the formation of these aggregates in solution, we designed an MS-based biophysical approach and applied it to the formation of soluble aggregates of the Aβ42 peptide, the proposed causative agent in Alzheimer’s disease. The approach incorporates pulsed hydrogen–deuterium exchange coupled with MS analysis. The combined approach provides evidence for a self-catalyzed aggregation with a lag phase, as observed previously by fluorescence methods. Unlike those approaches, pulsed hydrogen–deuterium exchange does not require modified Aβ42 (e.g., labeling with a fluorophore). Furthermore, the approach reveals that the center region of Aβ42 is first to aggregate, followed by the C and N termini. We also found that the lag phase in the aggregation of soluble species is affected by temperature and Cu2+ ions. This MS approach has sufficient structural resolution to allow interrogation of Aβ aggregation in physiologically relevant environments. This platform should be generally useful for investigating the aggregation of other amyloid-forming proteins and neurotoxic soluble peptide aggregates. PMID:23959898

  1. Aqueous ionic liquids and their influence on peptide conformations: denaturation and dehydration mechanisms.

    PubMed

    Diddens, Diddo; Lesch, Volker; Heuer, Andreas; Smiatek, Jens

    2017-08-09

    Low concentrated aqueous ionic liquids (ILs) and their influence on protein structures have attracted a lot of interest over the last few years. This can be mostly attributed to the fact that aqueous ILs, depending on the ion species involved, can be used as protein protectants or protein denaturants. Atomistic molecular dynamics (MD) simulations are performed in order to study the influence of different aprotic ILs on the properties of a short hairpin peptide. Our results reveal distinct binding and denaturation effects for 1-ethyl-3-methylimidazolium (EMIM) in combination with different anions, namely, chloride (CL), tetrafluoroborate (BF4) and acetate (ACE). The simulation outcomes demonstrate that the studied ILs with larger anions reveal a more pronounced accumulation behavior of the individual ion species around the peptide, which is accomplished by a stronger dehydration effect. We can relate these findings to the implications of the Kirkwood-Buff theory, which provides a thermodynamic explanation for the denaturation strength in terms of the IL accumulation behavior. The results for the spatial distribution functions, the binding energies and the local/bulk partition coefficients are in good agreement with metadynamics simulations in order to determine the energetically most stable peptide conformations. The free energy landscapes indicate a decrease of the denaturation strength in the order EMIM/ACE, EMIM/BF4 and EMIM/CL, which coincides with a decreasing size of the anion species. An analysis of the potential binding energies reveals that this effect is mainly of enthalpic nature.

  2. Dinuclear titanium(IV) complexes from amino acid bridged dicatechol ligands: formation, structure, and conformational analysis.

    PubMed

    Albrecht, M; Napp, M; Schneider, M; Weis, P; Fröhlich, R

    2001-09-17

    Amino acid bridged dicatechol ligands 3a-e-H4 form dinuclear double-stranded coordination compounds [(3a-e)2Ti2(OCH3)2]2- with titanium(IV) ions. Due to the directionality of the ligands, the chirality of the strand, and the chiral complex units, up to seven isomers, I-VII, can be obtained for the double-stranded complexes of ligands 3a-e-H4. The composition of the mixture of isomeric compounds in solution is strongly dependent on the conditions of complex formation. Under thermodynamic control, only a few isomers are obtained, one of which is the major component of the mixture. X-ray structure analyses were performed for K2[(3b)2Ti2(OH)2] and K2[(3d)2Ti2(OH)2] (type I), and for the meso complex Na2[(3e)(3e')Ti2(OCH3)2]. A conformational analysis that uses Ramachandrans method revealed that the conformation of the amino acids in the ligand strands can be compared with those found for amino acids in helical peptide structures. The most favored isomer of [(3)2Ti2(OCH3)2]2- appears to be of type I, with the catecholamide unit located at the N terminus of the ligand strand that binds to a lambda-configurated titanium(IV) complex unit and the dihydroxybenzyl group at the C terminus that coordinates to a delta-configurated titanium(IV) complex unit. The lambda configuration at the N terminus induces the conformation of a right-handed helix in the amino acid residue, while the delta configuration induces the less favored left-handed helix.

  3. Recent advances in the investigation of the bioactive conformation of peptides active at the micro-opioid receptor. conformational analysis of endomorphins.

    PubMed

    Gentilucci, Luca; Tolomelli, Alessandra

    2004-01-01

    Despite of the recent advances in the structural investigation of complex molecules, the comprehension of the 3D features responsible for the interaction between opioid peptides and micro-opioid receptors still remains an elusive task. This has to be attributed to the intrinsic nature of opioid peptides, which can assume a number of different conformations of similar energy, and to the flexibility of the receptorial cavity, which can modify its inner shape to host different ligands. Due to this inherent mobility of the ligand-receptor system, massive efforts devoted to the definition of a rigid bioactive conformation to be used as a template for the design of new pharmacologically active compounds might be overstressed. The future goal might be the design of peptide or nonpeptide ligands capable of maximizing specific hydrophobic interactions. This review covers the recent opinions emerged on the nature of the ligand-receptor interaction, and the development of suitable models for the determination of the bioactive conformation of peptide ligands active towards micro-opioid receptors.

  4. Amino Acid and Peptide Immobilization on Oxidized Nanocellulose: Spectroscopic Characterization

    PubMed Central

    Barazzouk, Saïd; Daneault, Claude

    2012-01-01

    In this work, oxidized nanocellulose (ONC) was synthesized and chemically coupled with amino acids and peptides using a two step coupling method at room temperature. First, ONC was activated by N-ethyl-N’-(3-dimethylaminopropyl) carbodiimide hydrochloride, forming a stable active ester in the presence of N-hydroxysuccinimide. Second, the active ester was reacted with the amino group of the amino acid or peptide, forming an amide bond between ONC and the grafted molecule. Using this method, the intermolecular interaction of amino acids and peptides was avoided and uniform coupling of these molecules on ONC was achieved. The coupling reaction was very fast in mild conditions and without alteration of the polysaccharide. The coupling products (ONC-amino acids and ONC-peptides) were characterized by transmission electron microscopy and by the absorption, emission, Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) spectroscopic techniques.

  5. Peptide nucleic acid probes with charged photocleavable mass markers

    PubMed Central

    Ball, Rachel J; Green, Philip S; Gale, Nittaya; Langley, G John

    2010-01-01

    Halogen-labelled peptide organic acid (HPOA) monomers have been synthesised and incorporated into sequence-specific peptide nucleic acid (PNA) probes. Three different types of probe have been prepared; the unmodified PNA probe, the PNA probe with a mass marker, and the PNA probe with photocleavable mass marker. All three types of probe have been used in model studies to develop a mass spectrometry-based hybridisation assay for detection of point mutations in DNA. PMID:21687524

  6. Ab initio-based exciton model of amide I vibrations in peptides: definition, conformational dependence, and transferability.

    PubMed

    Gorbunov, Roman D; Kosov, Daniil S; Stock, Gerhard

    2005-06-08

    Various aspects of the ab initio-based parametrization of an exciton model of amide I vibrations in peptides are discussed. Adopting "glycine dipeptide" (Ac-Gly-NHCH3) as a simple building-block model that describes the vibrational interaction between two peptide units, we perform comprehensive quantum-chemical calculations to investigate the effect and importance of the level of theory, the choice of local coordinates, and the localization method. A solvent continuum model description turns out important to obtain planar CONH peptide units when a full geometry optimization (which is necessary to obtain the correct frequencies) is performed. To study the conformational dependence of the amide I vibrations, we calculate (phi,psi) maps of the local-mode frequencies and couplings. Performing conformational averages of the (phi,psi) maps with respect to the most important peptide conformational states in solution (alpha, beta, P(II), and C5), we discuss the relation between these measurable quantities and the corresponding conformation of the peptide. Finally, the transferability of these maps to dipeptides with hydrophilic and hydrophobic side chains as well as to tripeptides with charged end groups is investigated.

  7. Examining Polyglutamine Peptide Length: A Connection between Collapsed Conformations and Increased Aggregation

    PubMed Central

    Walters, Robert H.; Murphy, Regina M.

    2009-01-01

    Abnormally expanded polyglutamine domains in proteins are associated with several neurodegenerative diseases, of which the best known is Huntington’s. Expansion of the polyglutamine domain facilitates aggregation of the affected protein, and several studies directly link aggregation to neurotoxicity. The age of onset of disease is inversely correlated with the length of the polyglutamine domain; this correlation motivates an examination of the role of the length of the domain on aggregation. In this investigation, peptides containing 8 to 24 glutamines were synthesized, and their conformational and aggregation properties were examined. All peptides lacked secondary structure. Fluorescence resonance energy transfer (FRET) studies revealed that the peptides became increasingly collapsed as the number of glutamine residues increased. The effective persistence length was estimated to decrease from ~11 Å to ~7 Å as the number of glutamines increased from 8 to 24. A comparison of our data with theoretical results suggests that phosphate-buffered saline is a good solvent for Q8 and Q12, a theta solvent for Q16, and a poor solvent for Q20 and Q24. By dynamic light scattering, we observed that Q16, Q20 and Q24, but not Q8 or Q12, immediately formed soluble aggregates upon dilution into phosphate buffered saline at 37°C. Thus, Q16 stands at the transition point between good and poor solvent, and between stable and aggregation-prone peptide. Examination of aggregates by transmission electron microscopy, along with kinetic assays for sedimentation, provided evidence indicating that soluble aggregates mature into sedimentable aggregates. Together, the data support a mechanism of aggregation in which monomer collapse is accompanied by formation of soluble oligomers; these soluble species lack regular secondary structure but appear morphologically similar to the sedimentable aggregates into which they eventually mature. PMID:19699209

  8. Conformational properties of surfactant-like peptides with variable glycine tails

    NASA Astrophysics Data System (ADS)

    Arkın, Handan

    2010-01-01

    The three-dimensional structures of surfactant-like peptides containing 4-10 glycines as the components of the hydrophobic tails and aspartic acids as the hydrophilic heads (G 4D 2, G 6D 2, G 8D 2, G 10D 2) are investigated by using the multicanonical simulation procedure. The thermodynamically most stable low energy structures of the sequences are determined. Ramachandran plots are prepared and analyzed to predict the secondary structure motifs of the molecules.

  9. Relationship between peptide amino acid sequence and membrane curvature generation

    NASA Astrophysics Data System (ADS)

    Schmidt, Nathan; Kuo, David; Hwee Lai, Ghee; Mishra, Abhijit; Wong, Gerard

    2012-02-01

    Amphipathic peptides and amphipathic domains in proteins can perturb and restructure biological membranes. For example, it is believed that the cationic, amphipathic motif found in membrane active antimicrobial peptides (AMPs) is responsible for their membrane disruption mechanisms of action. And ApoA-I, the main apolipoprotein in high density lipoprotein contains a series of amphipathic α-helical repeats which are responsible for its lipid associating properties. We use small angle x-ray scattering (SAXS) to investigate the interaction of model cell membranes with prototypical AMPs and consensus peptides derived from the helical structural motif of ApoA-I. The relationship between peptide sequence and the peptide-induced changes in membrane curvature and topology is examined. By comparing the membrane rearrangement and corresponding phase behavior induced by these two distinct classes of membrane restructuring peptides we will discuss the role of amino acid sequence on membrane curvature generation.

  10. Conformational studies on the beta subunits of human hemoglobin and their arginyl-COOH peptides.

    PubMed

    Bucci, C F; Bucci, E

    1975-10-07

    The beta subunits of hemoglobin upon alkylation of the cysteinyl residues with iodoacetamide showed a sedimentation velocity with an S20w, near 1.8 as for monomeric subunits. They reacted with alpha chains to give a tetrameric hemoglobin with a sedimentation constant near 4.4. Their CD spectrum was indistinguishable from that of untreated beta chains below 270 nm, otherwise they showed some deviation that became pronounced in the Soret region, where the optical activity of the alkylated subunits was definitely lower than that of the native subunits. Upon removal of the heme the apo-beta subunits showed a decreased optical activity in the far-uv region of the spectrum indicating a substantial loss of helical content. Their sedimentation behavior was consistent with the presence of large aggregates, which dissociates into monomers upon reconstitution with cyanoheme. The apo-beta subunits could be renatured from 6 M guanidine hydrochloride. They showed a stoichiometric reaction with heme in the molar ratio 1:1. Upon reconstitution with the heme their optical activity became similar to that of the native beta chains in the far-uv region of the spectrum, but remained lower in the near-uv and Soret regions. After acylation of the lysyl residues with citraconic anhydride the apo-beta subunits were digested with trypsin and the arginyl-COOH peptides beta(1-30), beta(31-40), beta(41-104), and beta(105-146) were separated by gel chromatography. With the exception of the peptide beta/105-146), which was insoluble at neutral pH, the sedimentation behavior of the other peptides showed the presence of small polymers. The sedimentation behavior of the peptide beta(31-40) was not tested. The percentage of alpha helix, beta conformation, and of random coil (or unordered structure) of the various proteins and peptides was measured fitting their CD spectra in the far-uv region with the parameter published by Y.H. Chen et al. ((1974), Biochemistry 13, 3350) and by N. Greenfield and G

  11. Proline Editing: A General and Practical Approach to the Synthesis of Functionally and Structurally Diverse Peptides. Analysis of Steric versus Stereoelectronic Effects of 4-Substituted Prolines on Conformation within Peptides

    PubMed Central

    Pandey, Anil K.; Naduthambi, Devan; Thomas, Krista M.; Zondlo, Neal J.

    2013-01-01

    Functionalized proline residues have diverse applications. Herein we describe a practical approach, proline editing, for the synthesis of peptides with stereospecifically modified proline residues. Peptides are synthesized by standard solid-phase-peptide-synthesis to incorporate Fmoc-Hydroxyproline (4R-Hyp). In an automated manner, the Hyp hydroxyl is protected and the remainder of the peptide synthesized. After peptide synthesis, the Hyp protecting group is orthogonally removed and Hyp selectively modified to generate substituted proline amino acids, with the peptide main chain functioning to “protect” the proline amino and carboxyl groups. In a model tetrapeptide (Ac-TYPN-NH2), 4R-Hyp was stereospecifically converted to 122 different 4-substituted prolyl amino acids, with 4R or 4S stereochemistry, via Mitsunobu, oxidation, reduction, acylation, and substitution reactions. 4-Substituted prolines synthesized via proline editing include incorporated structured amino acid mimetics (Cys, Asp/Glu, Phe, Lys, Arg, pSer/pThr), recognition motifs (biotin, RGD), electron-withdrawing groups to induce stereoelectronic effects (fluoro, nitrobenzoate), handles for heteronuclear NMR (19F:fluoro; pentafluorophenyl or perfluoro-tert-butyl ether; 4,4-difluoro; 77SePh) and other spectroscopies (fluorescence, IR: cyanophenyl ether), leaving groups (sulfonate, halide, NHS, bromoacetate), and other reactive handles (amine, thiol, thioester, ketone, hydroxylamine, maleimide, acrylate, azide, alkene, alkyne, aryl halide, tetrazine, 1,2-aminothiol). Proline editing provides access to these proline derivatives with no solution phase synthesis. All peptides were analyzed by NMR to identify stereoelectronic and steric effects on conformation. Proline derivatives were synthesized to permit bioorthogonal conjugation reactions, including azide-alkyne, tetrazinetrans-cyclooctene, oxime, reductive amination, native chemical ligation, Suzuki, Sonogashira, cross-metathesis, and Diels

  12. Fragmentation reactions of deprotonated peptides containing aspartic acid

    NASA Astrophysics Data System (ADS)

    Harrison, Alex G.; Young, Alex B.

    2006-09-01

    The fragmentation reactions of deprotonated peptides containing aspartic acid have been elucidated using MS2 and MS3 experiments and accurate mass measurements where necessary. The disposition of labile (N and O bonded) hydrogens in the fragmentation products has been studied by exchanging the labile hydrogens for deuterium whereby the [MD]- ion is formed on electrospray ionization. [alpha]-Aspartyl and [beta]-aspartyl dipeptides give very similar fragment ion spectra on collisional activation, involving for both species primarily formation of the y1 ion and loss of H2O from [MH]- followed by further fragmentation, thus precluding the distinction of the isomeric species by negative ion tandem mass spectrometry. Dipeptides of sequence HXxxAspOH give characteristic spectra different from the [alpha]- and [beta]-isomers. For larger peptides containing aspartic acid a common fragmentation reaction involves nominal cleavage of the NC bond N-terminal to the aspartic acid residue to form a c ion (deprotonated amino acid amide (c1) or peptide amide (cn)) and the complimentary product involving elimination of a neutral amino acid amide or peptide amide. When aspartic acid is in the C-terminal position this fragmentation reaction occurs from the [MH]- ion while when the aspartic acid is not in the C-terminal position the fragmentation reaction occurs mainly from the [MHH2O]- ion. The products of this NC bond cleavage reaction serve to identify the position of the aspartic acid residue in the peptide.

  13. Structural and conformational dynamics of self-assembling bioactive β-sheet peptide nanostructures decorated with multivalent RNA-binding peptides.

    PubMed

    Han, Sanghun; Kim, Donghun; Han, So-hee; Kim, Nam Hee; Kim, Sun Hee; Lim, Yong-beom

    2012-09-26

    Understanding the dynamic behavior of nanostructural systems is important during the development of controllable and tailor-made nanomaterials. This is particularly true for nanostructures that are intended for biological applications because biomolecules are usually highly dynamic and responsive to external stimuli. In this Article, we investigated the structural and conformational dynamics of self-assembling bioactive β-sheet peptide nanostructures using electron paramagnetic resonance (EPR) spectroscopy. The model peptide nanostructures are characterized by the cross-β spine of β-ribbon fibers and multiple RNA-binding bioactive peptides that constitute the shell of the nanostructures. We found first, that bioactive peptides at the shell of β-ribbon nanostructure have a mobility similar to that of an isolated monomeric peptide. Second, the periphery of the cross-β spine is more immobile than the distal part of surface-displayed bioactive peptides. Third, the rotational dynamics of short and long fibrils are similar; that is, the mobility is largely independent of the extent of aggregation. Fourth, peptides that constitute the shell are affected first by the external environment at the initial stage. The cross-β spine resists its external environment to a certain extent and abruptly disintegrates when the perturbation reaches a certain degree. Our results provide an overall picture of β-sheet peptide nanostructure dynamics, which should be useful in the development of dynamic self-assembled peptide nanostructures.

  14. VCD studies on cyclic peptides assembled from L-α-amino acids and a trans-2-aminocyclopentane- or trans-2-aminocyclohexane carboxylic acid.

    PubMed

    Vass, E; Strijowski, U; Wollschläger, K; Mándity, I M; Szilvágyi, G; Jewgiński, M; Gaus, K; Royo, S; Majer, Z; Sewald, N; Hollósi, M

    2010-11-01

    The increasing interest in peptidomimetics of biological relevance prompted us to synthesize a series of cyclic peptides comprising trans-2-aminocyclohexane carboxylic acid (Achc) or trans-2-aminocyclopentane carboxylic acid (Acpc). NMR experiments in combination with MD calculations were performed to investigate the three-dimensional structure of the cyclic peptides. These data were compared to the conformational information obtained by electronic circular dichroism (ECD) and vibrational circular dichroism (VCD) spectroscopy. Experimental VCD spectra were compared to theoretical VCD spectra computed quantum chemically at B3LYP/6-31G(d) density functional theory (DFT) level. The good agreement between the structural features derived from the VCD spectra and the NMR-based structures underlines the applicability of VCD in studying the conformation of small cyclic peptides.

  15. Spectral and biological evaluation of a synthetic antimicrobial peptide derived from 1-aminocyclohexane carboxylic acid.

    PubMed

    Abercrombie, J J; Leung, Kai P; Chai, Hanbo; Hicks, Rickey P

    2015-03-15

    Ac-GF(A6c)G(A6c)K(A6c)G(A6c)F(A6c)G(A6c)GK(A6c)KKKK-amide (A6c=1-aminocyclohexane carboxylic acid) is a synthetic antimicrobial peptide (AMP) that exhibits in vitro inhibitory activity against drug resistant strains of Staphylococcus aureus, Acinetobacter baumannii, Klebsiella pneumoniae, Pseudomonas aeruginosa, Enterobacter aerogenes, and Enterococcus faecium at concentrations ranging from 10.9 to 43μM. Spectroscopic investigations were conducted to determine how this AMP interacts with simple membrane model systems in order to provide insight into possible mechanisms of action. CD and 2D-(1)H NMR experiments indicated this AMP on binding to SDS and DPC micelles adopts conformations with varying percentages of helical and random coil conformers. CD investigations in the presence of three phospholipid SUVs consisting of POPC, 4:1 POPC/POPG, and 60% POPE/21%POPG/19%POPC revealed: (1) The interactions occurring with POPC SUVs have minimal effect on the conformational diversity of the AMP yielding conformations similar to those observed in buffer. (2) The interactions with 4:1 POPC/POPG, and 60% POPE/21%POPG/19%POPC SUVs exhibited a greater influence on the percentage of different conformers contributing to the CD spectra. (3) The presence of a high of percentage of helical conformers was not observed in the presence of SUVs as was the case with micelles. This data indicates that the diversity of surface bound conformations adopted by this AMP are very different from the diversity of conformations adopted by this AMP on insertion into the lipid bilayer. CD spectra of this AMP in the presence of SUVs consisting of LPS isolated from P. aeruginosa, K. pneumoniae and Escherichia coli exhibited characteristics associated with various helical conformations.

  16. The Conformational Stability of Nonfibrillar Amyloid-β Peptide Oligomers Critically Depends on the C-Terminal Peptide Length

    PubMed Central

    2014-01-01

    The amyloid-β (Aβ) peptide is one key molecule in the pathogenesis of Alzheimer’s disease. We investigated the conformational stability of a nonfibrillar tetrameric Aβ structure by molecular dynamics (MD) simulations revealing that the stability of the Aβ tetramer depends critically on the C-terminal length. In contrast to the Aβ17–40 tetramer, which proved to be instable, the simulations demonstrate structural integrity of the Aβ17–42 and Aβ17–43 tetramers. These differences in stability can be attributed to an extension of the middle strand of a three-stranded antiparallel β sheet through residues 41–43, only present in the longer Aβ species that aggregate faster and are more neurotoxic. Additional MD simulations demonstrate that this higher stability is also present in the monomers forming the tetramer. In conclusion, our findings suggest the existence of a nonfibrillar oligomer topology that is significantly more stable for the longer Aβ species, thus offering a structural explanation for their higher neurotoxicity. PMID:24494584

  17. Conformational dynamics of two natively unfolded fragment peptides: Comparison of the AMBER and CHARMM force fields

    PubMed Central

    Chen, Wei; Shi, Chuanyin; MacKerell, Alexander D.; Shen, Jana

    2015-01-01

    Physics-based force fields are the backbone of molecular dynamics simulations. In recent years, significant progress has been made in the assessment and improvement of commonly-used force fields for describing conformational dynamics of folded proteins. However, the accuracy for the unfolded states remains unclear. The latter is however important for detailed studies of protein folding pathways, conformational transitions involving unfolded states and dynamics of intrinsically disordered proteins. In this work we compare the three commonly-used force fields, AMBER ff99SB-ILDN, CHARMM22/CMAP and CHARMM36, for modeling the natively unfolded fragment peptides, NTL9(1-22) and NTL9(6-17), using explicit-solvent replica-exchange molecular dynamics simulations. All three simulations show that NTL9(6-17) is completely unstructured, while NTL9(1-22) transiently samples various β-hairpin states, reminiscent of the first β-hairpin in the structure of the intact NT9 protein. The radius of gyration of the two peptides is force field independent but likely underestimated due to the current deficiency of additive force fields. Compared to the CHARMM force fields, ff99SB-ILDN gives slightly higher β-sheet propensity and more native-like residual structures for NTL9(1-22), which may be attributed to its known β preference. Surprisingly, only two sequence-local pairs of charged residues make appreciable ionic contacts in the simulations of NTL9(1-22), which are sampled slightly more by the CHARMM force fields. Taken together, these data suggest that the current CHARMM and AMBER force fields are globally in agreement in modeling the unfolded states corresponding to β-sheet in the folded structure, while differing in details such as the native-likeness of the residual structures and interactions. PMID:26020564

  18. Solution Phase Conformation and Proteolytic Stability of Amide-Linked Neuraminic Acid Analogues

    PubMed Central

    Saludes, Jonel P.; Gregar, Travis Q.; Monreal, I. Abrrey; Cook, Brandan M.; Danan-Leon, Lieza M.; Gervay-Hague, Jacquelyn

    2013-01-01

    Amide-linked homopolymers of sialic acid offer the advantages of stable secondary structure and increased bioavailability making them useful constructs for pharmaceutical design and drug delivery. Defining the structural characteristics that give rise to secondary structure in aqueous solution is challenging in homopolymeric material due to spectral overlap in NMR spectra. Having previously developed computational tools for heteroologomers with resolved spectra, we now report that application of these methods in combination with circular dichroism, NH/ND NMR exchange rates and nOe data has enabled the structural determination of a neutral, δ-amide-linked homopolymer of a sialic acid analogue called Neu2en. The results show that the inherent planarity of the pyranose ring in Neu2en brought about by the α,β-conjugated amide bond serves as the primary driving force of the overall conformation of the homooligomer. This peptide surrogate has an excellent bioavailability profile, with half-life of ~12 hours in human blood serum, which offers a viable peptide scaffold that is resistant to proteolytic degradation. Furthermore, a proof-of-principle study illustrates that Neu2en oligomers are functionalizable with small molecule ligands using 1,3-dipolar cycloaddition chemistry. PMID:23765412

  19. Aromatic amino acids providing characteristic motifs in the Raman and SERS spectroscopy of peptides.

    PubMed

    Wei, Fang; Zhang, Dongmao; Halas, Naomi J; Hartgerink, Jeffrey D

    2008-07-31

    Raman and surface-enhanced Raman spectroscopies (SERS) are potentially important tools in the characterization of biomolecules such as proteins and DNA. In this work, SERS spectra of three cysteine-containing aromatic peptides: tryptophan-cysteine, tyrosine-cysteine, and phenylalanine-cysteine, bound to Au nanoshell substrates, were obtained, and compared to their respective normal Raman spectra. While the linewidths of the SERS peaks are significantly broadened (up to 70%), no significant spectral shifts (<6 cm (-1)) of the major Stokes modes were observed between the two modalities. We show that the Raman and SERS spectra of penetratin, a cell-penetrating peptide oligomer, can be comprised quite reliably from the spectra of its constituent aromatic amino acids except in the backbone regions where the spectral intensities are critically dependent on the length and conformations of the probed molecules. From this study we conclude that, together with protein backbone groups, aromatic amino acid residues provide the overwhelmingly dominant features in the Raman and SERS spectra of peptides and proteins when present. It follows that the Raman modes of these three small constructed peptides may likely apply to the assignment of Raman and SERS features in the spectra of other peptides and proteins.

  20. Intestinal absorption of amino acids and peptides in Hartnup disorder.

    PubMed

    Leonard, J V; Marrs, T C; Addison, J M; Burston, D; Clegg, K M; Lloyd, J K; Matthews, D M; Seakins, J W

    1976-04-01

    Absorption of free and peptide-bound amino acids was investigated in a girl with Hartnup disorder aged 26 months. Plasma levels of amino acids were followed after oral administration of (1) an amino acid mixture simulating casein and (2) an equivalent dose of a partial enzymic hydrolysate of casein containing oligopeptides in addition to free amino acids. The results suggested that many neutral amino acids were poorly absorbed when given in the free form, but much more readily absorbed when given as peptides. Unexpectedly, the results also suggested that glutamic acid was poorly absorbed when given in the free form. The results obtained with threonine could not be interpreted. There was an increased renal clearance of many neutral amino acids, including glycine, but clearance of proline was not increased. Most amino acids with an increased renal clearance also appeared to be poorly absorbed when given by mouth in the free form.

  1. Conformational equilibria and large-amplitude motions in dimers of carboxylic acids: rotational spectrum of acetic acid-difluoroacetic acid.

    PubMed

    Gou, Qian; Feng, Gang; Evangelisti, Luca; Caminati, Walther

    2014-10-06

    We report the rotational spectra of two conformers of the acetic acid-difluoroacetic acid adduct (CH3COOH-CHF2COOH) and supply information on its internal dynamics. The two conformers differ from each other, depending on the trans or gauche orientation of the terminal -CHF2 group. Both conformers display splittings of the rotational transitions, due to the internal rotation of the methyl group of acetic acid. The corresponding barriers are determined to be V3(trans)=99.8(3) and V3(gauche)=90.5(9) cm(-1) (where V3 is the methyl rotation barrier height). The gauche form displays a further doubling of the rotational transitions, due to the tunneling motion of the -CHF2 group between its two equivalent conformations. The corresponding B2 barrier is estimated to be 108(2) cm(-1). The increase in the distance between the two monomers upon OH→OD deuteration (the Ubbelohde effect) is determined.

  2. 4R- and 4S-iodophenyl hydroxyproline, 4R-pentynoyl hydroxyproline, and S-propargyl-4-thiolphenylalanine: conformationally biased and tunable amino acids for bioorthogonal reactions.

    PubMed

    Forbes, Christina R; Pandey, Anil K; Ganguly, Himal K; Yap, Glenn P A; Zondlo, Neal J

    2016-02-21

    Bioorthogonal reactions allow the introduction of new functionalities into peptides, proteins, and other biological molecules. The most readily accessible amino acids for bioorthogonal reactions have modest conformational preferences or bases for molecular interactions. Herein we describe the synthesis of 4 novel amino acids containing functional groups for bioorthogonal reactions. (2S,4R)- and (2S,4S)-iodophenyl ethers of hydroxyproline are capable of modification via rapid, specific Suzuki and Sonogashira reactions in water. The synthesis of these amino acids, as Boc-, Fmoc- and free amino acids, was achieved through succinct sequences. These amino acids exhibit well-defined conformational preferences, with the 4S-iodophenyl hydroxyproline crystallographically exhibiting β-turn (ϕ, ψ∼-80°, 0°) or relatively extended (ϕ, ψ∼-80°, +170°) conformations, while the 4R-diastereomer prefers a more compact conformation (ϕ∼-60°). The aryloxyproline diastereomers present the aryl groups in a highly divergent manner, suggesting their stereospecific use in molecular design, medicinal chemistry, and catalysis. Thus, the 4R- and 4S-iodophenyl hydroxyprolines can be differentially applied in distinct structural contexts. The pentynoate ester of 4R-hydroxyproline introduces an alkyne functional group within an amino acid that prefers compact conformations. The propargyl thioether of 4-thiolphenylalanine was synthesized via copper-mediated cross-coupling reaction of thioacetic acid with protected 4-iodophenylalanine, followed by thiolysis and alkylation. This amino acid combines an alkyne functional group with an aromatic amino acid and the ability to tune aromatic and side chain properties via sulfur oxidation. These amino acids provide novel loci for peptide functionalization, with greater control of conformation possible than with other amino acids containing these functional groups.

  3. Evaluation of basic amphipathic peptides for cellular delivery of antisense peptide nucleic acids.

    PubMed

    Maier, Martin A; Esau, Christine C; Siwkowski, Andrew M; Wancewicz, Edward V; Albertshofer, Klaus; Kinberger, Garth A; Kadaba, Neena S; Watanabe, Tanya; Manoharan, Muthiah; Bennett, C Frank; Griffey, Richard H; Swayze, Eric E

    2006-04-20

    Cellular permeation peptides have been used successfully for the delivery of a variety of cargoes across cellular membranes, including large hydrophilic biomolecules such as proteins, oligonucleotides, or plasmid DNA. For the present work, a series of short amphipathic peptides was designed to elucidate the structural requirements for efficient and nontoxic delivery of peptide nucleic acids (PNAs). On the basis of an idealized alpha-helical structure, the helical parameters were modulated systematically to yield peptides within a certain range of hydrophobicity and amphipathicity. The corresponding PNA conjugates were synthesized and characterized in terms of secondary structure, enzymatic stability, and antisense activity. The study revealed correlations between the physicochemical and biophysical properties of the conjugates and their biological activity and led to the development of potent peptide vectors for the cellular delivery of antisense PNAs. Two representative compounds were radiolabeled and evaluated for their biodistribution in healthy mice.

  4. Advances in the Determination of Nucleic Acid Conformational Ensembles

    PubMed Central

    Salmon, Loïc; Yang, Shan; Al-Hashimi, Hashim M.

    2016-01-01

    Conformational changes in nucleic acids play a key role in the way genetic information is stored, transferred, and processed in living cells. Here, we describe new approaches that employ a broad range of experimental data, including NMR derived chemical shifts and residual dipolar couplings, small angle X-ray scattering, and computational approaches such as molecular dynamics simulations, to determine ensembles of DNA and RNA at atomic resolution. We review the complimentary information that can be obtained from diverse sets of data and the various methods that have been developed to combine these data with computational methods to construct ensembles and assess their uncertainty. We conclude by surveying RNA and DNA ensembles determined using these methods, highlighting the unique physical and functional insights that have been obtained so far. PMID:24364917

  5. Acidic electrolysed water delays browning by destroying conformation of polyphenoloxidase.

    PubMed

    Sun, Jiangping; Wang, Meng; Liu, Haiquan; Xie, Jing; Pan, Yingjie; Xu, Changhua; Zhao, Yong

    2017-05-26

    Browning frequently occurs at fruits, vegetables and aquatic products during storage, and it drastically reduces the consumer's acceptability, with considerable financial loss. The objective of this paper was to investigate the effects of acidic electrolysed water (AEW) technology on polyphenoloxidase (PPO), which is an essential enzyme for browning. AEW ice exhibited a good ability in delaying browning in shrimp. Kinetic study revealed that AEW exhibited the mixed type inhibition of PPO with a Ki value of 1.96 mmol L(-1) . Moreover, both the circular dichroism spectrum and Fourier transform infrared spectroscopy analyses revealed that the α-helix in PPO decreased whereas random coil increased which indicates that PPO conformation was destroyed. Thus, this paper may provide a deeper understanding of the application of AEW technology for preventing browning in the food industry. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  6. Conformational analysis and vibrational assignments of benzohydroxamic acid and benzohydrazide

    NASA Astrophysics Data System (ADS)

    Al-Saadi, Abdulaziz A.

    2012-09-01

    The structures of benzohydroxamic acid (BHA) and benzohydrazide (BH) were investigated at the B3LYP, MP2 and MP4(SDQ) levels of theory and compared to the corresponding structures of formyl analogs. All levels of theory predicted the two molecules to exist predominantly in a near-planar structure adopting a cis conformation where the hydroxyl group of the acid and the amino group of the hydrazide eclipse the carbonyl bond. The stability of the near-planar structure is explained on the basis of mutual conjugation between the phenyl and the Nsbnd H moieties with the Cdbnd O group. The intramolecular interaction between the carbonyl group and the hydrogen atom of the hydroxyl group of the acid or the amino group of the hydrazide plays a significant role in stabilizing the near-cis form in both molecules. The degree of the non-planarity was predicted to increase as going from BHA to BH molecules. The computed vibrational frequencies of the near-cis structure were combined with experimental infrared and Raman data to provide reliable vibrational assignments for the two molecules.

  7. Conformational investigation of designed short linear peptides able to fold into beta-hairpin structures in aqueous solution.

    PubMed

    de Alba, E; Jiménez, M A; Rico, M; Nieto, J L

    1996-01-01

    Formation of secondary structure plays an important role in the early stages of protein folding. The conformational analysis of designed peptides has proved to be very useful for identifying the interactions responsible for the formation and stability of alpha-helices. However, very little is known about the factors leading to the formation of beta-hairpins. In order to get a good beta-hairpin-forming model peptide, two peptides were designed on the basis of beta-sheet propensities and individual statistical probabilities in the turn sites, together with solubility criteria. The conformational properties of the two peptides were analyzed by two-dimensional NMR methods. Long-range cross-correlations observed in NOE and ROE spectra, together with other NMR evidence, show that peptide IYSNPDGTWT forms a highly populated beta-hairpin in aqueous solution with a type I beta-turn plus a G1 beta-bulge conformation in the chain-bend region. The analogous peptide with a Pro5 substituted by Ser forms, in addition to the previous conformation, a second beta-hairpin with a standard type I beta-turn conformation, and the two forms are in fast dynamic equilibrium with one another. The effect of pH demonstrates the existence of a stabilizing interaction between the Asn and Asp sidechains. The populations of beta-hairpin conformations increase in the presence of trifluoroethanol (a structure-enhancing solvent). On the other hand, some residual structure persists at a high denaturant concentration (8 M urea). This work highlights the importance of the beta-turn residue composition in determining the particular type of beta-hairpin adopted by a peptide, though a role of interstrand sidechain interactions in the stabilization of the formed beta-hairpin is not discarded. The fact that trifluoroethanol can stabilize alpha-helices or beta-hairpins depending on the intrinsic properties of the peptide sequence is again shown. An additional example of the presence of residual structure

  8. Investigating the microstructure of keratin extracted from wool: Peptide sequence (MALDI-TOF/TOF) and protein conformation (FTIR)

    NASA Astrophysics Data System (ADS)

    Cardamone, Jeanette M.

    2010-04-01

    Investigations of keratins extracted from wool by reduction hydrolysis and by alkaline hydrolysis showed that their chemical compositions and secondary structures were similar to original wool. The keratin isolates were similar in amino acid, Amides I and II, and secondary structure to each other and to original wool. From SDS-PAGE electrophoresis, keratin isolated by reduction contained protein homologs of molecular weight, ˜40-60 kDa and keratin isolate from alkaline hydrolysis contained peptide fragments of ˜6-8 kDa. MALDI-TOF/TOF spectrometry confirmed that the reduction isolate contained Type II microfibrillar component 7C, hair Type II intermediate filament, Type I microfibrillar 48 kDa component 8C-1, and Type I microfibrillar 47.6 kDa protein homologs which contained alanine, glutamine, glutamic acid, leucine, serine, leucine, and cystine with highest amounts glutamic acid and leucine amino acids. FTIR spectroscopy was applied to examine secondary structure to confirm the content of α-helix/β-sheet/disordered regions for original wool (58.2%/37.9%/3.9%); keratin from reduction (36.7%/50.2%/13.1%); and keratin from alkaline hydrolysis (25.7%/51.8%/22.5%). The higher content of β-sheet secondary structure and intact α-helical conformation characterized these isolates as viable starting materials for chemical modification to form novel bio-based materials useful in industrial formulations and compositions. In particular keratin extracted by reduction with the molecular weight of original wool and the probability of useful mechanical properties can be transformed into stand-alone products of various shapes and forms such as porous foams, sponges, mats, and films for bio-based, adaptable structures.

  9. Near-infrared laser-induced generation of three rare conformers of glycolic acid.

    PubMed

    Halasa, Anna; Lapinski, Leszek; Reva, Igor; Rostkowska, Hanna; Fausto, Rui; Nowak, Maciej J

    2014-07-31

    Structural transformations were induced in conformers of glycolic acid by selective excitation with monochromatic tunable near-infrared laser light. For the compound isolated in Ar matrixes, near-IR excitation led to generation of two higher-energy conformers (GAC; AAT) differing from the most stable SSC form by 180° rotation around the C-C bond. A detailed investigation of this transformation revealed that one conformer (GAC) is produced directly from the near-IR-excited most stable conformer. The other higher-energy conformer (AAT) was effectively generated only upon excitation of the primary photoproduct (GAC) with another near-IR photon. Once these higher-energy conformers of glycolic acid were generated in an Ar matrix, they could be subsequently transformed into one another upon selective near-IR excitations. Interestingly, no repopulation of the initial most stable SSC conformer occurred upon near-IR excitation of the higher-energy forms of the compound isolated in solid Ar. A dramatically different picture of near-IR-induced conformational transformations was observed for glycolic acid isolated in N2 matrixes. In this case, upon near-IR excitation, the most stable SSC form converted solely into a new conformer (SST), where the acid OH group is rotated by 180°. This conformational transformation was found to be photoreversible. Moreover, SST conformer, photoproduced in the N2 matrix, spontaneously converted to the most stable SSC form of glycolic acid, when the matrix was kept at cryogenic temperature and in the dark.

  10. A Natural Interruption Displays Higher Global Stability and Local Conformational Flexibility than a Similar Gly Mutation Sequence in Collagen Mimic Peptides.

    PubMed

    Sun, Xiuxia; Chai, Yalin; Wang, Qianqian; Liu, Huanxiang; Wang, Shaoru; Xiao, Jianxi

    2015-10-06

    Natural interruptions in the repeating (Gly-X-Y)n amino acid sequence pattern are found normally in triple helix domains of all nonfibrillar collagens, while any Gly substitution in fibrillar collagens leads to pathological conditions. As revealed by our sequence analysis, two peptides, one modeling a natural G5G interruption (POALO) and the other one mimicking a pathological Gly-to-Ala substitution (LOAPO), are designed. Circular dichroism (CD), NMR, and computational simulation studies have discovered significant differences in stability, conformation, and folding between the two peptides. Compared with the Gly substitution sequence, the natural interruption maintains higher stability, higher triple helix content, and a higher folding rate while introducing more alterations in local triple helical conformation in terms of dihedral angles and hydrogen bonding. The conserved hydrophobic residues at the specific sites of interruptions may provide functional constraints for higher-order assembly as well as biomolecular interactions. These results suggest a molecular basis of different biological roles of natural interruptions and Gly substitutions and may guide the design of collagen mimic peptides containing functional natural interruptions.

  11. Synthesis of phosphonamidate peptides by Staudinger reactions of silylated phosphinic acids and esters.

    PubMed

    Wilkening, Ina; del Signore, Giuseppe; Hackenberger, Christian P R

    2011-01-07

    The Staudinger reaction of unprotected azido-peptides with silylated phosphinic acids and esters on the solid support offers a straightforward acid-free entry to different phosphonamidate peptide esters or acids under mild conditions in high purity and yield.

  12. Peptide length variants p2Ca and QL9 present distinct conformations to L(d)-specific T cells.

    PubMed

    Hornell, T M; Martin, S M; Myers, N B; Connolly, J M

    2001-10-15

    Recent advances have provided insights into how the TCR interacts with MHC/peptide complexes and a rationale to predict optimal epitopes for MHC binding and T cell recognition. For example, peptides of nine residues are predicted to be optimal for binding to H2-L(d), although 8 mer epitopes have also been identified. It has been predicted that 8 mer and 9 mer length variant peptides bound to L(d) present identical epitopes to T cells. However, in contrast to this prediction, we demonstrate here that the 8 mer peptide p2Ca and its 9 mer length variant QL9, extended by an N-terminal glutamine, assume distinct conformations when bound to L(d). We generated self-L(d)-restricted CTL clones specific for p2Ca that recognize L(d)/QL9 poorly if at all. This result is in sharp contrast to what has been observed with L(d)-alloreactive T cells that possess a much higher affinity for L(d)/QL9 than for L(d)/p2Ca. Alanine substitutions of the N-terminal residues of the QL9 peptide rescue detection by these self-L(d)/p2Ca-specific T cells, but decrease recognition by the L(d)-alloreactive 2C T cell clone. In addition, 2C T cell recognition of the p2Ca peptide is affected by different alanine substitutions compared with 2C T cell recognition of the QL9 peptide. These data clearly demonstrate that the p2Ca and QL9 peptides assume distinct conformations when bound to L(d) and, furthermore, demonstrate that there is flexibility in peptide binding within the MHC class I cleft.

  13. Sequence selective recognition of double-stranded RNA using triple helix-forming peptide nucleic acids.

    PubMed

    Zengeya, Thomas; Gupta, Pankaj; Rozners, Eriks

    2014-01-01

    Noncoding RNAs are attractive targets for molecular recognition because of the central role they play in gene expression. Since most noncoding RNAs are in a double-helical conformation, recognition of such structures is a formidable problem. Herein, we describe a method for sequence-selective recognition of biologically relevant double-helical RNA (illustrated on ribosomal A-site RNA) using peptide nucleic acids (PNA) that form a triple helix in the major grove of RNA under physiologically relevant conditions. Protocols for PNA preparation and binding studies using isothermal titration calorimetry are described in detail.

  14. Conformational behavior of polyalanine peptides with and without protecting groups of varying chain lengths: population of PP-II structure!

    PubMed

    Nandel, Fateh S; Garg, Mohan L; Shafique, Mohd

    2015-05-01

    Oculopharyngeal muscular dystrophy (OPMD), a polyalanine myopathy, occurs due to expansion of homo-polyalanine stretch in normal polyadenylating binding protein nuclear 1 (PABPN1) protein from Ala10 to Ala11-17. Therefore, the conformational behavior of polyalanine peptides with n = 10-17, with and without terminal protecting groups, have been investigated with different starting geometries in water by molecular dynamics simulation studies. Alanine peptides are shown to give rise to unordered structure irrespective of starting geometry and not more than two residues at a stretch have the same/similar set of φ, ψ values. However, the final structure with terminal protecting groups look like β-strand. Unprotected poly-Ala peptides adopt twisted β-hairpin/multi hairpin like structure with increasing chain length. The number of residues having φ, ψ values in collagen region is found to be less in peptides with unprotected termini as compared to peptides with protected termini of same chain length. The results have been supported by recent synchrotron radiation circular dichroism spectroscopy of polyproline II and unordered secondary structures. Opening of the helical structure in poly-Ala peptides with protecting groups has been shown to take place from C-terminal and in peptides without protecting groups opening of helix starts from both terminals. Further, opening of helix takes more time in poly-Ala peptides without terminal protecting groups. The deviations in amide bond planarity have been discussed and compared with available experimental and computational results.

  15. Effects of N-terminus modifications on the conformation and permeation activities of the synthetic peptide L1A.

    PubMed

    Zanin, Luciana Puia Moro; de Araujo, Alexandre Suman; Juliano, Maria Aparecida; Casella, Tiago; Nogueira, Mara Correa Lelles; Ruggiero Neto, João

    2016-06-01

    We investigate the effect of the N-terminus modification of the L1A, a synthetic octadecapeptide, on its helical content, affinity and lytic action in model membranes and on its hemolytic and antibacterial activities. L1A and its acetylated analog displayed a selective antibacterial activity to Gram-negative bacteria without being hemolytic. The covalently linked 2-aminobezoic acid to the N-terminus impaired the antibacterial efficacy and increased hemolysis. Despite their lower net charge (+2), N-terminus modifications resulted in enhanced affinity and improved lytic efficiency in anionic vesicles. The analogs also showed higher helical content and consequently higher amphipathicity in these vesicles. The conformational analysis by molecular dynamics simulations in 30 % of TFE/water showed that the hydrophobic faces of the peptides are in close contact with CF3 groups of TFE while the hydrophilic faces with water molecules. Due to the loss of the amino charge, the N-termini of the analogs are buried in TFE molecules. The analysis of the pair distribution functions, obtained for the center of mass of the charged groups, has evidenced that the state of the N-terminus has influenced the possibility of different ion-pairing. The higher complexity of the bacterial cells compared with anionic vesicles hampers to establish correlations structure-function for the analogs.

  16. Differential tapasin dependence of MHC class I molecules correlates with conformational changes upon peptide dissociation: A molecular dynamics simulation study

    SciTech Connect

    Sieker, Florian; Straatsma, TP; Springer, Sebastian; Zacharias, Martin W

    2008-08-01

    Efficiency of peptide loading to MHC class I molecules in the endoplasmatic reticulum depends on the class I allele and can involve interaction with tapasin and other proteins of the loading complex. Allele HLA-B*4402 (Asp at position 116) depends on tapasin for efficient peptide loading whereas HLA-B*4405 (identical to B*4402 except for Tyr116) can efficiently load peptides in the absence of tapasin. Both alleles adopt very similar structures in the presence of the same peptide. Molecular dynamics (MD) simulations on induced peptide termini dissociation from the α1/α2 peptide binding domains have been performed to characterize free energy changes and associated structural changes in the two alleles. A smooth free energy change along the distance dissociation coordinate was obtained for N terminus dissociation. A different shape and magnitude of the calculated free energy change and was obtained for induced peptide C terminus dissociation in case of the tapasin independent allele B*4405 compared to B*4402. Structural changes during C terminus dissociation occurred mainly in the first segment of the α2-1 helix that flanks the peptide C-terminus binding region (F-pocket) and contacts residue 116. This segment is also close to the proposed tapasin contact region. For B*4402, a stable shift towards an altered open F-pocket structure deviating significantly from the bound form was observed. In contrast, B*4405 showed only a transient opening of the F-pocket followed by relaxation towards a structure close to the bound form upon C terminus dissociation. The greater tendency for peptide-receptive conformation in the absence of peptide combined with a more long-range character of the interactions with the peptide C terminus facilitates peptide binding to B*4405 and could be responsible for the tapasin independence of this allele. A possible role of tapasin in case of HLA-B*4402 and other tapasin-dependent alleles could be the stabilization of a peptide receptive class I

  17. Importance of backbone angles versus amino acid configurations in peptide vibrational Raman optical activity spectra

    NASA Astrophysics Data System (ADS)

    Herrmann, Carmen; Ruud, Kenneth; Reiher, Markus

    2008-01-01

    In this work, we investigate whether the differential scattering of right- and left-circularly polarized light in peptide Raman optical activity spectra are uniquely dominated by the backbone conformation, or whether the configurations of the individual amino acid also play a significant role. This is achieved by calculating Raman optical activity spectra using density functional theory for four structurally related peptides with a common backbone conformation, but with different sequences of amino acid configurations. Furthermore, the ROA signals of the amide normal modes are decomposed into contributions from groups of individual atoms. It is found that the amino acid configuration has a considerable influence on the ROA peaks in the amide I, II, and III regions, although the local decomposition reveals that the side-chain atoms only contribute to those peaks directly in the case of the amide II vibrations. Furthermore, small changes in the amide normal modes may lead to large and irregular modifications in the ROA intensity differences, making it difficult to establish transferable ROA intensity differences even for structurally similar vibrations.

  18. Interaction of sweet proteins with their receptor. A conformational study of peptides corresponding to loops of brazzein, monellin and thaumatin.

    PubMed

    Tancredi, Teodorico; Pastore, Annalisa; Salvadori, Severo; Esposito, Veronica; Temussi, Piero A

    2004-06-01

    The mechanism of interaction of sweet proteins with the T1R2-T1R3 sweet taste receptor has not yet been elucidated. Low molecular mass sweeteners and sweet proteins interact with the same receptor, the human T1R2-T1R3 receptor. The presence on the surface of the proteins of "sweet fingers", i.e. protruding features with chemical groups similar to those of low molecular mass sweeteners that can probe the active site of the receptor, would be consistent with a single mechanism for the two classes of compounds. We have synthesized three cyclic peptides corresponding to the best potential "sweet fingers" of brazzein, monellin and thaumatin, the sweet proteins whose structures are well characterized. NMR data show that all three peptides have a clear tendency, in aqueous solution, to assume hairpin conformations consistent with the conformation of the same sequences in the parent proteins. The peptide corresponding to the only possible loop of brazzein, c[CFYDEKRNLQC(37-47)], exists in solution in a well ordered hairpin conformation very similar to that of the same sequence in the parent protein. However, none of the peptides has a sweet taste. This finding strongly suggests that sweet proteins recognize a binding site different from the one that binds small molecular mass sweeteners. The data of the present work support an alternative mechanism of interaction, the "wedge model", recently proposed for sweet proteins [Temussi, P. A. (2002) FEBS Lett.526, 1-3.].

  19. Ceragenins: cholic acid-based mimics of antimicrobial peptides.

    PubMed

    Lai, Xin-Zhong; Feng, Yanshu; Pollard, Jacob; Chin, Judy N; Rybak, Michael J; Bucki, Robert; Epand, Raquel F; Epand, Richard M; Savage, Paul B

    2008-10-01

    The prevalence of drug-resistant bacteria drives the quest for new antimicrobials, including those that are not expected to readily engender resistance. One option is to mimic Nature's most ubiquitous means of controlling bacterial growth, antimicrobial peptides, which have evolved over eons. In general, bacteria remain susceptible to these peptides. Human antimicrobial peptides play a central role in innate immunity, and deficiencies in these peptides have been tied to increased rates of infection. However, clinical use of antimicrobial peptides is hampered by issues of cost and stability. The development of nonpeptide mimics of antimicrobial peptides may provide the best of both worlds: a means of using the same mechanism chosen by Nature to control bacterial growth without the problems associated with peptide therapeutics. The ceragenins were developed to mimic the cationic, facially amphiphilic structures of most antimicrobial peptides. These compounds reproduce the required morphology using a bile-acid scaffolding and appended amine groups. The resulting compounds are actively bactericidal against both gram-positive and gram-negative organisms, including drug-resistant bacteria. This antimicrobial activity originates from selective association of the ceragenins with negatively charged bacterial membrane components. Association has been studied with synthetic models of bacterial membrane components, with bacterial lipopolysaccharide, with vesicles derived from bacterial phospholipids, and with whole cells. Comparisons of the antimicrobial activities of ceragenins and representative antimicrobial peptides suggest that these classes of compounds share a mechanism of action. Rapid membrane depolarization is caused by both classes as well as blebbing of bacterial membranes. Bacteria express the same genes in response to both classes of compounds. On the basis of the antibacterial activities of ceragenins and preliminary in vivo studies, we expect these compounds to

  20. Water compatible photoarylation of amino acids and peptides.

    PubMed

    Sudakow, Alex; Papke, Uli; Lindel, Thomas

    2014-08-11

    A novel photoarylation of amino acids and peptides is described, which tolerates the presence of water. Irradiation of Boc-protected amino acids in the presence of N-protected 2-azidobenzimidazoles leads to selective arylation of carboxy termini or side chains. The new reaction also works for peptides. Irradiation of the nonapeptide H-SPSYVYHQF-OH also resulted in selective arylation of the tyrosine side chains, as indicated by ESI-MS/MS fragmentation. Chemo- and regioselectivity could add the title reaction to the repertoire of photoaffinity labeling methods. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Conformational assembly and biological properties of collagen mimetic peptides and their thermally responsive polymer conjugates

    NASA Astrophysics Data System (ADS)

    Krishna, Ohm Divyam

    2011-12-01

    Collagens are one of the most abundant proteins found in body tissues and organs, endowing structural integrity, mechanical strength, and multiple biological functions. Destabilized collagen inside human body leads to various degenerative diseases (ex. osteoarthritis) and ageing. This has continued to motivate the design of synthetic peptides and bio-synthetic polypeptides to closely mimic the native collagens in terms of triple helix structure and stability, potential for higher order assembly, and biological properties. However, the widespread application of de novo collagens has been limited in part by the need for hydroxylated proline in the formation of stable triple helical structures. To address this continued need, a hydroxyproline-free, thermally stable collagen-mimetic peptide (CLP-Cys) was rationally designed via the incorporation of electrostatically stabilized amino acid triplets. CLP-Cys was synthesized via solid phase peptide synthesis. The formation and stability of the triple helical structure were indicated via circular dichroism (CD) experiments and confirmed via differential scanning calorimetry (DSC) results. CLP-Cys also self-assembled into nano-rods and micro-fibrils, as evidenced via a combination of dynamic light scattering and transmission electron microscopy. Given the high thermal stability and its propensity for higher-order assembly, CLP-Cys was further functionalized at both the ends with a thermally responsive polymer, poly(diethylene glycol methyl ether methacrylate), (PDEGMEMA) to synthesize a biohybrid triblock copolymer. The CD results indicated that the triple helical form is retained, the thermal unfolding is sustained and helix to coil transition is reversible in the triblock hybrid context. The LCST of PDEGMEMA homopolymer (26 °C) is increased (to 35 °C) upon conjugation to the hydrophilic collagen peptide domain. Further, a combination of static light scattering, Cryo-SEM, TEM and confocal microscopy elucidated that the

  2. Conformational state of a 25-mer peptide from the cyclophilin-binding loop of the HIV type 1 capsid protein.

    PubMed Central

    Reimer, U; Drewello, M; Jakob, M; Fischer, G; Schutkowski, M

    1997-01-01

    Recently a 25-residue part of Gag polyprotein from HIV type 1 (HIV-1) was reported to bind to the cytosolic 18 kDa cyclophilin (Cyp18) with an IC50 value of 180 microM. This peptide corresponds to the Cyp18-binding domain of HIV-1 Gag. A replacement of Gly with Ala in the cyclophilin-binding loop of HIV-1 Gag polyprotein results in the prevention of the packaging of Cyp18 into virions. We found only two conformers of this peptide among 16 possible expected conformers, owing to cis/trans isomerization of four peptidyl-prolyl bonds. Although this finding implicates the existence of a stabilizing structure, we were not able to detect secondary structure formation by 1H-NMR and CD spectroscopy. We characterized the peptide as a substrate for Cyp18 by two-dimensional exchange 1H-NMR spectroscopy. Surprisingly, we found similar binding characteristics for a peptide corresponding to 25-mer peptide containing the above-mentioned Gly to Ala substitution. PMID:9337866

  3. Conformational landscape of isolated capped amino acids: on the nature of non-covalent interactions*

    NASA Astrophysics Data System (ADS)

    González, Jorge; Martínez, Rodrigo; Fernández, José A.; Millan, Judith

    2017-08-01

    The intramolecular interactions for isolated capped amino acids were investigated computationally by characterizing the conformers for selected amino acids with charged (arginine), polar (asparagine and glutamine), non-polar (alanine, valine and isoleucine), and aromatic (phenylalanine, tryptophan and tyrosine) side chains. The computational method applied combined a molecular mechanics conformational search (with an MMFFs forced field) followed by structural and vibrational density-functional calculations (M06-2X with a triple- ζ Pople's basis set). The intramolecular forces in each amino acid were analyzed with the Non-Covalent Interactions (NCI) analysis. The results for the 15 most stable conformers studied showed that the structure of isolated capped amino acids resembles those found in proteins. In particular, the two most stable conformers of the nine amino acids investigated exhibit γ L and β L conformations with 7- and 5-membered rings, respectively, as a result of the balance between non-covalent interactions (hydrogen bonds and van der Waals).

  4. Biologically relevant conformational features of linear and cyclic proteolipid protein (PLP) peptide analogues obtained by high-resolution nuclear magnetic resonance and molecular dynamics.

    PubMed

    Kordopati, Golfo G; Tzoupis, Haralambos; Troganis, Anastassios N; Tsivgoulis, Gerasimos M; Golic Grdadolnik, Simona; Simal, Carmen; Tselios, Theodore V

    2017-07-29

    Proteolipid protein (PLP) is one of the main proteins of myelin sheath that are destroyed during the progress of multiple sclerosis (MS). The immunodominant PLP139-151 epitope is known to induce experimental autoimmune encephalomyelitis (EAE, animal model of MS), wherein residues 144 and 147 are recognized by T cell receptor (TCR) during the formation of trimolecular complex with peptide-antigen and major histocompability complex. The conformational behavior of linear and cyclic peptide analogues of PLP, namely PLP139-151 and cyclic (139-151) (L(144), R(147)) PLP139-151, have been studied in solution by means of nuclear magnetic resonance (NMR) methods in combination with unrestrained molecular dynamics simulations. The results indicate that the side chains of mutated amino acids in the cyclic analogue have different spatial orientation compared with the corresponding side chains of the linear analogue, which can lead to reduced affinity to TCR. NMR experiments combined with theoretical calculations pave the way for the design and synthesis of potent restricted peptides of immunodominant PLP139-151 epitope as well as non peptide mimetics that rises as an ultimate goal.

  5. Biologically relevant conformational features of linear and cyclic proteolipid protein (PLP) peptide analogues obtained by high-resolution nuclear magnetic resonance and molecular dynamics

    NASA Astrophysics Data System (ADS)

    Kordopati, Golfo G.; Tzoupis, Haralambos; Troganis, Anastassios N.; Tsivgoulis, Gerasimos M.; Golic Grdadolnik, Simona; Simal, Carmen; Tselios, Theodore V.

    2017-07-01

    Proteolipid protein (PLP) is one of the main proteins of myelin sheath that are destroyed during the progress of multiple sclerosis (MS). The immunodominant PLP139-151 epitope is known to induce experimental autoimmune encephalomyelitis (EAE, animal model of MS), wherein residues 144 and 147 are recognized by T cell receptor (TCR) during the formation of trimolecular complex with peptide-antigen and major histocompability complex. The conformational behavior of linear and cyclic peptide analogues of PLP, namely PLP139-151 and cyclic (139-151) (L144, R147) PLP139-151, have been studied in solution by means of nuclear magnetic resonance (NMR) methods in combination with unrestrained molecular dynamics simulations. The results indicate that the side chains of mutated amino acids in the cyclic analogue have different spatial orientation compared with the corresponding side chains of the linear analogue, which can lead to reduced affinity to TCR. NMR experiments combined with theoretical calculations pave the way for the design and synthesis of potent restricted peptides of immunodominant PLP139-151 epitope as well as non peptide mimetics that rises as an ultimate goal.

  6. peptide fibrillar architectures controlled by conformational constraints of the monomer.

    PubMed

    Brännström, Kristoffer; Ohman, Anders; Olofsson, Anders

    2011-01-01

    Anomalous self-assembly of the Aβ peptide into fibrillar amyloid deposits is strongly correlated with the development of Alzheimer's disease. Aβ fibril extension follows a template guided "dock and lock" mechanism where polymerisation is catalysed by the fibrillar ends. Using surface plasmon resonance (SPR) and quenched hydrogen-deuterium exchange NMR (H/D-exchange NMR), we have analysed the fibrillar structure and polymerisation properties of both the highly aggregation prone Aβ1-40 Glu22Gly (Aβ(40Arc)) and wild type Aβ1-40 (Aβ(40WT)). The solvent protection patterns from H/D exchange experiments suggest very similar structures of the fibrillar forms. However, through cross-seeding experiments monitored by SPR, we found that the monomeric form of Aβ(40WT) is significantly impaired to acquire the fibrillar architecture of Aβ(40Arc). A detailed characterisation demonstrated that Aβ(40WT) has a restricted ability to dock and isomerise with high binding affinity onto Aβ(40Arc) fibrils. These results have general implications for the process of fibril assembly, where the rate of polymerisation, and consequently the architecture of the formed fibrils, is restricted by conformational constraints of the monomers. Interestingly, we also found that the kinetic rate of fibril formation rather than the thermodynamically lowest energy state determines the overall fibrillar structure.

  7. Synthesis and structural characterization of monomeric and dimeric peptide nucleic acids prepared by using microwave-promoted multicomponent reactions.

    PubMed

    Ovadia, Reuben; Lebrun, Aurélien; Barvik, Ivan; Vasseur, Jean-Jacques; Baraguey, Carine; Alvarez, Karine

    2015-12-07

    A solution phase synthesis of peptide nucleic acid monomers and dimers was developed by using microwave-promoted Ugi multicomponent reactions. A mixture of a functionalized amine, a carboxymethyl nucleobase, paraformaldehyde and an isocyanide as building blocks generates PNA monomers which are then partially deprotected and used in a second Ugi 4CC reaction, leading to PNA dimers. Conformational rotamers were identified by using NMR and MD simulations.

  8. Histidine-lysine peptides as carriers of nucleic acids.

    PubMed

    Leng, Qixin; Goldgeier, Lisa; Zhu, Jingsong; Cambell, Patricia; Ambulos, Nicholas; Mixson, A James

    2007-03-01

    With their biodegradability and diversity of permutations, peptides have significant potential as carriers of nucleic acids. This review will focus on the sequence and branching patterns of peptide carriers composed primarily of histidines and lysines. While lysines within peptides are important for binding to the negatively charged phosphates, histidines are critical for endosomal lysis enabling nucleic acids to reach the cytosol. Histidine-lysine (HK) polymers by either covalent or ionic bonds with liposomes augment transfection compared to liposome carriers alone. More recently, we have examined peptides as sole carriers of nucleic acids because of their intrinsic advantages compared to the bipartite HK/liposome carriers. With a protocol change and addition of a histidine-rich tail, HK peptides as sole carriers were more effective than liposomes alone in several cell lines. While four-branched polymers with a primary repeating sequence pattern of -HHK- were more effective as carriers of plasmids, eight-branched polymers with a sequence pattern of -HHHK- were more effective as carriers of siRNA. Compared to polyethylenimine, HK carriers of siRNA and plasmids had reduced toxicity. When injected intravenously, HK polymers in complex with plasmids encoding antiangiogenic proteins significantly decreased tumor growth. Furthermore, modification of HK polymers with polyethylene glycol and vascular-specific ligands increased specificity of the polyplex to the tumor by more than 40-fold. Together with further development and insight on the structure of HK polyplexes, HK peptides may prove to be useful as carriers of different forms of nucleic acids both in vitro and in vivo.

  9. Synthesis of Nucleoside N-Phosphoamino Acids and Peptide Formation

    NASA Astrophysics Data System (ADS)

    Lin, Changxue; Fu, Hua; Zhao, Yufen; Cheng, Changmei

    2005-02-01

    Nucleoside N-phosphoamino acids were synthesized through Atherton-Todd reaction of nucleoside H-phosphonate with amino acids, and their structures were confirmed by NMR and ESI-MS. After nucleoside N-phosphoamino acid was incubated in anhydrous methanol at 40 °C for 72 h, di- to tetra-peptide derivatives were detected by ESI-MS, and their structures were further identified by multistage mass spectrometry. These and previously published studies in aqueous solution suggest that nucleoside N-phosphoamino acids could have been prebiotic precursors of oligopeptides.

  10. Conformational Analysis of 3,3,3-TRIFLUORO-2-(TRIFLUOROMETHYL)PROPANOIC Acid

    NASA Astrophysics Data System (ADS)

    Thomas, Javix; Carrillo, Michael J.; Serrato, Agapito, III; Schnitzler, Elijah G.; Jäger, Wolfgang; Xu, Yunjie; Lin, Wei

    2017-06-01

    Partially fluorinated carboxylic acids exhibit rich conformational landscapes. We report the first high-resolution spectroscopic study of 3,3,3-trifluoro-2-(trifluoromethyl)propanoic acid. Its rotational spectrum was measured using both broadband chirped-pulse and narrow-band cavity-based Fourier transform microwave spectrometers. Two dominant conformers were observed, and their structures confirmed with the aid of quantum chemical calculations. Both conformers take on the Z form of the carboxylic acid group. Similarities and differences between this and other fluorinated carboxylic acids are discussed.

  11. Conformational behavior of fragments of adrenocorticotropin and their antisense peptides determined by NMR spectroscopy and CD spectropolarimetry.

    PubMed

    Najem, E S; Corigliano-Murphy, A; Ferretti, J A

    1989-07-03

    An 'antisense' peptide ('HTCA'), whose sequence was generated by reading the antisense RNA sequence corresponding to ACTH (1-24) was shown to bind ACTH (1-24) with a Kd of 0.3 nM in a solid-matrix binding assay [( 1986) Biochem. J. 234, 679 683]. Two-dimensional NMR spectra were used to examine the conformational behavior in methanol and in water solution of two fragments of adrenocorticotropin, ACTH(1-24) and ACTH (1-13), as well as their antisense peptides, HTCA and HTCA(12-24). The conformations are extended chains in these solutions, both as isolated molecules and when mixed with their antisense complements. The Kd values are greater than 1 mM.

  12. Chiral separation of amino acids and peptides by capillary electrophoresis.

    PubMed

    Wan, H; Blomberg, L G

    2000-04-14

    Chiral separation of amino acids and peptides by capillary electrophoresis (CE) is reviewed regarding the separation principles of different approaches, advantages and limitations, chiral recognition mechanisms and applications. The direct approach details various chiral selectors with an emphasis on cyclodextrins and their derivatives, antibiotics and chiral surfactants as the chiral selectors. The indirect approach deals with various chiral reagents applied for diastereomer formation and types of separation media such as micelles and polymeric pseudo-stationary phases. Many derivatization reagents used for high sensitivity detection of amino acids and peptides are also discussed and their characteristics are summarized in tables. A large number of relevant examples is presented illustrating the current status of enantiomeric and diastereomeric separation of amino acids and peptides. Strategies to enhance the selectivity and optimize separation parameters by the application of experimental designs are described. The reversal of enantiomeric elution order and the effects of organic modifiers on the selectivity are illustrated in both direct and indirect methods. Some applications of chiral amino acid and peptide analysis, in particular, regarding the determination of trace enantiomeric impurities, are given. This review selects more than 200 articles published between 1988 and 1999.

  13. Incorporation of 2,3-diaminopropionic acid into linear cationic amphipathic peptides produces pH-sensitive vectors.

    PubMed

    Lan, Yun; Langlet-Bertin, Bérangère; Abbate, Vincenzo; Vermeer, Louic S; Kong, Xiaole; Sullivan, Kelly E; Leborgne, Christian; Scherman, Daniel; Hider, Robert C; Drake, Alex F; Bansal, Sukhvinder S; Kichler, Antoine; Mason, A James

    2010-06-14

    Nonviral vectors that harness the change in pH in endosomes, are increasingly being used to deliver cargoes, including nucleic acids, into mammalian cells. Here we present evidence that the pK(a) of the beta-NH(2) in 2,3-diaminopropionic acid (Dap) is sufficiently lowered, when Dap is incorporated into peptides, that its protonation state is sensitive to the pH changes that occur during endosomal acidification. The lowered pK(a) of around 6.3 is stabilized by the increased electron-withdrawing effect of the peptide bonds, by intermolecular hydrogen bonding and from contributions arising from the peptide conformation. These include mixed polar/apolar environments, Coulombic interactions and intermolecular hydrogen bonding. Changes in the charged state are therefore expected between pH 5 and 7, and large-scale conformational changes are observed in Dap-rich peptides, in contrast to analogues containing lysine or ornithine, when the pH is altered through this range. These physical properties confer a robust gene-delivery capability on designed cationic amphipathic peptides that incorporate Dap.

  14. Synthesis and structural characterization of sialic acid-glutamic acid hybrid foldamers as conformational surrogates of alpha-2,8-linked polysialic acid.

    PubMed

    Saludes, Jonel P; Ames, James B; Gervay-Hague, Jacquelyn

    2009-04-22

    Surface expression of alpha-(2,8)-linked polymers of sialic acid in adult tissues has been correlated with metastasis of several human cancers. One approach to chemotherapeutic intervention against the spread of these cancers involves the development of immunogenic molecules that elicit an antibody response against alpha-(2,8)-linked polysialic acids. Naturally occurring polysialic acids are not viable candidates because they are present during embryonic development and are recognized as self by the immune system. These natural polymers also have poor pharmacokinetic properties because they are readily degraded by neuraminidase enzymes. We have been interested in developing structural surrogates of polysialic acids in an effort to overcome these limitations. Reported herein are microwave-assisted solid-phase peptide syntheses and structural characterization studies of a series of alpha/delta hybrid peptides derived from Fmoc-Neu2en and Fmoc-Glu(OtBu)-OH. Conformational experiments including circular dichroism, NH/ND exchange, and ROESY in aqueous solution were performed to study the secondary structures of these hybrid foldamers. ROESY data were analyzed with the assistance of XPLOR-NIH that was modified to include parameter and topology files to accommodate unnatural amino acids and the delta amide linkages. The results indicate that stable secondary structure is dependent upon both the amino acid sequence and the configuration of Glu. The most stable foldamer was composed of a total of 6 residues beginning with L-Glu at the carboxy terminus and alternating Neu2en and L-Glu residues. In water, this foldamer adopts a right-handed helical conformation with 3.7 residues per turn, 7.4 A pitch, 5.8 A diameter, and a length of 18.5 A, which is stabilized by both classical C=O...H-N backbone interactions and by pyranose ring O and L-Glu HN H-bonding. These structural features orient the L-Glu carboxylates along the helical backbone with a periodicity that matches the

  15. Conformational analysis of short polar side-chain amino-acids through umbrella sampling and DFT calculations.

    PubMed

    Ramos, Javier; Cruz, Victor L

    2016-11-01

    Molecular and quantum mechanics calculations were carried out in a series of tripeptides (GXG, where X = D, N and C) as models of the unfolded states of proteins. The selected central amino acids, especially aspartic acid (D) and asparagine (N) are known to present significant average conformations in partially allowed areas of the Ramachandran plot, which have been suggested to be important in unfolded protein regions. In this report, we present the calculation of the propensity values through an umbrella sampling procedure in combination with the calculation of the NMR J-coupling constants obtained by a DFT model. The experimental NMR observations can be reasonably explained in terms of a conformational distribution where PPII and β basins sum up propensities above 0.9. The conformational analysis of the side chain dihedral angle (χ1), along with the computation of (3)J(H(α)H(β)), revealed a preference for the g - and g + rotamers. These may be connected with the presence of intermolecular H-bonding and carbonyl-carbonyl interactions sampled in the PPII and β basins. Taking into account all those results, it can be established that these residues show a similar behavior to other amino acids in short peptides regarding backbone φ,ψ dihedral angle distribution, in agreement with some experimental analysis of capped dipeptides.

  16. Construction of peptides with nucleobase amino acids: design and synthesis of the nucleobase-conjugated peptides derived from HIV-1 Rev and their binding properties to HIV-1 RRE RNA.

    PubMed

    Takahashi, T; Hamasaki, K; Ueno, A; Mihara, H

    2001-04-01

    In order to develop a novel molecule that recognizes a specific structure of RNA, we have attempted to design peptides having L-alpha-amino acids with a nucleobase at the side chain (nucleobase amino acid (NBA)), expecting that the function of a nucleobase which can specifically recognize a base in RNA is regulated in a peptide conformation. In this study, to demonstrate the applicability of the NBA units in the peptide to RNA recognition, we designed and synthesized a variety of NBA-conjugated peptides, derived from HIV-1 Rev. Circular dichroism study revealed that the conjugation of the Rev peptide with an NBA unit did not disturb the peptide conformation. RNA-binding affinities of the designed peptides with RRE IIB RNA were dependent on the structure of the nucleobase moieties in the peptides. The peptide having the cytosine NBA at the position of the Asn40 site in the Rev showed a higher binding ability for RRE IIB RNA, despite the diminishing the Asn40 function. Furthermore, the peptide having the guanine NBA at the position of the Arg44 site, which is the most important residue for the RNA binding in the Rev, bound to RRE IIB RNA in an ability similar to Rev34-50 with native sequence. These results demonstrate that an appropriate NBA unit in the peptide plays an important role in the RNA binding with a specific contact such as hydrogen bonding, and the interaction between the nucleobase in the peptide and the base in the RNA can enhance the RNA-binding affinity and specificity.

  17. Characteristics of Cell-Penetrating Peptide/Nucleic Acid Nanoparticles.

    PubMed

    Margus, Helerin; Arukuusk, Piret; Langel, Ülo; Pooga, Margus

    2016-01-04

    Nucleic acids are highly promising candidates for the treatment of various genetic diseases. However, due to the large size and negative charge, nucleic acids are not efficiently taken up by cells, and thus, their clinical potential remains limited so far. Therefore, various delivery vehicles have been designed to assist the cellular uptake of nucleic acids. Among these, cell-penetrating peptides (CPPs) have gained increasing popularity as efficient and nontoxic delivery vectors. CPPs can be coupled to nucleic acids either by covalent or noncovalent association. Noncovalent coupling, which is based on the formation of nanoparticle-like nanocomplexes (NP), has received much attention in recent years, and the number of studies employing the strategy is explosively increasing due to the high therapeutic potential. However, the properties of CPP/nucleic acid NPs have not been characterized in sufficient detail yet. We performed a comprehensive analysis of the size and morphology of nucleic acid nanoparticles with novel transfection peptides, PepFects (PFs) and NickFects (NFs), using negative staining transmission electron microscopy (TEM). In addition, we examined whether the attachment of fluorescence or (nano)gold label to nucleic acid affects the nanocomplex formation or its morphology. We demonstrated that transportan-10-based new generation CPPs from PF and NF families condense nucleic acids to NPs of homogeneous size and shape. The size and shape of assembled nanoparticles depend on the type of the complexed nucleic acid and the sequence of the used peptide, whereas the label on the nucleic acid does not influence the gross characteristics of formed NPs.

  18. Exogenous loading of a tapasin-dependent peptide onto HLA-B*44:02 can be restored by acid treatment or fixation of target cells

    PubMed Central

    Stroobant, Vincent; Demotte, Nathalie; Luiten, Rosalie M.; Leonhardt, Ralf M.; Cresswell, Peter; Bonehill, Aude; Michaux, Alexandre; Ma, Wenbin; Mulder, Arend; Van den Eynde, Benoît J.; van der Bruggen, Pierre; Vigneron, Nathalie

    2013-01-01

    Anti-tumor CTLs recognize peptides derived from cellular proteins and presented on MHC class I. One category of peptides recognized by these CTLs is derived from proteins encoded by “cancer-germline” genes, which are specifically expressed in tumors, and therefore represent optimal targets for cancer immunotherapy. Here, we identify an antigenic peptide, which is derived from the MAGE-A1-encoded protein (160-169) and presented to CTLs by HLA-B*44:02. Although this peptide is encoded by MAGE-A1, processed endogenously and presented by tumor cells, the corresponding synthetic peptide is hardly able to sensitize target cells to CTL recognition when pulsed exogenously. Endogenous processing and presentation of this peptide is strictly dependent on the presence of tapasin, which is believed to help peptide loading by stabilizing a peptide-receptive form of HLA-B*44:02. Exogenous loading of the peptide can be dramatically improved by paraformaldehyde fixation of surface molecules or by peptide loading at acidic pH. Either strategy allows efficient exogenous loading of the peptide, presumably by generating or stabilizing a peptide-receptive, empty conformation of the HLA. Altogether, our results indicate a potential drawback of short peptide-based vaccination strategies and offer possible solutions regarding the use of problematic epitopes such as the one described here. PMID:22678898

  19. Screening of peptides with a high affinity to bile acids using peptide arrays and a computational analysis.

    PubMed

    Takeshita, Toshikazu; Okochi, Mina; Kato, Ryuji; Kaga, Chiaki; Tomita, Yasuyuki; Nagaoka, Satoshi; Honda, Hiroyuki

    2011-07-01

    Bile acid binding peptides have attracted attention for the improvement and prevention of hypercholesterolemia. In this study, screening of bile acid high affinity peptides was investigated using computationally-assisted peptide array analysis. Starting with the screening data obtained from a limited, random 6-mer library (2212 sequences), the peptides with a high affinity to bile acid were characterized by comparison of high- and low-affinity peptides using fuzzy neural network (FNN) analysis. The physical properties of amino acids at specific positions that contribute to bile acid binding activity were extracted as the structural rule; optimization was carried out using three repeated screening cycles of the rule extraction. The extracted structural rule indicates that Trp, Tyr, Phe, Leu, Ile and Val are enriched in bile acid binding peptides. The yields of bile acid binding peptides with an affinity of above the VAWWMY peptide (soystatin, control sequence) were significantly higher in the optimized structural rule (32.5%) compared to that of the random library (3.1%), and 6 peptides were obtained with above 2.0-fold increased binding activity. Copyright © 2011 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  20. Prediction of conformational states of amino acids using a Ramachandran plot.

    PubMed

    Kolaskar, A S; Sawant, S

    1996-01-01

    (phi, psi) data from crystal structures of 221 proteins having high resolution and sequence similarity cut-off at the 25% level were analysed by dividing the Ramachandran plot in three regions representing three conformational states: (i) conformational state 1: conformations in the (phi, psi) range from (-140 degrees, -100 degrees) to (0 degrees, 0 degrees); (ii) conformational state 2: conformations with (phi, psi) from (-180 degrees, 80 degrees) to (0 degrees, 180 degrees); and (iii) conformational state 3: all the remaining conformations in the (phi, psi) plane which are not included in the above two conformational states. Normalized probability values of the occurrence of single amino acid residues in conformational regions 1-3 and similar values for dipeptides were calculated. Comparisons of single residue and dipeptide normalized probability values have shown that short-range interactions, although strong, destabilize conformational states of only 44 dipeptides out of the 400 x 9 possible states. However, dipeptide frequency values provide better resolving power than single-residue potentials when used to predict conformational states of residues in a protein from its primary structure. The simple approach used in the present study to predict conformational states yields an accuracy of > 70% for 14 proteins and an accuracy in the range of 50-70% for 247 proteins. Thus these studies point out yet another use of the Ramachandran plot and the role of tertiary interactions in protein folding.

  1. Fourier transform vibrational circular dichroism as a decisive tool for conformational studies of peptides containing tyrosyl residues.

    PubMed

    Borics, Attila; Murphy, Richard F; Lovas, Sándor

    2003-01-01

    Previous UV-circular dichroism (UV-CD) and NMR studies showed that Ac-AAAAAAAEAAKA-NH(2) has an alpha-helical structure in 50% (v/v) aqueous trifluoroethanol. Replacement of Ala(1) to Ala(6) with Tyr results in spectra that show an apparent loss of helicity in the same solvent. This apparent loss of helicity could be attributed to the coupling of the tyrosyl side chain chromophore with the backbone amide. However, such electronic coupling does not affect the vibrational CD (VCD) spectra. The VCD spectra of the peptides with tyrosyl residues were identical to that of the peptide containing no Tyr, which shows the same alpha-helical structure. Because it is now clear that Tyr replacement does not change the backbone conformation of peptides, UV-CD measurements should be complemented by VCD to determine the secondary structure when electronic effects can disturb the UV-CD spectrum of the inherent structure. Copyright 2002 Wiley Periodicals, Inc.

  2. Diffusion and conformation of peptide-functionalized polyphenylene dendrimers studied by fluorescence correlation and 13C NMR spectroscopy.

    PubMed

    Koynov, K; Mihov, G; Mondeshki, M; Moon, C; Spiess, H W; Müllen, K; Butt, H-J; Floudas, G

    2007-05-01

    We report on the combined use of fluorescence correlation spectroscopy (FCS) and 1H and 13C NMR spectroscopy to detect the size and type of peptide secondary structures in a series of poly-Z-L-lysine functionalized polyphenylene dendrimers bearing the fluorescent perylenediimide core in solution. In dilute solution, the size of the molecule as detected from FCS and 1H NMR diffusion measurements matches nicely. We show that FCS is a sensitive probe of the core size as well as of the change in the peptide secondary structure. However, FCS is less sensitive to functionality. A change in the peptide secondary conformation from beta-sheets to alpha-helices detected by 13C NMR spectroscopy gives rise to a steep increase in the hydrodynamic radii for number of residues n > or = 16. Nevertheless, helices are objects of low persistence.

  3. How Amino Acids and Peptides Shaped the RNA World

    PubMed Central

    van der Gulik, Peter T.S.; Speijer, Dave

    2015-01-01

    The “RNA world” hypothesis is seen as one of the main contenders for a viable theory on the origin of life. Relatively small RNAs have catalytic power, RNA is everywhere in present-day life, the ribosome is seen as a ribozyme, and rRNA and tRNA are crucial for modern protein synthesis. However, this view is incomplete at best. The modern protein-RNA ribosome most probably is not a distorted form of a “pure RNA ribosome” evolution started out with. Though the oldest center of the ribosome seems “RNA only”, we cannot conclude from this that it ever functioned in an environment without amino acids and/or peptides. Very small RNAs (versatile and stable due to basepairing) and amino acids, as well as dipeptides, coevolved. Remember, it is the amino group of aminoacylated tRNA that attacks peptidyl-tRNA, destroying the bond between peptide and tRNA. This activity of the amino acid part of aminoacyl-tRNA illustrates the centrality of amino acids in life. With the rise of the “RNA world” view of early life, the pendulum seems to have swung too much towards the ribozymatic part of early biochemistry. The necessary presence and activity of amino acids and peptides is in need of highlighting. In this article, we try to bring the role of the peptide component of early life back into focus. We argue that an RNA world completely independent of amino acids never existed. PMID:25607813

  4. β-Amino acids containing peptides and click-cyclized peptide as β-turn mimics: a comparative study with 'conventional' lactam- and disulfide-bridged hexapeptides.

    PubMed

    Larregola, Maud; Lequin, Olivier; Karoyan, Philippe; Guianvarc'h, Dominique; Lavielle, Solange

    2011-09-01

    The increasing interest in click chemistry and its use to stabilize turn structures led us to compare the propensity for β-turn stabilization of different analogs designed as mimics of the β-turn structure found in tendamistat. The β-turn conformation of linear β-amino acid-containing peptides and triazole-cyclized analogs were compared to 'conventional' lactam- and disulfide-bridged hexapeptide analogs. Their 3D structures and their propensity to fold in β-turns in solution, and for those not structured in solution in the presence of α-amylase, were analyzed by NMR spectroscopy and by restrained molecular dynamics with energy minimization. The linear tetrapeptide Ac-Ser-Trp-Arg-Tyr-NH(2) and both the amide bond-cyclized, c[Pro-Ser-Trp-Arg-Tyr-D-Ala] and the disulfide-bridged, Ac-c[Cys-Ser-Trp-Arg-Tyr-Cys]-NH(2) hexapeptides adopt dominantly in solution a β-turn conformation closely related to the one observed in tendamistat. On the contrary, the β-amino acid-containing peptides such as Ac-(R)-β(3) -hSer-(S)-Trp-(S)-β(3) -hArg-(S)-β(3) -hTyr-NH(2) , and the triazole cyclic peptide, c[Lys-Ser-Trp-Arg-Tyr-βtA]-NH(2) , both specifically designed to mimic this β-turn, do not adopt stable structures in solution and do not show any characteristics of β-turn conformation. However, these unstructured peptides specifically interact in the active site of α-amylase, as shown by TrNOESY and saturation transfer difference NMR experiments performed in the presence of the enzyme, and are displaced by acarbose, a specific α-amylase inhibitor. Thus, in contrast to amide-cyclized or disulfide-bridged hexapeptides, β-amino acid-containing peptides and click-cyclized peptides may not be regarded as β-turn stabilizers, but can be considered as potential β-turn inducers.

  5. Combining conformational sampling and selection to identify the binding mode of zinc-bound amyloid peptides with bifunctional molecules

    NASA Astrophysics Data System (ADS)

    Xu, Liang; Gao, Ke; Bao, Chunyu; Wang, Xicheng

    2012-08-01

    The pathogenesis of Alzheimer's disease (AD) has been suggested to be related with the aggregation of amyloid β (Aβ) peptides. Metal ions (e.g. Cu, Fe, and Zn) are supposed to induce the aggregation of Aβ. Recent development of bifunctional molecules that are capable of interacting with Aβ and chelating biometal ions provides promising therapeutics to AD. However, the molecular mechanism for how Aβ, metal ions, and bifunctional molecules interact with each other is still elusive. In this study, the binding mode of Zn2+-bound Aβ with bifunctional molecules was investigated by the combination of conformational sampling of full-length Aβ peptides using replica exchange molecular dynamics simulations (REMD) and conformational selection using molecular docking and classical MD simulations. We demonstrate that Zn2+-bound Aβ(1-40) and Aβ(1-42) exhibit different conformational ensemble. Both Aβ peptides can adopt various conformations to recognize typical bifunctional molecules with different binding affinities. The bifunctional molecules exhibit their dual functions by first preferentially interfering with hydrophobic residues 17-21 and/or 30-35 of Zn2+-bound Aβ. Additional interactions with residues surrounding Zn2+ could possibly disrupt interactions between Zn2+ and Aβ, which then facilitate these small molecules to chelate Zn2+. The binding free energy calculations further demonstrate that the association of Aβ with bifunctional molecules is driven by enthalpy. Our results provide a feasible approach to understand the recognition mechanism of disordered proteins with small molecules, which could be helpful to the design of novel AD drugs.

  6. Recent Developments in Peptide-Based Nucleic Acid Delivery

    PubMed Central

    Veldhoen, Sandra; Laufer, Sandra D.; Restle, Tobias

    2008-01-01

    Despite the fact that non-viral nucleic acid delivery systems are generally considered to be less efficient than viral vectors, they have gained much interest in recent years due to their superior safety profile compared to their viral counterpart. Among these synthetic vectors are cationic polymers, branched dendrimers, cationic liposomes and cell-penetrating peptides (CPPs). The latter represent an assortment of fairly unrelated sequences essentially characterised by a high content of basic amino acids and a length of 10–30 residues. CPPs are capable of mediating the cellular uptake of hydrophilic macromolecules like peptides and nucleic acids (e.g. siRNAs, aptamers and antisense-oligonucleotides), which are internalised by cells at a very low rate when applied alone. Up to now, numerous sequences have been reported to show cell-penetrating properties and many of them have been used to successfully transport a variety of different cargos into mammalian cells. In recent years, it has become apparent that endocytosis is a major route of internalisation even though the mechanisms underlying the cellular translocation of CPPs are poorly understood and still subject to controversial discussions. In this review, we will summarise the latest developments in peptide-based cellular delivery of nucleic acid cargos. We will discuss different mechanisms of entry, the intracellular fate of the cargo, correlation studies of uptake versus biological activity of the cargo as well as technical problems and pitfalls. PMID:19325804

  7. Dissecting a role of a charge and conformation of Tat2 peptide in allosteric regulation of 20S proteasome.

    PubMed

    Witkowska, Julia; Karpowicz, Przemysław; Gaczynska, Maria; Osmulski, Pawel A; Jankowska, Elżbieta

    2014-08-01

    Proteasome is a 'proteolytic factory' that constitutes an essential part of the ubiquitin-proteasome pathway. The involvement of proteasome in regulation of all major aspects of cellular physiology makes it an attractive drug target. So far, only inhibitors of the proteasome entered the clinic as anti-cancer drugs. However, proteasome regulators may also be useful for treatment of inflammatory and neurodegenerative diseases. We established in our previous studies that the peptide Tat2, comprising the basic domain of HIV-1 Tat protein: R(49) KKRRQRR(56) , supplemented with Q(66) DPI(69) fragment, inhibits the 20S proteasome in a noncompetitive manner. Mechanism of Tat2 likely involves allosteric regulation because it competes with the proteasome natural 11S activator for binding to the enzyme noncatalytic subunits. In this study, we performed alanine walking coupled with biological activity measurements and FTIR and CD spectroscopy to dissect contribution of a charge and conformation of Tat2 to its capability to influence peptidase activity of the proteasome. In solution, Tat2 and most of its analogs with a single Ala substitution preferentially adopted a conformation containing PPII/turn structural motifs. Replacing either Asp10 or two or more adjacent Arg/Lys residues induced a random coil conformation, probably by disrupting ionic interactions responsible for stabilization of the peptides ordered structure. The random coil Tat2 analogs lost their capability to activate the latent 20S proteasome. In contrast, inhibitory properties of the peptides more significantly depended on their positive charge. The data provide valuable clues for the future optimization of the Tat2-based proteasome regulators. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.

  8. D-amino acid substitution enhances the stability of antimicrobial peptide polybia-CP.

    PubMed

    Jia, Fengjing; Wang, Jiayi; Peng, Jinxiu; Zhao, Ping; Kong, Ziqing; Wang, Kairong; Yan, Wenjin; Wang, Rui

    2017-10-01

    With the increasing emergence of resistant microbes toward conventional antimicrobial agents, there is an urgent need for the development of antimicrobial agents with novel action mode. Antimicrobial peptides (AMPs) are believed to be one kind of ideal alternatives. However, AMPs can be easily degraded by protease, which limited their therapeutic use. In the present study, D-amino acid substitution strategy was employed to enhance the stability of polybia-CP. We investigated the stability of peptides against the degradation of trypsin and chymotrypsin by determining the antimicrobial activity or determining the HPLC profile of peptides after incubation with proteases. Our results showed that both the all D-amino acid derivative (D-CP) and partial D-lysine substitution derivative (D-lys-CP) have an improved stability against trypsin and chymotrypsin. Although D-CP takes left-hand α-helical conformation and D-lys-CP loses some α-helical content, both of the D-amino acid-substituted derivatives maintain their parental peptides' membrane active action mode. In addition, D-lys-CP showed a slight weaker antimicrobial activity than polybia-CP, but the hemolytic activity decreased greatly. These results suggest that D-CP and D-lys-CP can offer strategy to improve the property of AMPs and may be leading compounds for the development of novel antimicrobial agents. © The Author 2017. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  9. Stapling mimics noncovalent interactions of γ-carboxyglutamates in conantokins, peptidic antagonists of N-methyl-D-aspartic acid receptors.

    PubMed

    Platt, Randall J; Han, Tiffany S; Green, Brad R; Smith, Misty D; Skalicky, Jack; Gruszczynski, Pawel; White, H Steve; Olivera, Baldomero; Bulaj, Grzegorz; Gajewiak, Joanna

    2012-06-08

    Conantokins are short peptides derived from the venoms of marine cone snails that act as antagonists of the N-methyl-D-aspartate (NMDA) receptor family of excitatory glutamate receptors. These peptides contain γ-carboxyglutamic acid residues typically spaced at i,i+4 and/or i,i+7 intervals, which by chelating divalent cations induce and stabilize helical conformation of the peptide. Introduction of a dicarba bridge (or a staple) can covalently stabilize peptide helicity and improve its pharmacological properties. To test the hypothesis that stapling can effectively replace γ-carboxyglutamic acid residues in stabilizing the helical conformation of conantokins, we designed, synthesized, and characterized several stapled analogs of conantokin G (conG), with varying connectivities in terms of staple length and location along the face of the α-helix. NMR studies confirmed that the ring-closing metathesis reaction yielded a single product with the Z configuration of the olefinic bond. Based on circular dichroism and molecular modeling, the stapled analogs exhibited significantly enhanced helicity compared with the native peptide in a metal-free environment. Stapling i,i+4 was benign with respect to effects on in vitro and in vivo pharmacological properties. One analog, namely conG[11-15,S(i,i+4)S(8)], blocked NR2B-containing NMDA receptors with IC(50) = 0.7 μm and provided significant protection in the 6-Hz psychomotor model of pharmacoresistant epilepsy in mice. Remarkably, unlike native conG, conG[11-15,S(i,i+4)S(8)] produced no behavioral motor toxicity. Our results extend the applications of peptide stapling to helical peptides with extracellular targets and provide a means for engineering conantokins with improved pharmacological properties.

  10. Unnatural amino acid analogues of membrane-active helical peptides with anti-mycobacterial activity and improved stability.

    PubMed

    Khara, Jasmeet Singh; Priestman, Miles; Uhía, Iria; Hamilton, Melissa Shea; Krishnan, Nitya; Wang, Ying; Yang, Yi Yan; Langford, Paul R; Newton, Sandra M; Robertson, Brian D; Ee, Pui Lai Rachel

    2016-08-01

    The emergence of MDR-TB, coupled with shrinking antibiotic pipelines, has increased demands for new antimicrobials with novel mechanisms of action. Antimicrobial peptides have increasingly been explored as promising alternatives to antibiotics, but their inherent poor in vivo stability remains an impediment to their clinical utility. We therefore systematically evaluated unnatural amino acid-modified peptides to design analogues with enhanced anti-mycobacterial activities. Anti-mycobacterial activities were evaluated in vitro and intracellularly against drug-susceptible and MDR isolates of Mycobacterium tuberculosis using MIC, killing efficacy and intracellular growth inhibition studies. Toxicity profiles were assessed against mammalian cells to verify cell selectivity. Anti-mycobacterial mechanisms were investigated using microfluidic live-cell imaging with time-lapse fluorescence microscopy and confocal laser-scanning microscopy. Unnatural amino acid incorporation was well tolerated without an appreciable effect on toxicity profiles and secondary conformations of the synthetic peptides. The modified peptides also withstood proteolytic digestion by trypsin. The all d-amino acid peptide, i(llkk)2i (II-D), displayed superior activity against all six mycobacterial strains tested, with a 4-fold increase in selectivity index as compared with the unmodified l-amino acid peptide in broth. II-D effectively reduced the intracellular bacterial burden of both drug-susceptible and MDR clinical isolates of M. tuberculosis after 4 days of treatment. Live-cell imaging studies demonstrated that II-D permeabilizes the mycobacterial membrane, while confocal microscopy revealed that II-D not only permeates the cell membrane, but also accumulates within the cytoplasm. Unnatural amino acid modifications not only decreased the susceptibility of peptides to proteases, but also enhanced mycobacterial selectivity. © The Author 2016. Published by Oxford University Press on behalf of the

  11. The Role of the Local Conformation of a Cyclically Constrained β-AMINO Acid in the Secondary Structures of a Mixed α/β Diastereomer Pair

    NASA Astrophysics Data System (ADS)

    Blodgett, Karl N.; Zwier, Timothy S.

    2017-06-01

    Synthetic foldamers are non-natural polymers designed to fold into unique secondary structures that either mimic nature's preferred secondary structures, or expand their possibilities. Among the most studied synthetic foldamers are β-peptides, which lengthen the distance between amide groups from the single substituted carbon spacer in α-peptides by one (β) additional carbon. Cyclically constrained β-amino acids can impart rigidity to the secondary structure of oligomers by locking in a particular conformation. The β-residue cis-2-aminocyclohexanecarboxylic acid (cis-ACHC) is one such amino acid which has been shown to drive vastly different secondary structures as a function of the local conformation of the cyclohexane ring. We present data on two diastereomers of the mixed α/β tri-peptide Ac-Ala-β_{ACHC}-Ala-NHBn which differ from one another by the chirality along the ACHC residue (SRSS vs. SSRS). The first oligomer is known to crystallize to a 9/11 mixed helix while the second forms no intramolecular hydrogen bonds in the crystal state. This talk will describe the conformation-specific IR and UV spectroscopy of the above two diastereomers under jet cooled conditions in the gas phase. Assignments based on comparison with calculations show the presence of incipient 9/11 mixed helices and competing structures containing more tightly folded hydrogen-bonded networks. The calculated global minimum structures are observed in each case, and in each case these folded structures are reminiscent of a β-turn.

  12. Determining the Conformation of an Adsorbed Br-PEG-Peptide by Long Period X-ray Standing Wave Fluorescence

    PubMed Central

    Crot, Carrie A.; Wu, Chunping; Schlossman, Mark L.; Trainor, Thomas P.; Eng, Peter J.; Hanley, Luke

    2008-01-01

    Long period x-ray standing wave fluorescence (XSW) and x-ray reflectivity techniques are employed to probe the conformation of a Br-polyethylene glycol (PEG)-peptide adsorbate at the hydrated interface of a polystyrene substrate. The Br atom on this Br-PEG-peptide construct serves as a marker atom allowing determination by XSW of its position and distribution with respect to the adsorption surface with angstrom resolution. Adsorption occurs on native or ion beam modified polystyrene films that are spin coated onto a Si substrate and display either nonpolar or polar surfaces, respectively. A compact, oriented monolayer of Br-PEG-peptide can be formed with the peptide end adsorbed onto the polar surface and the PEG end terminating with the Br tag extending into the aqueous phase. The 108 – 141 Å distance of the Br atom from the polystyrene surface in this oriented monolayer is similar to the estimated ~150 Å length of the extended Br-PEG-peptide. This Br-polystyrene distance depends upon adsorption time and surface properties prior to adsorption. Incomplete multilayers form on the polar surface after sufficient adsorption time elapses. By contrast, adsorption onto the nonpolar surface is submonolayer, patchy, and highly disordered with an isotropic Br distribution. Overall, this combination of x-ray surface scattering techniques with a novel sample preparation strategy has several advantages as a real space probe of adsorbed or covalently bound biomolecules at the liquid-solid interface. PMID:16089398

  13. Purification of polyclonal anti-conformational antibodies for use in affinity selection from random peptide phage display libraries: A study using the hydatid vaccine EG95

    PubMed Central

    Read, A.J.; Gauci, C.G.; Lightowlers, M.W.

    2009-01-01

    The use of polyclonal antibodies to screen random peptide phage display libraries often results in the recognition of a large number of peptides that mimic linear epitopes on various proteins. There appears to be a bias in the use of this technology toward the selection of peptides that mimic linear epitopes. In many circumstances the correct folding of a protein immunogen is required for conferring protection. The use of random peptide phage display libraries to identify peptide mimics of conformational epitopes in these cases requires a strategy for overcoming this bias. Conformational epitopes on the hydatid vaccine EG95 have been shown to result in protective immunity in sheep, whereas linear epitopes are not protective. In this paper we describe a strategy that results in the purification of polyclonal antibodies directed against conformational epitopes while eliminating antibodies directed against linear epitopes. These affinity purified antibodies were then used to select a peptide from a random peptide phage display library that has the capacity to mimic conformational epitopes on EG95. This peptide was subsequently used to affinity purify monospecific antibodies against EG95. PMID:19349218

  14. Cholesterol accelerates the binding of Alzheimer's β-amyloid peptide to ganglioside GM1 through a universal hydrogen-bond-dependent sterol tuning of glycolipid conformation

    PubMed Central

    Fantini, Jacques; Yahi, Nouara; Garmy, Nicolas

    2013-01-01

    Age-related alterations of membrane lipids in brain cell membranes together with high blood cholesterol are considered as major risk factors for Alzheimer's disease. Yet the molecular mechanisms by which these factors increase Alzheimer's risk are mostly unknown. In lipid raft domains of the plasma membrane, neurotoxic Alzheimer's beta-amyloid (Abeta) peptides interact with both cholesterol and ganglioside GM1. Recent data also suggested that cholesterol could stimulate the binding of Abeta to GM1 through conformational modulation of the ganglioside headgroup. Here we used a combination of physicochemical and molecular modeling approaches to decipher the mechanisms of cholesterol-assisted binding of Abeta to GM1. With the aim of decoupling the effect of cholesterol on GM1 from direct Abeta-cholesterol interactions, we designed a minimal peptide (Abeta5-16) containing the GM1-binding domain but lacking the amino acid residues involved in cholesterol recognition. Using the Langmuir technique, we showed that cholesterol (but not phosphatidylcholine or sphingomyelin) significantly accelerates the interaction of Abeta5-16 with GM1. Molecular dynamics simulations suggested that Abeta5-16 interacts with a cholesterol-stabilized dimer of GM1. The main structural effect of cholesterol is to establish a hydrogen-bond between its own OH group and the glycosidic-bond linking ceramide to the glycone part of GM1, thereby inducing a tilt in the glycolipid headgroup. This fine conformational tuning stabilizes the active conformation of the GM1 dimer whose headgroups, oriented in two opposite directions, form a chalice-shaped receptacle for Abeta. These data give new mechanistic insights into the stimulatory effect of cholesterol on Abeta/GM1 interactions. They also support the emerging concept that cholesterol is a universal modulator of protein-glycolipid interactions in the broader context of membrane recognition processes. PMID:23772214

  15. Cholesterol accelerates the binding of Alzheimer's β-amyloid peptide to ganglioside GM1 through a universal hydrogen-bond-dependent sterol tuning of glycolipid conformation.

    PubMed

    Fantini, Jacques; Yahi, Nouara; Garmy, Nicolas

    2013-01-01

    Age-related alterations of membrane lipids in brain cell membranes together with high blood cholesterol are considered as major risk factors for Alzheimer's disease. Yet the molecular mechanisms by which these factors increase Alzheimer's risk are mostly unknown. In lipid raft domains of the plasma membrane, neurotoxic Alzheimer's beta-amyloid (Abeta) peptides interact with both cholesterol and ganglioside GM1. Recent data also suggested that cholesterol could stimulate the binding of Abeta to GM1 through conformational modulation of the ganglioside headgroup. Here we used a combination of physicochemical and molecular modeling approaches to decipher the mechanisms of cholesterol-assisted binding of Abeta to GM1. With the aim of decoupling the effect of cholesterol on GM1 from direct Abeta-cholesterol interactions, we designed a minimal peptide (Abeta5-16) containing the GM1-binding domain but lacking the amino acid residues involved in cholesterol recognition. Using the Langmuir technique, we showed that cholesterol (but not phosphatidylcholine or sphingomyelin) significantly accelerates the interaction of Abeta5-16 with GM1. Molecular dynamics simulations suggested that Abeta5-16 interacts with a cholesterol-stabilized dimer of GM1. The main structural effect of cholesterol is to establish a hydrogen-bond between its own OH group and the glycosidic-bond linking ceramide to the glycone part of GM1, thereby inducing a tilt in the glycolipid headgroup. This fine conformational tuning stabilizes the active conformation of the GM1 dimer whose headgroups, oriented in two opposite directions, form a chalice-shaped receptacle for Abeta. These data give new mechanistic insights into the stimulatory effect of cholesterol on Abeta/GM1 interactions. They also support the emerging concept that cholesterol is a universal modulator of protein-glycolipid interactions in the broader context of membrane recognition processes.

  16. Structural Analysis of a β-Helical Protein Motif Stabilized by Targeted Replacements with Conformationally Constrained Amino Acids

    PubMed Central

    Ballano, Gema; Zanuy, David; Jiménez, Ana I.; Cativiela, Carlos; Nussinov, Ruth; Alemán, Carlos

    2009-01-01

    Here we study conformational stabilization induced in a β-helical nanostructure by position-specific mutations. The nanostructure is constructed through the self-assembly of the β-helical building block excised from E. coli galactoside acetyltransferase (PDB code 1krr, chain A; residues 131-165). The mutations involve substitutions by cyclic, conformationally constrained amino acids. Specifically, a complete structural analysis of the Pro-Xaa-Val sequence [with Xaa being Gly, Ac3c (1-aminocyclopropane-1-carboxylic acid) and Ac5c (1-aminocyclopentane-1-carboxylic acid)], corresponding to the 148-150 loop region in the wild-type (Gly) and mutated (Ac3c and Ac5c) 1krr, has been performed using Molecular Dynamics simulations and X-ray crystallography. Simulations have been performed for the wild-type and mutants of three different systems, namely the building block, the nanoconstruct and the isolated Pro-Xaa-Val tripeptide. Furthermore, the crystalline structures of five peptides of Pro-Xaa-Val or Xaa-Val sequences have been solved by X-ray diffraction analysis and compared with theoretical predictions. Both the theoretical and crystallographic studies indicate that the Pro-Acnc-Val sequences exhibit a high propensity to adopt turn-like conformations, and this propensity is little affected by the chemical environment. Overall, the results indicate that replacement of Gly149 by Ac3c or Ac5c significantly reduce the conformational flexibility of the target site enhancing the structural specificity of the building block and the nanoconstruct derived from the 1krr β-helical motif. PMID:18811190

  17. Comprehensive Analysis of Contributions from Protein Conformational Stability and Major Histocompatibility Complex Class II-Peptide Binding Affinity to CD4+ Epitope Immunogenicity in HIV-1 Envelope Glycoprotein

    PubMed Central

    Li, Tingfeng; Steede, N. Kalaya; Nguyen, Hong-Nam P.; Freytag, Lucy C.; McLachlan, James B.; Mettu, Ramgopal R.; Robinson, James E.

    2014-01-01

    ABSTRACT Helper T-cell epitope dominance in human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein gp120 is not adequately explained by peptide binding to major histocompatibility complex (MHC) proteins. Antigen processing potentially influences epitope dominance, but few, if any, studies have attempted to reconcile the influences of antigen processing and MHC protein binding for all helper T-cell epitopes of an antigen. Epitopes of gp120 identified in both humans and mice occur on the C-terminal flanks of flexible segments that are likely to be proteolytic cleavage sites. In this study, the influence of gp120 conformation on the dominance pattern in gp120 from HIV strain 89.6 was examined in CBA mice, whose MHC class II protein has one of the most well defined peptide-binding preferences. Only one of six dominant epitopes contained the most conserved element of the I-Ak binding motif, an aspartic acid. Destabilization of the gp120 conformation by deletion of single disulfide bonds preferentially enhanced responses to the cryptic I-Ak motif-containing sequences, as reported by T-cell proliferation or cytokine secretion. Conversely, inclusion of CpG in the adjuvant with gp120 enhanced responses to the dominant CD4+ T-cell epitopes. The gp120 destabilization affected secretion of some cytokines more than others, suggesting that antigen conformation could modulate T-cell functions through mechanisms of antigen processing. IMPORTANCE CD4+ helper T cells play an essential role in protection against HIV and other pathogens. Thus, the sites of helper T-cell recognition, the dominant epitopes, are targets for vaccine design; and the corresponding T cells may provide markers for monitoring infection and immunity. However, T-cell epitopes are difficult to identify and predict. It is also unclear whether CD4+ T cells specific for one epitope are more protective than T cells specific for other epitopes. This work shows that the three-dimensional (3D) structure of an

  18. Peptide-binding dependent conformational changes regulate the transcriptional activity of the quorum-sensor NprR.

    PubMed

    Zouhir, Samira; Perchat, Stéphane; Nicaise, Magali; Perez, Javier; Guimaraes, Beatriz; Lereclus, Didier; Nessler, Sylvie

    2013-09-01

    The transcriptional regulator NprR controls the expression of genes essential for the adaptative response of Bacillus cereus. NprR belongs to the RNPP family of directly regulated quorum sensors from Gram-positive bacteria. It is activated by the re-imported signaling peptide NprX. To elucidate the activation mechanism of this quorum-sensing system, we analyzed the conformation changes induced on binding of NprX. We solved the crystal structure of the NprR/NprX binary complex and characterized the apo form of NprR in solution. We demonstrated that apo NprR is a dimer that switches to a tetramer in the presence of NprX. Mutagenesis, and functional analysis allowed us to identify the protein and peptide residues directly involved in the NprR activation process. Based on the comparison with the Rap proteins, we propose a model for the peptide-induced conformational change allowing the apo dimer to switch to an active tetramer specifically recognizing target DNA sequences.

  19. Synthesis of a conformationally constrained δ-amino acid building block.

    PubMed

    O'Reilly, Elaine; Pes, Lara; Ortin, Yannick; Müller-Bunz, Helge; Paradisi, Francesca

    2013-02-01

    Conformationally restricted amino acids are important components in peptidomimetics and drug design. Herein, we describe the synthesis of a novel, non-proteinogenic constrained delta amino acid containing a cyclobutane ring, cis-3(aminomethyl)cyclobutane carboxylic acid (ACCA). The synthesis of the target amino acid was achieved in seven steps, with the key reaction being a base induced intramolecular nucleophilic substitution. A small library of dipeptides was prepared through the coupling of ACCA with proteinogenic amino acids.

  20. The bioactive acidic serine- and aspartate-rich motif peptide.

    PubMed

    Minamizaki, Tomoko; Yoshiko, Yuji

    2015-01-01

    The organic component of the bone matrix comprises 40% dry weight of bone. The organic component is mostly composed of type I collagen and small amounts of non-collagenous proteins (NCPs) (10-15% of the total bone protein content). The small integrin-binding ligand N-linked glycoprotein (SIBLING) family, a NCP, is considered to play a key role in bone mineralization. SIBLING family of proteins share common structural features and includes the arginine-glycine-aspartic acid (RGD) motif and acidic serine- and aspartic acid-rich motif (ASARM). Clinical manifestations of gene mutations and/or genetically modified mice indicate that SIBLINGs play diverse roles in bone and extraskeletal tissues. ASARM peptides might not be primary responsible for the functional diversity of SIBLINGs, but this motif is suggested to be a key domain of SIBLINGs. However, the exact function of ASARM peptides is poorly understood. In this article, we discuss the considerable progress made in understanding the role of ASARM as a bioactive peptide.

  1. Structure Analysis and Conformational Transitions of the Cell Penetrating Peptide Transportan 10 in the Membrane-Bound State

    PubMed Central

    Strandberg, Erik; Verdurmen, Wouter P. R.; Bürck, Jochen; Ehni, Sebastian; Mykhailiuk, Pavel K.; Afonin, Sergii; Gerthsen, Dagmar; Komarov, Igor V.; Brock, Roland; Ulrich, Anne S.

    2014-01-01

    Structure analysis of the cell-penetrating peptide transportan 10 (TP10) revealed an exemplary range of different conformations in the membrane-bound state. The bipartite peptide (derived N-terminally from galanin and C-terminally from mastoparan) was found to exhibit prominent characteristics of (i) amphiphilic α-helices, (ii) intrinsically disordered peptides, as well as (iii) β-pleated amyloid fibrils, and these conformational states become interconverted as a function of concentration. We used a complementary approach of solid-state 19F-NMR and circular dichroism in oriented membrane samples to characterize the structural and dynamical behaviour of TP10 in its monomeric and aggregated forms. Nine different positions in the peptide were selectively substituted with either the L- or D-enantiomer of 3-(trifluoromethyl)-bicyclopent-[1.1.1]-1-ylglycine (CF3-Bpg) as a reporter group for 19F-NMR. Using the L-epimeric analogs, a comprehensive three-dimensional structure analysis was carried out in lipid bilayers at low peptide concentration, where TP10 is monomeric. While the N-terminal region is flexible and intrinsically unstructured within the plane of the lipid bilayer, the C-terminal α-helix is embedded in the membrane with an oblique tilt angle of ∼55° and in accordance with its amphiphilic profile. Incorporation of the sterically obstructive D-CF3-Bpg reporter group into the helical region leads to a local unfolding of the membrane-bound peptide. At high concentration, these helix-destabilizing C-terminal substitutions promote aggregation into immobile β-sheets, which resemble amyloid fibrils. On the other hand, the obstructive D-CF3-Bpg substitutions can be accommodated in the flexible N-terminus of TP10 where they do not promote aggregation at high concentration. The cross-talk between the two regions of TP10 thus exerts a delicate balance on its conformational switch, as the presence of the α-helix counteracts the tendency of the unfolded N

  2. Theoretical study of γ-aminobutyric acid conformers: Intramolecular interactions and ionization energies

    NASA Astrophysics Data System (ADS)

    Wang, Ke-Dong; Wang, Mei-Ting; Meng, Ju

    2014-10-01

    Allowing for all combinations of internal single-bond rotamers, 1,296 unique trial structures of γ-Aminobutyric acid (GABA) are obtained. All of these structures are optimized at the M06-2X level of theory and a total of 68 local minimal conformers are found. The nine low-lying conformers are used for further studies. According to the calculated relative Gibbs free energies at M06-2X level of theory, we find that the dispersion is important for the relative energy of GABA. The intramolecular hydrogen bonds and hyperconjugative interaction and their effects on the conformational stability are studied. The results show that both of them have great influence on the conformers. The vertical ionization energies (VIE) are calculated and match the experimental data well. The results show that the neutral GABA in the gas phase is a multi-conformer system and at least four conformations exist.

  3. Conformational equilibrium of phenylacetic acid and its halogenated analogues through theoretical studies, NMR and IR spectroscopy

    NASA Astrophysics Data System (ADS)

    Levandowski, Mariana N.; Rozada, Thiago C.; Melo, Ulisses Z.; Basso, Ernani A.; Fiorin, Barbara C.

    2017-03-01

    This paper presents a study on the conformational preferences of phenylacetic acid (PA) and its halogenated analogues (FPA, CPA, BPA). To clarify the effects that rule these molecules' behaviour, theoretical calculations were used, for both the isolated phase and solution, combined with nuclear magnetic resonance (NMR) and infrared (IR) spectroscopy. Most conformations of phenylacetic acid and its halogenated derivatives are stabilized through the hyperconjugative effect, which rules the conformational preference. NMR analyses showed that even with the variation in medium polarity, there was no significant change in the conformation population. Infrared spectroscopy showed similar results for all compounds under study. In most spectra, two bands were found through the carbonyl deconvolution, which is in accordance with the theoretical data. It was possible to prove that variation in the nature of the substituent in the ortho position had no significant influence on the conformational equilibrium.

  4. Determination of conformation and orientation of immobilized peptides and proteins at buried interfaces

    NASA Astrophysics Data System (ADS)

    Shen, Lei; Ulrich, Nathan W.; Mello, Charlene M.; Chen, Zhan

    2015-01-01

    Surface immobilized peptides/proteins have important applications such as antimicrobial coating and biosensing. We report a study of such peptides/proteins using sum frequency generation vibrational spectroscopy and ATR-FTIR. Immobilization on surfaces via physical adsorption and chemical coupling revealed that structures of chemically immobilized peptides are determined by immobilization sites, chemical environments, and substrate surfaces. In addition, controlling enzyme orientation by engineering the surface immobilization site demonstrated that structures can be well-correlated to measured chemical activity. This research facilitates the development of immobilized peptides/proteins with improved activities by optimizing their surface orientation and structure.

  5. Conformations, energies, and intramolecular hydrogen bonds in dicarboxylic acids: implications for the design of synthetic dicarboxylic acid receptors.

    PubMed

    Nguyen, Thanh Ha; Hibbs, David E; Howard, Siân T

    2005-09-01

    The various conformers of the dicarboxylic acids HO2C--(CH2)n--CO2H, n = 1-4, were obtained using density functional methods (DFT), both in the gas phase and in the aqueous phase using a polarized continuum model (PCM). Several new conformers were identified, particularly for the two larger molecules glutaric (n = 3) and adipic acid (n =4). The PCM results show that the stability of most conformers were affected, many becoming unstable in the aqueous phase; and the energy ordering of conformers is also different. The results suggest that conformational preferences could be important in determining the design and stability of appropriate synthetic receptors for glutaric and adipic acid. Geometry changes between gas and aqueous phases were most marked in those conformers containing an intramolecular hydrogen bond. Additional calculations have probed the strength of intramolecular hydrogen bonds in these dicarboxylic acids. In the cases of glutaric and adipic acid, the strength of the intramolecular hydrogen bond were estimated to be around 28-29 kJ/mol, without any vibrational energy correction. The intramolecular hydrogen bond energies in malonic and succinic acid were also estimated from the calculated H-bond distances using an empirical relationship. Intramolecular H-bond redshifts of 170-250 cm(-1) have been estimated from the results of the harmonic frequency analyses.

  6. Interrelationships among biological activity, disulfide bonds, secondary structure, and metal ion binding for a chemically synthesized 34-amino-acid peptide derived from alpha-fetoprotein.

    PubMed

    MacColl, R; Eisele, L E; Stack, R F; Hauer, C; Vakharia, D D; Benno, A; Kelly, W C; Mizejewski, G J

    2001-10-03

    A 34-amino-acid peptide has been chemically synthesized based on a sequence from human alpha-fetoprotein. The purified peptide is active in anti-growth assays when freshly prepared in pH 7.4 buffer at 0.20 g/l, but this peptide slowly becomes inactive. This functional change is proven by mass spectrometry to be triggered by the formation of an intrapeptide disulfide bond between the two cysteine residues on the peptide. Interpeptide cross-linking does not occur. The active and inactive forms of the peptide have almost identical secondary structures as shown by circular dichroism (CD). Zinc ions bind to the active peptide and completely prevents formation of the inactive form. Cobalt(II) ions also bind to the peptide, and the UV-Vis absorption spectrum of the cobalt-peptide complex shows that: (1) a near-UV sulfur-to-metal-ion charge-transfer band had a molar extinction coefficient consistent with two thiolate bonds to Co(II); (2) the lowest-energy visible d-d transition maximum at 659 nm, also, demonstrated that the two cysteine residues are ligands for the metal ion; (3) the d-d molar extinction coefficient showed that the metal ion-ligand complex was in a distorted tetrahedral symmetry. The peptide has two cysteines, and it is speculated that the other two metal ion ligands might be the two histidines. The Zn(II)- and Co(II)-peptide complexes had similar peptide conformations as indicated by their ultraviolet CD spectra, which differed very slightly from that of the free peptide. Surprisingly, the cobalt ions acted in the reverse of the zinc ions in that, instead of stabilizing anti-growth form of the peptide, they catalyzed its loss. Metal ion control of peptide function is a saliently interesting concept. Calcium ions, in the conditions studied, apparently do not bind to the peptide. Trifluoroethanol and temperature (60 degrees C) affected the secondary structure of the peptide, and the peptide was found capable of assuming various conformations in solution

  7. Biophysical and morphological studies on the dual interaction of non-octarepeat prion protein peptides with copper and nucleic acids.

    PubMed

    Chaves, Juliana A P; Sanchez-López, Carolina; Gomes, Mariana P B; Sisnande, Tháyna; Macedo, Bruno; de Oliveira, Vanessa End; Braga, Carolina A C; Rangel, Luciana P; Silva, Jerson L; Quintanar, Liliana; Cordeiro, Yraima

    2014-08-01

    Conversion of prion protein (PrP) to an altered conformer, the scrapie PrP (PrP(Sc)), is a critical step in the development of transmissible spongiform encephalopathies. Both Cu(II) and nucleic acid molecules have been implicated in this conversion. Full-length PrP can bind up to six copper ions; four Cu(II) binding sites are located in the octarepeat domain (residues 60-91), and His-96 and His-111 coordinate two additional copper ions. Experimental evidence shows that PrP binds different molecules, resulting in diverse cellular signaling events. However, there is little information about the interaction of macromolecular ligands with Cu(II)-bound PrP. Both RNA and DNA sequences can bind PrP, and this interaction results in reciprocal conformational changes. Here, we investigated the interaction of Cu(II) and nucleic acids with amyloidogenic non-octarepeat PrP peptide models (comprising human PrP residues 106-126 and hamster PrP residues 109-149) that retain His-111 as the copper-anchoring residue. The effect of Cu(II) and DNA or RNA sequences in the aggregation, conformation, and toxicity of PrP domains was investigated at low and neutral pH. Circular dichroism and EPR spectroscopy data indicate that interaction of the PrP peptides with Cu(II) and DNA occurs at pH 7. This dual interaction induces conformational changes in the peptides, modulating their aggregation, and affecting the morphology of the aggregated species, resulting in different cytotoxic effects. These results provide new insights into the role of Cu(II) and nucleic acid sequences in the structural conversion and aggregation of PrP, which are both critical events related to prion pathogenesis.

  8. Characterization of the HIV N-terminal fusion peptide-containing region in context of key gp41 fusion conformations.

    PubMed

    Sackett, Kelly; Wexler-Cohen, Yael; Shai, Yechiel

    2006-08-04

    Central to our understanding of human immunodeficiency virus-induced fusion is the high resolution structure of fragments of the gp41 fusion protein folded in a low energy core conformation. However, regions fundamental to fusion, like the fusion peptide (FP), have yet to be characterized in the context of the cognate protein regardless of its conformation. Based on conformation-specific monoclonal antibody recognition, we identified the polar region consecutive to the N36 fragment as a stabilizer of trimeric coiled-coil assembly, thereby enhancing inhibitory potency. This tertiary organization is retained in the context of the hydrophobic FP (N70 fragment). Our data indicate that the N70 fragment recapitulates the expected organization of this region in the viral fusion intermediate (N-terminal half of the pre-hairpin intermediate (N-PHI)), which happens to be the prime target for fusion inhibitors. Regarding the low energy conformation, we show for the first time core formation in the context of the FP (N70 core). The alpha-helical and coiled-coil stabilizing polar region confers substantial thermal stability to the core, whereas the hydrophobic FP does not add further stability. For the two key fusion conformations, N-PHI and N70 core, we find that the FP adopts a nonhelical structure and directs higher order assembly (assembly of coiled coils in N-PHI and assembly of bundles in the N70 core). This supra-molecular organization of coiled coils or folded cores is seen only in the context of the FP. This study is the first to characterize the FP region in the context of the folded core and provides a basic understanding of the role of the elusive FP for key gp41 fusion conformations.

  9. Synthesis, conformational analysis, and biological activities of cyclic enkephalins and model cyclic peptides containing thioamides as amide bond replacements

    SciTech Connect

    Sherman, D.B.

    1988-01-01

    This thesis describes the first applications of the thioamide surrogate ({psi}(CSNH)) in model cyclic peptides and cyclic enkephalins. The solution phase synthesis and conformational analysis of two model cyclic endothiopentapeptides, cyclo(D-Phe-Pro{psi} (CSNH)Gly-Pro-Gly) (I) and cyclo(D-Phe-Pro-Gly-Pro{psi}(CSNH)Gly) (II), are reported. The conformations of I and II were analyzed using several 1 and 2D NMR techniques, such as INDOR, temperature and concentration dependence, {sup 1}H-{sup 1}H and {sup 1}H-{sup 13}C COSY, ROESY, and magnetization transfer. Compound I displayed the same general conformation as its parent in CDCl{sub 3}, but the {gamma}-turn hydrogen bond appeared to be weaker, while the {beta}-turn hydrogen bond appeared to be stronger. In DMSO-d{sub 6}, this molecule existed in two conformations in a ratio of 2:1. The major conformer appeared to be the same as that in CDCl{sub 3}, while the second contained at least one cis X-Pro bond, most likely at the Gly{sup 1}-Pro{sup 2} position. Compound II displayed the same conformation as its parent in both solvents. The {gamma}-turn hydrogen bond again appeared to be weaker in CDCl{sub 3}, but comparable with the parent in DMSO-d{sub 6}. Molecular modeling studies of I and II indicated the Pro {psi} angle increased by {approx}6{degree} when a thiocarbonyl was present, thereby reducing steric interactions with the Pro {beta} methylene.

  10. Conformational stability of digestion-resistant peptides of peanut conglutins reveals the molecular basis of their allergenicity

    PubMed Central

    Apostolovic, Danijela; Stanic-Vucinic, Dragana; de Jongh, Harmen H. J.; de Jong, Govardus A. H.; Mihailovic, Jelena; Radosavljevic, Jelena; Radibratovic, Milica; Nordlee, Julie A.; Baumert, Joseph L.; Milcic, Milos; Taylor, Steve L.; Garrido Clua, Nuria; Cirkovic Velickovic, Tanja; Koppelman, Stef J.

    2016-01-01

    Conglutins represent the major peanut allergens and are renowned for their resistance to gastro-intestinal digestion. Our aim was to characterize the digestion-resistant peptides (DRPs) of conglutins by biochemical and biophysical methods followed by a molecular dynamics simulation in order to better understand the molecular basis of food protein allergenicity. We have mapped proteolysis sites at the N- and C-termini and at a limited internal segment, while other potential proteolysis sites remained unaffected. Molecular dynamics simulation showed that proteolysis only occurred in the vibrant regions of the proteins. DRPs appeared to be conformationally stable as intact conglutins. Also, the overall secondary structure and IgE-binding potency of DRPs was comparable to that of intact conglutins. The stability of conglutins toward gastro-intestinal digestion, combined with the conformational stability of the resulting DRPs provide conditions for optimal exposure to the intestinal immune system, providing an explanation for the extraordinary allergenicity of peanut conglutins. PMID:27377129

  11. Peptide bond formation does not involve acid-base catalysis by ribosomal residues.

    PubMed

    Bieling, Peter; Beringer, Malte; Adio, Sarah; Rodnina, Marina V

    2006-05-01

    Ribosomes catalyze the formation of peptide bonds between aminoacyl esters of transfer RNAs within a catalytic center composed of ribosomal RNA only. Here we show that the reaction of P-site formylmethionine (fMet)-tRNA(fMet) with a modified A-site tRNA substrate, Phelac-tRNA(Phe), in which the nucleophilic amino group is replaced with a hydroxyl group, does not show the pH dependence observed with small substrate analogs such as puromycin and hydroxypuromycin. This indicates that acid-base catalysis by ribosomal residues is not important in the reaction with the full-size substrate. Rather, the ribosome catalyzes peptide bond formation by positioning the tRNAs, or their 3' termini, through interactions with rRNA that induce and/or stabilize a pH-insensitive conformation of the active site and provide a preorganized environment facilitating the reaction. The rate of peptide bond formation with unmodified Phe-tRNA(Phe) is estimated to be >300 s(-1).

  12. Conformational studies of peptides corresponding to the coeliac-activating regions of wheat alpha-gliadin.

    PubMed Central

    Tatham, A S; Marsh, M N; Wieser, H; Shewry, P R

    1990-01-01

    The structures of four peptides corresponding to parts of the coeliac-activating protein A-gliadin were studied by structure prediction and c.d. spectroscopy. Three of the peptides corresponded to parts of the coeliac-activating N-terminal region (residues 3-55, 3-19 and 39-45) and contained two tetrapeptide motifs common to all coeliac-active regions (Pro-Ser-Gln-Gln and Gln-Gln-Gln-Pro). The Pro-Ser-Gln-Gln sequence was also present in the fourth peptide, on the basis of the C-terminal part of the molecule (211-217). These studies showed that beta-reverse turns were the predominant structural feature in all peptides and were predominantly of type I/III in two of the N-terminal peptides and type II in the C-terminal peptide. These turns form when the peptide is dissolved in solvents of low dielectric constant (trifluoroethanol) and high dielectric constant (water and iso-osmotic saline), although their presence in the N-terminal peptides may be masked in the latter solvents due to equilibrium with a poly-L-proline II structure favoured at lower temperatures. PMID:2400392

  13. A conformational study of the opioid peptide dermorphin by one-dimensional and two-dimensional nuclear magnetic resonance spectroscopy.

    PubMed Central

    Pastore, A; Temussi, P A; Salvadori, S; Tomatis, R; Mascagni, P

    1985-01-01

    Dermorphin, a natural peptide opioid containing a D-Ala2 residue, has been studied in dimethyl sulfoxide (DMSO) solution by means of several one-dimensional and two-dimensional 1H nuclear magnetic resonance (NMR) methods at various fields from 80 to 600 MHz. The combined use of conventional NMR parameters and of nuclear Overhauser effect effects points to an essentially extended structure. This conformation may be, in part, the result of strong interaction of the amide groups with DMSO molecules. PMID:4052557

  14. Solution conformations of the gamma-carboxyglutamic acid domain of bovine prothrombin fragment 1, residues 1-65.

    PubMed

    Charifson, P S; Darden, T; Tulinsky, A; Hughey, J L; Hiskey, R G; Pedersen, L G

    1991-01-15

    Molecular dynamics simulations have been performed (AMBER version 3.1) on solvated residues 1-65 of bovine prothrombin fragment 1 (BF1) by using the 2.8-A resolution crystallographic coordinates as the starting conformation for understanding calcium ion-induced conformational changes that precede experimentally observable phospholipid binding. Simulations were performed on the non-metal-bound crystal structure, the form resulting from addition of eight calcium ions to the 1-65 region of the crystal structure, the form resulting from removal of calcium ions after 107 ps and continuing the simulation, and an isolated hexapeptide loop (residues 18-23). In all cases, the 100-ps time scale seemed adequate to sample an ensemble of solution conformers within a particular region of conformation space. The non-metal-containing BF1 did not unfold appreciably during a 106-ps simulation starting from the crystallographic geometry. The calcium ion-containing structure (Ca-BF1) underwent an interesting conformational reorganization during its evolution from the crystal structure: during the time course of a 107-ps simulation, Ca-BF1 experienced a trans----cis isomerization of the gamma-carboxyglutamic acid-21 (Gla-21)-Pro-22 peptide bond. Removal of the calcium ions from this structure followed by 114 ps of additional molecular dynamics showed significant unfolding relative to the final 20-ps average structure of the 107-ps simulation; however, the Gla-21-Pro-22 peptide bond remained cis. A 265-ps simulation on the termini-protected hexapeptide loop (Cys-18 to Cys-23) containing two calcium ions also did not undergo a trans----cis isomerization. It is believed that the necessary activation energy for the transitional event observed in the Ca-BF1 simulation was largely supplied by global conformational events with a possible assist from relief of intermolecular crystal packing forces. The presence of a Gla preceding Pro-22, the inclusion of Pro-22 in a highly strained loop

  15. Protein-peptide molecular docking with large-scale conformational changes: the p53-MDM2 interaction

    NASA Astrophysics Data System (ADS)

    Ciemny, Maciej Pawel; Debinski, Aleksander; Paczkowska, Marta; Kolinski, Andrzej; Kurcinski, Mateusz; Kmiecik, Sebastian

    2016-12-01

    Protein-peptide interactions are often associated with large-scale conformational changes that are difficult to study either by classical molecular modeling or by experiment. Recently, we have developed the CABS-dock method for flexible protein-peptide docking that enables large-scale rearrangements of the protein chain. In this study, we use CABS-dock to investigate the binding of the p53-MDM2 complex, an element of the cell cycle regulation system crucial for anti-cancer drug design. Experimental data suggest that p53-MDM2 binding is affected by significant rearrangements of a lid region - the N-terminal highly flexible MDM2 fragment; however, the details are not clear. The large size of the highly flexible MDM2 fragments makes p53-MDM2 intractable for exhaustive binding dynamics studies using atomistic models. We performed extensive dynamics simulations using the CABS-dock method, including large-scale structural rearrangements of MDM2 flexible regions. Without a priori knowledge of the p53 peptide structure or its binding site, we obtained near-native models of the p53-MDM2 complex. The simulation results match well the experimental data and provide new insights into the possible role of the lid fragment in p53 binding. The presented case study demonstrates that CABS-dock methodology opens up new opportunities for protein-peptide docking with large-scale changes of the protein receptor structure.

  16. A Novel Four Amino Acid Determinant Defines Conformational Freedom within Chorionic Gonadotropin β-Subunits†

    PubMed Central

    Wilken, Jason A; Bedows, Elliott

    2008-01-01

    Based on apparent molecular mass heterogeneity following reducing versus non-reducing SDS-PAGE, we determined that the β-subunit of macaque (Macaca fascicularis) chorionic gonadotropin (mCG-β) is more conformationally constrained than is human chorionic gonadotropin (hCG-β). These two subunits share 81% amino acid identity. To determine the conformational variance source, which was not due to glycosylation differences, we generated a series of h-/mCG-β chimeras and identified domains that contributed to CG-β conformational freedom. We discovered that the CG-β 54-101 domain contained a small sub-domain, residues 74-77, that regulated the conformational freedom of the β-subunit, i.e., when residues 74-77 were of macaque origin (PGVD), mutated hCG-β subunit displayed macaque-like conformational rigidity; when residues 74-77 were of human origin (RGVN), mutated mCG-β subunit displayed human-like conformational freedom and microheterogeneity. Additionally, CG-β N-terminal domain residues (8, 18, 42, 46-48) were also found to influence CG-β conformational freedom when residues 74-77 were of human, but not macaque origin. The biological significance of the CG-β conformational variance was tested using a biological assay that showed that the hα/hβ CG heterodimer facilitated human CG receptor-mediated cAMP-driven luciferase reporter gene activity in HEK cells nearly an order of magnitude more effectively than did the hα/mCG-β chimera. Together, these data demonstrate that two essential amino acid residues contained within a four amino acid sub-domain regulated CG-β conformational freedom and that a conformational difference between hCG-β and mCG-β was recapitulated in the context of receptor-mediated CG heterodimer signal transduction activation. PMID:17358049

  17. Intra- and intermolecular forces dependent main chain conformations of esters of α,β-dehydroamino acids

    NASA Astrophysics Data System (ADS)

    Siodłak, Dawid; Bujak, Maciej; Staś, Monika

    2013-09-01

    Esters of dehydroamino acids occur in nature. To investigate their conformational properties, the low-temperature structures of Ac-ΔAla-OMe, Ac-ΔVal-OMe, Z-(Z)-ΔAbu-OMe, and Z-(Z)-ΔAbu-NHMe were studied by single-crystal X-ray diffraction. The ΔAla ester prefers the fully extended conformation C5. Both the ΔVal and (Z)-ΔAbu esters assume the conformation β, whereas the amide analogue of the latter prefers the conformation α. For the conformations found, DFT calculations using B3LYP/6-311++G(d,p) with the SCRF-PCM and M062X/6-311++G(d,p) with the SCRF-SMD method were applied to mimicking chloroform and water environment. The tendency of the ΔVal and (Z)-ΔAbu esters towards the conformation β, and their amide analogues towards the conformation α, with increase of the polarity of environment was found. The analysis of both intra- and intermolecular interactions including hydrogen bonds, carbonyl dipole attraction, and π-electron conjugation, enabled to understand and elucidate the conformational preferences of studied compounds. The studies show how the molecular structure, and in consequence, the conformation adopted by molecules is influenced by the different intra- and intermolecular forces.

  18. Conformational lability in the class II MHC 310 helix and adjacent extended strand dictate HLA-DM susceptibility and peptide exchange

    PubMed Central

    Painter, Corrie A.; Negroni, Maria P.; Kellersberger, Katherine A.; Zavala-Ruiz, Zarixia; Evans, James E.; Stern, Lawrence J.

    2011-01-01

    HLA-DM is required for efficient peptide exchange on class II MHC molecules, but its mechanism of action is controversial. We trapped an intermediate state of class II MHC HLA-DR1 by substitution of αF54, resulting in a protein with increased HLA-DM binding affinity, weakened MHC-peptide hydrogen bonding as measured by hydrogen-deuterium exchange mass spectrometry, and increased susceptibility to DM-mediated peptide exchange. Structural analysis revealed a set of concerted conformational alterations at the N-terminal end of the peptide-binding site. These results suggest that interaction with HLA-DM is driven by a conformational change of the MHC II protein in the region of the α-subunit 310 helix and adjacent extended strand region, and provide a model for the mechanism of DM-mediated peptide exchange. PMID:22084083

  19. Conformational lability in the class II MHC 310 helix and adjacent extended strand dictate HLA-DM susceptibility and peptide exchange.

    PubMed

    Painter, Corrie A; Negroni, Maria P; Kellersberger, Katherine A; Zavala-Ruiz, Zarixia; Evans, James E; Stern, Lawrence J

    2011-11-29

    HLA-DM is required for efficient peptide exchange on class II MHC molecules, but its mechanism of action is controversial. We trapped an intermediate state of class II MHC HLA-DR1 by substitution of αF54, resulting in a protein with increased HLA-DM binding affinity, weakened MHC-peptide hydrogen bonding as measured by hydrogen-deuterium exchange mass spectrometry, and increased susceptibility to DM-mediated peptide exchange. Structural analysis revealed a set of concerted conformational alterations at the N-terminal end of the peptide-binding site. These results suggest that interaction with HLA-DM is driven by a conformational change of the MHC II protein in the region of the α-subunit 3(10) helix and adjacent extended strand region, and provide a model for the mechanism of DM-mediated peptide exchange.

  20. Multiple peptide synthesis on acid-labile handle derivatized polyethylene supports.

    PubMed

    Valerio, R M; Bray, A M; Maeji, N J

    1994-08-01

    Using the multipin peptide synthesis approach, a range of peptides with native amide and carboxylate C-termini were generated using an acid-labile approach. Polyethylene crowns grafted with hydroxyethylmethacrylate (HEMA) polymer were functionalized with either 4-hydroxymethylphenoxyacetic acid for the generation of peptide-carboxylate or p-[(R,S)-alpha-[1-(9H-fluoren-9-yl)methoxyformamido]-2,4-dim ethoxy- benzyl]phenoxyacetic acid for peptide-amide. A range of known peptide hormone sequences and other peptides with native C-termini were assembled by sequential incorporation of N alpha-Fmoc protected amino acids. Peptides were sidechain deprotected and cleaved from crowns with TFA/scavengers within 2 mL centrifuge tubes, and isolated by a series of ether/petrol wash and centrifugation steps. In this way it was possible to avoid a cleavage and isolation botteneck, allowing rapid processing of large numbers of peptides.

  1. Polar Hinges as Functionalized Conformational Constraints in (Bi)cyclic Peptides.

    PubMed

    van de Langemheen, Helmus; Korotkovs, Valerijs; Bijl, Joachim; Wilson, Claire; Kale, Sangram S; Heinis, Christian; Liskamp, Rob M J

    2017-02-16

    Two polar hinges for cyclization of peptides have been developed, leading to bicyclic peptides and cyclized peptides with improved solubility and biological activity. Increasingly, we note that a good aqueous solubility of peptides is an absolute prerequisite, not only to allow handling and purification of our target peptides but also being crucial for biological activity characteristics. Compared to earlier hinges, the 1,1',1"-(1,3,5-triazinane-1,3,5-triyl)tris(2-bromoethanone) (TATB) and 2,4,6-tris(bromomethyl)-s-triazine (TBMT), each containing three nitrogen atoms are structurally similar but chemically very different. Both were accessible in a one-step fashion from bromoacetonitrile. TATB and TBMT are very suitable for the preparation of more soluble bicyclic peptides. Azide-modified TATB and TBMT derivatives provide hinges for the preparation of cyclized peptides for incorporation on scaffolds to afford protein mimics. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. The research on conformal acid etching process of glass ceramic

    NASA Astrophysics Data System (ADS)

    Wang, Kepeng; Guo, Peiji

    2014-08-01

    A series of experiments have been done to explore the effect of different conditions on the hydrofluoric acid etching. The hydrofluoric acid was used to etch the glass ceramic called "ZERODUR", which is invented by SCHOTT in Germany. The glass ceramic was processed into cylindrical samples. The hydrofluoric acid etching was done in a plastic beaker. The concentration of hydrofluoric acid and the etching time were changed to measure the changes of geometric tolerance and I observed the surface using a microscope in order to find an appropriate condition of hydrofluoric acid etching.

  3. Characterization of natural peptide ligands for HLA-B*4402 and -B*4403: implications for peptide involvement in allorecognition of a single amino acid change in the HLA-B44 heavy chain.

    PubMed

    Fleischhauer, K; Avila, D; Vilbois, F; Traversari, C; Bordignon, C; Wallny, H J

    1994-11-01

    This study describes the characterization of endogenous peptides associated with the two major subtypes of HLA-B44. The two subtypes differ for a single amino acid substitution from Asp (HLA-B*4402) to Leu (HLA-B*4403) in position 156 of the alpha 2 domain, causing strong alloreactivity in vivo. In order to study the involvement of peptides in this phenomenon, the peptide motifs of the two subtypes were determined from natural peptide pools using Edman degradation. The motif was found to be essentially identical for HLA-B*4402 and -B*4403, with a strong predominance for Glu at position 2, Tyr or Phe at positions 9 and 10 and hydrophobic residues, especially Met, at position 3. Two individual naturally processed ligands of HLA-B*4403 were sequenced and shown to be derived from intracellularly expressed proteins found in protein sequence databases. The sequence of these natural peptide ligands conform well to the determined motif. These data will allow the prediction of HLA-B44 restricted peptide epitopes from viral and tumor antigens of known amino acid sequences. Moreover, they indicate that the peptide repertoire presented by HLA-B*4402 and -B*4403 is very similar, suggesting that the strong alloresponse between these two subtypes is not due to presentation of a different set of self peptides.

  4. Kojic Acid Peptide: A New Compound with Anti-Tyrosinase Potential

    PubMed Central

    Singh, Birendra Kumar; Park, Seok Hoon; Lee, Hyang-Bok; Goo, Young-Aae; Kim, Hyoung Shik; Cho, Seung Hee; Lee, Jeong Hun; Ahn, Ghe Whan; Kim, Jin Pyo; Kang, Su Myoung

    2016-01-01

    Background Kojic acid was used for decades in the cosmetic industry as an antimelanogenic agent. However, there are two major drawbacks of Kojic acid, one is cytotoxicity and second are instability on storage. These limitations led the scientist to synthesize the active Kojic acid peptides. Objective In the present study, we synthesize and investigate the effect of five Kojic acid peptides to overcome the limitation of Kojic acid. Methods The peptide was analyzed and purified by high-performance liquid chromatography and matrix-assisted laser desorption ionization time of flight mass spectroscopy. Further, the tyrosinase activities of the Kojic acid and Kojic acid peptides were compared. The toxicity was measured and the melanin content is recorded in B16F10 mouse melanoma cells. Results Maximum tyrosinase activity was measured by Kojic acid peptides. Therefore, Kojic acid peptides were subjected to melanin assay and cytotoxicity assay and finally the stability of the Kojic acid peptide was measured. Conclusion It was observed that this newly synthesized Kojic acid peptide is stable and potent to inhibit the tyrosinase activity and melanin content of B16F10 mouse melanoma cells without exhibiting cell toxicity. Together, these preliminary results suggest that a further exploration is being needed to establish Kojic acid peptide as antimelanogenic agent. PMID:27746633

  5. Determination of conformational free energies of peptides by multidimensional adaptive umbrella sampling

    NASA Astrophysics Data System (ADS)

    Wang, Jun; Gu, Yan; Liu, Haiyan

    2006-09-01

    We improve the multidimensional adaptive umbrella sampling method for the computation of conformational free energies of biomolecules. The conformational transition between the α-helical and β-hairpin conformational states of an alanine decapeptide is used as an example. Convergence properties of the weighted-histogram-analysis-based adaptive umbrella sampling can be improved by using multiple replicas in each adaptive iteration and by using adaptive updating of the bounds of the umbrella potential. Using positional root-mean-square deviations from structures of the α-helical and β-hairpin reference states as reaction coordinates, we obtained well-converged free energy surfaces of both the in-vacuum and in-solution decapeptide systems. From the free energy surfaces well-converged relative free energies between the two conformational states can be derived. Advantages and disadvantages of different methods for obtaining conformational free energies as well as implications of our results in studying conformational transitions of proteins and in improving force field are discussed.

  6. NMR based solvent exchange experiments to understand the conformational preference of intrinsically disordered proteins using FG-nucleoporin peptide as a model.

    PubMed

    Heisel, Kurt A; Krishnan, V V

    2014-01-01

    The conformational preference of a peptide with three phenylalanine-glycine (FG) repeats from the intrinsically disordered domain of nucleoporin 159 (nup159) from the yeast nucleopore complex is studied. Conformational states of this FG-peptide in dimethyl sulfoxide (DMSO), a non-native solvent, are first studied. A solvent exchange scheme is designed and performed to understand how the conformational preferences of the peptide are altered as the solvent shifts from DMSO to water. An ensemble of structures of a 19-residue peptide is determined based on (13)Cα, (1)Hα, and (1)HN chemical shifts and with inter-proton distances. An experimental model is then presented where chemical shifts and amide-proton temperature dependence is probed at changing DMSO to water ratios. These co-solvent experiments provide evidence of a conformational change as the fraction of water increases by the stark change in the behavior of amide protons under varied temperature. This investigation provides a NMR based experimental method in the field of intrinsically disordered proteins to realize conformational transitions from a non-native set of structures (in DMSO) to a native set of disordered conformers (in water). Copyright © 2013 Wiley Periodicals, Inc.

  7. Template Based Design of Anti-Metastatic Drugs from the Active Conformation of Laminin Peptide II

    DTIC Science & Technology

    2001-01-01

    bind to positively charged side-chains (blue) and residues with negatively charged Primaria @ tissue culture 96-well plates as charged side-chains (red... Primaria " plate. Binding of the syn- heparan sulfate followed by peptide 11 elutions, thetic LBP205-2 9 peptide was quantitated using our LBP 2°5-229...Dynex Technologies, Inc. (Chantilly, VA). Positively na State University. The IgG fraction was purified using charged Primaria ®’ and standard Falcon

  8. [SPC/E and TIP4P models for investigation of the conformational mobility of the insulin superfamily peptides].

    PubMed

    Ksenofontova, O I

    2014-01-01

    In this work we carried out a comparative analysis of the two most popular water models-SPC/E and TIP4P and estimated the ability of using ones for insulin superfamily peptides-proinsulin and insulin-like growth factors (IGF1 and IGF2). It was shown that root-mean-square deviations and radius of gyration had tend to be in reversed phase when both water models were used. Only IGF1 had a plateau after 9000 ps. In addition, it was shown that in spite of the general nature of insulin-like packing maintenance, there were some differences in the secondary structures, when we used TIP4P and SPC/E. These differences could influence on the overall molecule dynamics and the ability to accept necessary conformation for interaction with cognate receptors. On the basis of the received data we concluded that it is necessary to use several, not one, water models for the study of the peptides conformational mobility.

  9. Antimicrobial Peptide Potency is Facilitated by Greater Conformational Flexibility when Binding to Gram-negative Bacterial Inner Membranes

    PubMed Central

    Amos, Sarah-Beth T. A.; Vermeer, Louic S.; Ferguson, Philip M.; Kozlowska, Justyna; Davy, Matthew; Bui, Tam T.; Drake, Alex F.; Lorenz, Christian D.; Mason, A. James

    2016-01-01

    The interaction of antimicrobial peptides (AMPs) with the inner membrane of Gram-negative bacteria is a key determinant of their abilities to exert diverse bactericidal effects. Here we present a molecular level understanding of the initial target membrane interaction for two cationic α-helical AMPs that share structural similarities but have a ten-fold difference in antibacterial potency towards Gram-negative bacteria. The binding and insertion from solution of pleurocidin or magainin 2 to membranes representing the inner membrane of Gram-negative bacteria, comprising a mixture of 128 anionic and 384 zwitterionic lipids, is monitored over 100 ns in all atom molecular dynamics simulations. The effects of the membrane interaction on both the peptide and lipid constituents are considered and compared with new and published experimental data obtained in the steady state. While both magainin 2 and pleurocidin are capable of disrupting bacterial membranes, the greater potency of pleurocidin is linked to its ability to penetrate within the bacterial cell. We show that pleurocidin displays much greater conformational flexibility when compared with magainin 2, resists self-association at the membrane surface and penetrates further into the hydrophobic core of the lipid bilayer. Conformational flexibility is therefore revealed as a key feature required of apparently α-helical cationic AMPs for enhanced antibacterial potency. PMID:27874065

  10. Antimicrobial Peptide Potency is Facilitated by Greater Conformational Flexibility when Binding to Gram-negative Bacterial Inner Membranes

    NASA Astrophysics Data System (ADS)

    Amos, Sarah-Beth T. A.; Vermeer, Louic S.; Ferguson, Philip M.; Kozlowska, Justyna; Davy, Matthew; Bui, Tam T.; Drake, Alex F.; Lorenz, Christian D.; Mason, A. James

    2016-11-01

    The interaction of antimicrobial peptides (AMPs) with the inner membrane of Gram-negative bacteria is a key determinant of their abilities to exert diverse bactericidal effects. Here we present a molecular level understanding of the initial target membrane interaction for two cationic α-helical AMPs that share structural similarities but have a ten-fold difference in antibacterial potency towards Gram-negative bacteria. The binding and insertion from solution of pleurocidin or magainin 2 to membranes representing the inner membrane of Gram-negative bacteria, comprising a mixture of 128 anionic and 384 zwitterionic lipids, is monitored over 100 ns in all atom molecular dynamics simulations. The effects of the membrane interaction on both the peptide and lipid constituents are considered and compared with new and published experimental data obtained in the steady state. While both magainin 2 and pleurocidin are capable of disrupting bacterial membranes, the greater potency of pleurocidin is linked to its ability to penetrate within the bacterial cell. We show that pleurocidin displays much greater conformational flexibility when compared with magainin 2, resists self-association at the membrane surface and penetrates further into the hydrophobic core of the lipid bilayer. Conformational flexibility is therefore revealed as a key feature required of apparently α-helical cationic AMPs for enhanced antibacterial potency.

  11. Natively unfolded human prothymosin alpha adopts partially folded collapsed conformation at acidic pH.

    PubMed

    Uversky, V N; Gillespie, J R; Millett, I S; Khodyakova, A V; Vasiliev, A M; Chernovskaya, T V; Vasilenko, R N; Kozlovskaya, G D; Dolgikh, D A; Fink, A L; Doniach, S; Abramov, V M

    1999-11-09

    Prothymosin alpha has previously been shown to be unfolded at neutral pH, thus belonging to a growing family of "natively unfolded" proteins. The structural properties and conformational stability of recombinant human prothymosin alpha were characterized at neutral and acidic pH by gel filtration, SAXS, circular dichroism, ANS fluorescence, (1)H NMR, and resistance to urea-induced unfolding. Interestingly, prothymosin alpha underwent a cooperative transition from the unfolded state into a partially folded conformation on lowering the pH. This conformation of prothymosin alpha is a compact denatured state, with structural properties different from those of the molten globule. The formation of alpha-helical structure by the glutamic acid-rich elements of the protein accompanied by the partial hydrophobic collapse is expected at lower pH due to the neutralization of the negatively charged residues. It is possible that such conformational changes may be associated with the protein function.

  12. Peptide nucleic acid - an opportunity for bio-nanotechnology.

    PubMed

    Anstaett, Philipp; Gasser, Gilles

    2014-01-01

    DNA is a major player in the field of bio-nanotechnology and many interesting applications have been realized using this oligonucleotide. In contrast, the use of peptide nucleic acid (PNA), which is a non-natural, neutral analogue of DNA with superior hybridization strengths compared to DNA, is still in its infancy in bio-nanotechnology. However, as demonstrated in this short review using selected studies, promising examples demonstrating the tremendous opportunities that PNA can offer for bio-nanotechnology were recently described.

  13. Conformational preferences of γ-aminobutyric acid in the gas phase and in water

    NASA Astrophysics Data System (ADS)

    Song, Il Keun; Kang, Young Kee

    2012-09-01

    The conformational study of γ-aminobutyric acid (GABA) has been carried out at the M06-2X/cc-pVTZ level of theory in the gas phase and the SMD M06-2X/cc-pVTZ level of theory in water. In the gas phase, the folded conformation gG1 with gauche- and gauche+ conformations for the Cβsbnd Cα and Cγsbnd Cβ bonds, respectively, is found to be lowest in energy and enthalpy, which can be ascribed to the favored hyperconjugative n → π* interaction between the lone electron pair of the amine nitrogen atom and the Cdbnd O bond of the carboxylic group and the favored antiparallel dipole-dipole interaction between the Nsbnd H bond and the Cdbnd O bond. In addition, the intramolecular hydrogen bonds between the carboxylic group and the amine Nsbnd H group have contributed to stabilize some low-energy conformers. However, the most preferred conformation is found to be tG1 and more stable by 0.4 kcal/mol in ΔG than the conformer gG1, in which the favored entropic term due to the conformational flexibility and the other favored n → σ*, σ → σ*, and π → σ* interactions seem to play a role. The conformational preferences of the neutral GABA calculated by ΔG's are reasonably consistent with the populations deduced from FT microwave spectroscopy in supersonic jets combined with laser ablation. In water, the two folded conformers Gg and gG of the zwitterionic GABA are dominantly populated, each of which has the population of 47%, and the hydrogen bond between the ammonium Nsbnd H group and the lone electron pair of the Csbnd O- group seems to be crucial in stabilizing these conformers. Our calculated result that the folded conformers preferentially exist in water is consistent with the 1H NMR experiments in D2O.

  14. Helical Conformation of the SEVI Precursor Peptide PAP248-286, a Dramatic Enhancer of HIV Infectivity, Promotes Lipid Aggregation and Fusion

    PubMed Central

    Brender, Jeffrey R.; Hartman, Kevin; Gottler, Lindsey M.; Cavitt, Marchello E.; Youngstrom, Daniel W.; Ramamoorthy, Ayyalusamy

    2009-01-01

    In previous in vivo studies, amyloid fibers formed from a peptide ubiquitous in human seminal fluid (semen-derived enhancer of viral infection (SEVI)) were found to dramatically enhance the infectivity of the HIV virus (3–5 orders of magnitude by some measures). To complement those studies, we performed in vitro assays of PAP248-286, the most active precursor to SEVI, and other polycationic polymers to investigate the physical mechanisms by which the PAP248-286 promotes the interaction with lipid bilayers. At acidic (but not at neutral) pH, freshly dissolved PAP248-286 catalyzes the formation of large lipid flocculates in a variety of membrane compositions, which may be linked to the promotion of convective transport in the vaginal environment rather than transport by a random Brownian motion. Furthermore, PAP248-286 is itself fusiogenic and weakens the integrity of the membrane in such a way that may promote fusion by the HIV gp41 protein. An α-helical conformation of PAP248-286, lying parallel to the membrane surface, is implicated in promoting bridging interactions between membranes by the screening of the electrostatic repulsion that occurs when two membranes are brought into close contact. This suggests that nonspecific binding of monomeric or small oligomeric forms of SEVI in a helical conformation to lipid membranes may be an additional mechanism by which SEVI enhances the infectivity of the HIV virus. PMID:19883590

  15. CD and NMR conformational studies of a peptide encompassing the Mid Loop interface of Ship2-Sam.

    PubMed

    Mercurio, Flavia A; Scognamiglio, Pasqualina L; Di Natale, Concetta; Marasco, Daniela; Pellecchia, Maurizio; Leone, Marilisa

    2014-11-01

    The lipid phosphatase Ship2 is a protein that intervenes in several diseases such as diabetes, cancer, neurodegeneration, and atherosclerosis. It is made up of a catalytic domain and several protein docking modules such as a C-terminal Sam (Sterile alpha motif) domain. The Sam domain of Ship2 (Ship2-Sam) binds to the Sam domains of the EphA2 receptor (EphA2-Sam) and the PI3K effector protein Arap3 (Arap3-Sam). These heterotypic Sam-Sam interactions occur through formation of dimers presenting the canonical "Mid Loop/End Helix" binding mode. The central region of Ship2-Sam, spanning the C-terminal end of α2, the α3 and α4 helices together with the α2α3 and α3α4 interhelical loops, forms the Mid Loop surface that is needed to bind partners Sam domains. A peptide encompassing most of the Ship2-Sam Mid Loop interface (Shiptide) capable of binding to both EphA2-Sam and Arap3-Sam, was previously identified. Here we investigated the conformational features of this peptide, through solution CD and NMR studies in different conditions. These studies reveal that the peptide is highly flexible in aqueous buffer, while it adopts a helical conformation in presence of 2,2,2-trifluoroethanol. The discovered structural insights and in particular the identification of a helical motif, may lead to the design of more constrained and possibly cell permeable Shiptide analogs that could work as efficient antagonists of Ship2-Sam heterotypic interactions and embrace therapeutic applications.

  16. Fatty Acids Change the Conformation of Uncoupling Protein 1 (UCP1)*

    PubMed Central

    Divakaruni, Ajit S.; Humphrey, Dickon M.; Brand, Martin D.

    2012-01-01

    UCP1 catalyzes proton leak across the mitochondrial inner membrane to disengage substrate oxidation from ATP production. It is well established that UCP1 is activated by fatty acids and inhibited by purine nucleotides, but precisely how this regulation occurs remains unsettled. Although fatty acids can competitively overcome nucleotide inhibition in functional assays, fatty acids have little effect on purine nucleotide binding. Here, we present the first demonstration that fatty acids induce a conformational change in UCP1. Palmitate dramatically changed the binding kinetics of 2′/3′-O-(N-methylanthraniloyl)-GDP, a fluorescently labeled nucleotide analog, for UCP1. Furthermore, palmitate accelerated the rate of enzymatic proteolysis of UCP1. The altered kinetics of both processes indicate that fatty acids change the conformation of UCP1, reconciling the apparent discrepancy between existing functional and ligand binding data. Our results provide a framework for how fatty acids and nucleotides compete to regulate the activity of UCP1. PMID:22952235

  17. The nature of peptide interactions with acid end-group PLGAs and facile aqueous-based microencapsulation of therapeutic peptides

    PubMed Central

    Sophocleous, Andreas M.; Desai, Kashappa-Goud H.; Mazzara, J. Maxwell; Tong, Ling; Cheng, Ji-Xin; Olsen, Karl F.; Schwendeman, Steven P.

    2013-01-01

    An important poorly understood phenomenon in controlled-release depots involves the strong interaction between common cationic peptides and low Mw free acid end-group poly(lactic-co-glycolic acids) (PLGAs) used to achieve continuous peptide release kinetics. The kinetics of peptide sorption to PLGA was examined by incubating peptide solutions of 0.2-4 mM octreotide or leuprolide acetate salts in 0.1 M HEPES buffer, pH 7.4, with polymer particles or films at 4-37 °C for 24 h. The extent of absorption/loading of peptides in PLGA particles/films was assayed by two-phase extraction and amino acid analysis. Confocal Raman microspectroscopy and stimulated Raman scattering (SRS) and laser scanning confocal imaging techniques were used to examine peptide penetration in the polymer phase. The release of sorbed peptide from leuprolide-PLGA particles was evaluated both in vitro (PBST + 0.02% sodium azide, 37 °C) and in vivo (male Sprague-Dawley rats). We found that when the PLGA-COOH chains are sufficiently mobilized, therapeutic peptides not only bind at the surface, a common belief to date, but can also internalized and distributed throughout the polymer phase at physiological temperature forming a salt with low-molecular weight PLGA-COOH. Importantly, absorption of leuprolide into low MW PLGA-COOH particles yielded ~17 wt% leuprolide loading in the polymer (i.e., ~70% of PLGA-COOH acids occupied), and the absorbed peptide was released from the polymer for > 2 weeks in a controlled fashion in vitro and as indicated by sustained testosterone suppression in male Sprague-Dawley rats. This new approach, which bypasses the traditional encapsulation method and associated production cost, opens up the potential for facile production of low-cost controlled-release injectable depots for leuprolide and related peptides. PMID:24021356

  18. The nature of peptide interactions with acid end-group PLGAs and facile aqueous-based microencapsulation of therapeutic peptides.

    PubMed

    Sophocleous, Andreas M; Desai, Kashappa-Goud H; Mazzara, J Maxwell; Tong, Ling; Cheng, Ji-Xin; Olsen, Karl F; Schwendeman, Steven P

    2013-12-28

    An important poorly understood phenomenon in controlled-release depots involves the strong interaction between common cationic peptides and low Mw free acid end-group poly(lactic-co-glycolic acids) (PLGAs) used to achieve continuous peptide release kinetics. The kinetics of peptide sorption to PLGA was examined by incubating peptide solutions of 0.2-4mM octreotide or leuprolide acetate salts in a 0.1M HEPES buffer, pH7.4, with polymer particles or films at 4-37°C for 24h. The extent of absorption/loading of peptides in PLGA particles/films was assayed by two-phase extraction and amino acid analysis. Confocal Raman microspectroscopy, stimulated Raman scattering (SRS) and laser scanning confocal imaging, and microtome sectioning techniques were used to examine peptide penetration into the polymer phase. The release of sorbed peptide from leuprolide-PLGA particles was evaluated both in vitro (PBST+0.02% sodium azide, 37°C) and in vivo (male Sprague-Dawley rats). We found that when the PLGA-COOH chains are sufficiently mobilized, therapeutic peptides not only bind at the surface, a common belief to date, but also can be internalized and distributed throughout the polymer phase at physiological temperature forming a salt with low-molecular weight PLGA-COOH. Importantly, absorption of leuprolide into low MW PLGA-COOH particles yielded ~17 wt.% leuprolide loading in the polymer (i.e., ~70% of PLGA-COOH acids occupied), and the absorbed peptide was released from the polymer for >2 weeks in a controlled fashion in vitro and as indicated by sustained testosterone suppression in male Sprague-Dawley rats. This new approach, which bypasses the traditional encapsulation method and associated production cost, opens up the potential for facile production of low-cost controlled-release injectable depots for leuprolide and related peptides. © 2013.

  19. Real-time Measurement of Membrane Conformational States Induced by Antimicrobial Peptides: Balance Between Recovery and Lysis

    PubMed Central

    Hall, Kristopher; Lee, Tzong-Hsien; Mechler, Adam I.; Swann, Marcus J.; Aguilar, Marie-Isabel

    2014-01-01

    The disruption of membranes by antimicrobial peptides is a multi-state process involving significant structural changes in the phospholipid bilayer. However, direct measurement of these membrane structural changes is lacking. We used a combination of dual polarisation interferometry (DPI), surface plasmon resonance spectroscopy (SPR) and atomic force microscopy (AFM) to measure the real-time changes in membrane structure through the measurement of birefringence during the binding of magainin 2 (Mag2) and a highly potent analogue in which Ser8, Gly13 and Gly18 has been replaced with alanine (Mag-A). We show that the membrane bilayer undergoes a series of structural changes upon peptide binding before a critical threshold concentration is reached which triggers a significant membrane disturbance. We also propose a detailed model for antimicrobial peptide action as a function of the degree of bilayer disruption to provide an unprecedented in-depth understanding of the membrane lysis in terms of the interconversion of different membrane conformational states in which there is a balance between recovery and lysis. PMID:24969959

  20. Control of Heme Coordination and Catalytic Activity by Conformational Changes in Peptide-Amphiphile Assemblies.

    PubMed

    Solomon, Lee A; Kronenberg, Jacob B; Fry, H Christopher

    2017-06-28

    Self-assembling peptide materials have gained significant attention, due to well-demonstrated applications, but they are functionally underutilized. To advance their utility, we use noncovalent interactions to incorporate the biological cofactor heme-B for catalysis. Heme-proteins achieve differing functions through structural and coordinative variations. Here, we replicate this phenomenon by highlighting changes in heme reactivity as a function of coordination, sequence, and morphology (micelles versus fibers) in a series of simple peptide amphiphiles with the sequence c16-xyL3K3-CO2H where c16 is a palmitoyl moiety and xy represents the heme binding region: AA, AH, HH, and MH. The morphology of this peptide series is characterized using transmission electron and atomic force microscopies as well as dynamic light scattering. Within this small library of peptide constructs, we show that three spectroscopically (UV/visible and electron paramagnetic resonance) distinct heme environments were generated: noncoordinated/embedded high-spin, five-coordinate high-spin, and six-coordinate low-spin. The resulting material's functional dependence on sequence and supramolecular morphology is highlighted 2-fold. First, the heme active site binds carbon monoxide in both micelles and fibers, demonstrating that the heme active site in both morphologies is accessible to small molecules for catalysis. Second, peroxidase activity was observed in heme-containing micelles yet was significantly reduced in heme-containing fibers. We briefly discuss the implications these findings have in the production of functional, self-assembling peptide materials.

  1. Conformation specific spectroscopy in the complexity gap: beta-peptides and flexible bichromophores

    NASA Astrophysics Data System (ADS)

    Baquero, Esteban Edwin

    Flexible biomolecules with many degrees of freedom have the ability to sample a great number of structural minima. The intrinsic structural preferences of these molecules are driven by stabilizations due to intramolecular interactions such as hydrogen bonding and interatomic attractive forces. These interactions become increasingly important as the atoms in the molecule come in close proximity to one another. The present work describes the conformational preferences of several model bio-relevant molecules of different sizes where different kinds of intramolecular interactions can occur. Conformation specific ultraviolet and infrared spectroscopies were utilized to obtain the spectroscopic signatures of different conformations of the jet cooled biomolecules. These experiments allowed for the assignment of conformational families and the development of a general protocol to identify the infrared signatures of amide NH stretches, which were found to vary in frequency due to their immediate chemical and structural environment. Specific examples of systems studied include molecules containing unnatural polypeptide chains and a flexible bichromophore. The unnatural polypetides (Ac-beta3-hPhe-NHMe, Ac-beta 3-hTyr-NHMe Ac-beta3-hPhe-beta3-hAla-NHMe and Ac-beta3-hAla-beta3-hPhe-NHMe) were found to have a rich conformational potential energy landscape that contained many kinds of intramolecular hydrogen bonded minima. The flexible bichromophore (HNBPA), containing two spectroscopically distinguishable chromophores (Phenol and Phenyl), was found to be an interesting case study for conformational specific electronic energy transfer and testing how well different theoretical methods manage non-covalent interactions, such as those between side-chain atoms and aromatic pi-clouds.

  2. Effects of osmolytes on the helical conformation of model peptide: Molecular dynamics simulation

    NASA Astrophysics Data System (ADS)

    Mehrnejad, Faramarz; Ghahremanpour, Mohammad Mehdi; Khadem-Maaref, Mahmoud; Doustdar, Farahnoosh

    2011-01-01

    Co-solvents such as glycerol and sorbitol are small organic molecules solvated in the cellular solutions that can have profound effects on the protein structures. Here, the molecular dynamics simulations and comparative structural analysis of magainin, as a peptide model, in pure water, 2,2,2-trifluoroethanol/water, glycerol/water, and sorbitol/water are reported. Our results show that the peptide NMR structure is largely maintained its native structure in osmolytes-water mixtures. The simulation data indicates that the stabilizing effect of glycerol and sorbitol is induced by preferential accumulation of glycerol and sorbitol molecules around the nonpolar and aromatic residues. Thus, the presence of glycerol and sorbitol molecules decreases the interactions of water molecules with the hydrophobic residues of the peptide, and the alpha helical structure is stabilized.

  3. Identification of a Novel Parallel β‐Strand Conformation within Molecular Monolayer of Amyloid Peptide

    PubMed Central

    Liu, Lei; Li, Qiang; Zhang, Shuai; Wang, Xiaofeng; Hoffmann, Søren Vrønning; Li, Jingyuan; Liu, Zheng

    2016-01-01

    The differentiation of protein properties and biological functions arises from the variation in the primary and secondary structure. Specifically, in abnormal assemblies of protein, such as amyloid peptide, the secondary structure is closely correlated with the stable ensemble and the cytotoxicity. In this work, the early Aβ33‐42 aggregates forming the molecular monolayer at hydrophobic interface are investigated. The molecular monolayer of amyloid peptide Aβ33‐42 consisting of novel parallel β‐strand‐like structure is further revealed by means of a quantitative nanomechanical spectroscopy technique with force controlled in pico‐Newton range, combining with molecular dynamic simulation. The identified parallel β‐strand‐like structure of molecular monolayer is distinct from the antiparallel β‐strand structure of Aβ33‐42 amyloid fibril. This finding enriches the molecular structures of amyloid peptide aggregation, which could be closely related to the pathogenesis of amyloid disease. PMID:27818898

  4. Molecular self-assembly using peptide nucleic acids.

    PubMed

    Berger, Or; Gazit, Ehud

    2017-01-01

    Peptide nucleic acids (PNAs) are extensively studied for the control of genetic expression since their design in the 1990s. However, the application of PNAs in nanotechnology is much more recent. PNAs share the specific base-pair recognition characteristic of DNA together with material-like properties of polyamides, both proteins and synthetic polymers, such as Kevlar and Nylon. The first application of PNA was in the form of PNA-amphiphiles, resulting in the formation of either lipid integrated structures, hydrogels or fibrillary assemblies. Heteroduplex DNA-PNA assemblies allow the formation of hybrid structures with higher stability as compared with pure DNA. A systematic screen for minimal PNA building blocks resulted in the identification of guanine-containing di-PNA assemblies and protected guanine-PNA monomer spheres showing unique optical properties. Finally, the co-assembly of PNA with thymine-like three-faced cyanuric acid allowed the assembly of poly-adenine PNA into fibers. In summary, we believe that PNAs represent a new and important family of building blocks which converges the advantages of both DNA- and peptide-nanotechnologies.

  5. Reactivity of consecutive basic amino acid residues in peptides.

    PubMed

    Bayle-Lacoste, M; De Tinguy-Moreaud, E; Geoffre, S; Neuzil, E

    1987-03-01

    Different tetrapeptides of general formula L-Ala-X-X-Gly, possessing a basic doublet in the second and third position (X = Arg or Lys), have been synthesized as free or N-acetylated molecules. The chemical reactivity of the arginine guanidino group and of the lysine epsilon-amino group were studied using respectively the Sakaguchi and the ortho-diacetylbenzene reactions, in the tetrapeptides as well as in related molecules. In both cases, the colour yield is markedly influenced by the length of the polypeptide chain and by the relative positions of the arginine and lysine residues, suggesting the occurrence of intramolecular bonds within the tetrapeptide molecule. Tryptic hydrolysis of the tetrapeptides was followed by evaluating the amino acids or peptides which appear to be specific for the different possible cleavages at the arginyl or at the lysyl bonds. The susceptibility to trypsin of the carboxylic group of the second basic amino acid decreases progressively in the order Lys-Arg greater than Arg-Arg much greater than Lys-Lys greater than Arg-Lys, which shows a fair correlation with the intra-cellular cleavage of the bonds observed during the processing of preproteins of of the precursors of several physiologically active peptides.

  6. Conformational changes in the PBX homeodomain and C-terminal extension upon binding DNA and HOX-derived YPWM peptides.

    PubMed

    Sprules, T; Green, N; Featherstone, M; Gehring, K

    2000-08-15

    PBX is a member of the three amino acid loop extension (TALE) class of homeodomains. PBX binds DNA cooperatively with HOX homeodomain proteins that contain a conserved YPWM motif. The amino acids immediately C-terminal to the PBX homeodomain increase the affinity of the homeodomain for its DNA site and HOX proteins. We have determined the structure of the free PBX homeodomain using NMR spectroscopy. Both the PBX homeodomain and the extended PBX homeodomain make identical contacts with a 5'-TGAT-3' DNA site and a YPWM peptide. A fourth alpha-helix, which forms upon binding to DNA, stabilizes the extended PBX structure. Variations in DNA sequence selectivity of heterodimeric PBX-HOX complexes depend on the HOX partner; however, a comparison of five different HOX-derived YPWM peptides showed that each bound to PBX in the same way, differing only in the strength of the association.

  7. Delineation of Alternative Conformational States in E.coli Peptide Deformylase via Thermodynamic Studies for the Binding of Actinonin†

    PubMed Central

    Berg, Alexander K.; Srivastava, D. K.

    2009-01-01

    We investigated the binding of a naturally occurring antibiotic, actinonin, to the Ni2+ reconstituted recombinant form of E. coli peptide deformylase (PDFEc) via isothermal titration microcalorimetry. The binding data conformed to both exothermic and endothermic phases with magnitudes of ΔG°, ΔH°, and TΔS° being equal to −12, −2.7, and 9.3, and −8.7, 3.9, and 12.6 kcal/mol, respectively. Evidently, although both phases are dominated by favorable entropic changes, the exothermic phase has about 6.7 kcal/mol enthalpic advantage over the endothermic phase. We observed that the removal of bound Ni2+ from PDFEc abolished the exothermic phase without affecting the endothermic phase, but it was regained upon addition of Zn2+. In conjunction with metal analysis data, we propose that the recombinant form of PDFEc is expressed in two stable conformational states that yield markedly distinct ITC profiles (i.e., exothermic versus endothermic) upon interaction with actinonin. The existence of two conformational states of PDFEc is further supported by the observation of two distinct and independent transitions during the thermal unfolding of the enzyme. In addition, the thermodynamic data reveals that the formation of the PDFEc-actinonin complex results in the transfer of one H+ from the enzyme phase to the bulk solvent at pH 6.3. Both exothermic and endothermic phases produce highly negative ΔCp° values, but there is no apparent enthalpy-entropy compensation effect upon formation of the PDFEc-actinonin complex. In view of the known structural features of the enzyme, arguments are presented that the alternative conformational states of PDFEc are modulated by the metal ligation at the enzyme site. PMID:19191548

  8. Equilibrium conformational ensemble of the intrinsically disordered peptide n16N: linking subdomain structures and function in nacre.

    PubMed

    Brown, Aaron H; Rodger, P Mark; Evans, John Spencer; Walsh, Tiffany R

    2014-12-08

    n16 is a framework protein family associated with biogenic mineral stabilization, thought to operate at three key interfaces in nacre: protein/β-chitin, protein/protein, and protein/CaCO3. The N-terminal half of this protein, n16N, is known to be active in conferring this mineral stabilization and organization. While some details relating to the stabilization and organization of the mineral are known, the molecular mechanisms that underpin these processes are not yet established. To provide these molecular-scale details, here we explore current hypotheses regarding the possible subdomain organization of n16N, as related to these three interfaces in nacre, by combining outcomes of Replica Exchange with Solute Tempering molecular dynamics simulations with NMR experiments, to investigate the conformational ensemble of n16N in solution. We verify that n16N lacks a well-defined secondary structure, both with and without the presence of Ca(2+) ions, as identified from previous experiments. Our data support the presence of three different, functional subdomains within n16N. Our results reveal that tyrosine, chiefly located in the center of the peptide, plays a multifunctional role in stabilizing conformations of n16N, for intrapeptide and possibly interpeptide interactions. Complementary NMR spectroscopy data confirm the participation of tyrosine in this stabilization. The C-terminal half of n16N, lacking in tyrosine and highly charged, shows substantive conformational diversity and is proposed as a likely site for nucleation of calcium carbonate. Finally, dominant structures from our predicted conformational ensemble suggest the presentation of key residues thought to be critical to the selective binding to β-chitin surfaces.

  9. Purification, Conformational Analysis, and Properties of a Family of Tigerinin Peptides from Skin Secretions of the Crowned Bullfrog Hoplobatrachus occipitalis.

    PubMed

    McLaughlin, Christopher M; Lampis, Sandrina; Mechkarska, Milena; Coquet, Laurent; Jouenne, Thierry; King, Jay D; Mangoni, Maria Luisa; Lukic, Miodrag L; Scorciapino, Mariano A; Conlon, J Michael

    2016-09-23

    Four host-defense peptides belonging to the tigerinin family (tigerinin-1O: RICTPIPFPMCY; tigerinin-2O: RTCIPIPLVMC; tigerinin-3O: RICTAIPLPMCL; and tigerinin-4O: RTCIPIPPVCF) were isolated from skin secretions of the African crowned bullfrog Hoplobatrachus occipitalis. In aqueous solution at pH 4.8, the cyclic domain of tigerinin-2O adopts a rigid amphipathic conformation that incorporates a flexible N-terminal tail. The tigerinins lacked antimicrobial (MIC > 100 μM) and hemolytic (LC50 > 500 μM) activities but, at a concentration of 20 μg/mL, significantly (P < 0.05) inhibited production of interferon-γ (IFN-γ) by peritoneal cells from C57BL/6 mice without affecting production of IL-10 and IL-17. Tigerinin-2O and -4O inhibited IFN-γ production at concentrations as low as 1 μg/mL. The tigerinins significantly (P ≤ 0.05) stimulated the rate of insulin release from BRIN-BD11 clonal β-cells without compromising the integrity of the plasma membrane. Tigerinin-1O was the most potent (threshold concentration 1 nM) and the most effective (395% increase over basal rate at a concentration of 1 μM). Tigerinin-4O was the most potent and effective peptide in stimulating the rate of glucagon-like peptide-1 release from GLUTag enteroendocrine cells (threshold concentration 10 nM; 289% increase over basal rate at 1 μM). Tigerinin peptides have potential for development into agents for the treatment of patients with type 2 diabetes.

  10. Acidic pH triggers conformational changes at the NH2-terminal propeptide of the precursor of pulmonary surfactant protein B to form a coiled coil structure.

    PubMed

    Bañares-Hidalgo, A; Pérez-Gil, J; Estrada, P

    2014-07-01

    Pulmonary surfactant protein SP-B is synthesized as a larger precursor, proSP-B. We report that a recombinant form of human SP-BN forms a coiled coil structure at acidic pH. The protonation of a residue with pK=4.8±0.06 is the responsible of conformational changes detected by circular dichroism and intrinsic fluorescence emission. Sedimentation velocity analysis showed protein oligomerisation at any pH condition, with an enrichment of the species compatible with a tetramer at acidic pH. Low 2,2,2,-trifluoroethanol concentration promoted β-sheet structures in SP-BN, which bind Thioflavin T, at acidic pH, whereas it promoted coiled coil structures at neutral pH. The amino acid stretch predicted to form β-sheet parallel association in SP-BN overlaps with the sequence predicted by several programs to form coiled coil structure. A synthetic peptide ((60)W-E(85)) designed from the sequence of the amino acid stretch of SP-BN predicted to form coiled coil structure showed random coil conformation at neutral pH but concentration-dependent helical structure at acidic pH. Sedimentation velocity analysis of the peptide indicated monomeric state at neutral pH (s20, w=0.55S; Mr~3kDa) and peptide association (s20, w=1.735S; Mr=~14kDa) at acidic pH, with sedimentation equilibrium fitting to a Monomer-Nmer-Mmer model with N=6 and M=4 (Mr=14692Da). We propose that protein oligomerisation through coiled-coil motifs could then be a general feature in the assembly of functional units in saposin-like proteins in general and in the organization of SP-B in a functional surfactant, in particular.

  11. [Amino acid composition of polynucleotide-peptide complexes isolated from algae].

    PubMed

    Pusheva, M A; Khoreva, M A

    1977-01-01

    The amino acid composition of the peptide moiety of sulphur containing polynucleotide-peptide complexes (S-PNPC) was studied with four species of green algae and two species of blue-green algae. S-PNPC contained almost all amino acids which were usually encountered in acid hydrolysates of proteins, and also unidentified ninhydrin-positive compounds. The amino acid composition of the peptide moiety of S-PNPC was rather similar in the case of different algae, and was characterized by the prevailence of acid amino acids.

  12. Tuning the biological properties of amphipathic alpha-helical antimicrobial peptides: rational use of minimal amino acid substitutions.

    PubMed

    Zelezetsky, Igor; Pag, Ulrike; Sahl, Hans-Georg; Tossi, Alessandro

    2005-12-01

    In nature, alpha-helical antimicrobial peptides present the small and flexible residue glycine at positions 7 or 14 with a significant frequency. Based on the sequence of the non-proteinogenic alpha-helical model peptide P1(Aib7), with a potent, broad spectrum antimicrobial activity, six peptides were designed by effecting a single amino acid substitution to investigate how tuning the structural characteristics at position 7 could lead to optimization of selectivity without affecting antimicrobial activity against a broad panel of multidrug resistant bacterial and yeast indicator strains. The relationship between structural features (size/hydrophobicity of the side chain as well as conformation and flexibility) and biological activity, in terms of minimum inhibitory concentration, membrane permeabilization kinetics and lysis of red blood cells are discussed. On conversion of the peptide to proteinogenic residues, these principles allowed development of a potent antimicrobial peptide with a reduced cytotoxicity. However, while results suggest that both hydrophobicity of residue 7 and chain flexibility at this position can be modulated to improve selectivity, position 14 is less tolerant of substitutions.

  13. Ligand specificity and conformational stability of human fatty acid-binding proteins.

    PubMed

    Zimmerman, A W; van Moerkerk, H T; Veerkamp, J H

    2001-09-01

    Fatty acid binding proteins (FABPs) are small cytosolic proteins with virtually identical backbone structures that facilitate the solubility and intracellular transport of fatty acids. At least eight different types of FABP occur, each with a specific tissue distribution and possibly with a distinct function. To define the functional characteristics of all eight human FABPs, viz. heart (H), brain (B), myelin (M), adipocyte (A), epidermal (E), intestinal (I), liver (L) and ileal lipid-binding protein (I-LBP), we studied their ligand specificity, their conformational stability and their immunological crossreactivity. Additionally, binding of bile acids to I-LBP was studied. The FABP types showed differences in fatty acid binding affinity. Generally, the affinity for palmitic acid was lower than for oleic and arachidonic acid. All FABP types, except E-FABP, I-FABP and I-LBP interacted with 1-anilinonaphtalene-8-sulphonic acid (ANS). Only L-FABP, I-FABP and M-FABP showed binding of 11-((5-dimethylaminonaphtalene-1-sulfonyl)amino)undecanoic acid (DAUDA). I-LBP showed increasing binding of bile acids in the order taurine-conjugated>glycine-conjugated>unconjugated bile acids. A hydroxylgroup of bile acids at position 7 decreased and at position 12 increased the binding affinity to I-LBP. The fatty acid-binding affinity and the conformation of FABP types were differentially affected in the presence of urea. Our results demonstrate significant differences in ligand binding, conformational stability and surface properties between different FABP types which may point to a specific function in certain cells and tissues. The preference of I-LBP (but not L-FABP) for conjugated bile acids is in accordance with a specific role in bile acid reabsorption in the ileum.

  14. Conjugation of synthetic peptides to proteins: quantitation from S-carboxymethylcysteine released upon acid hydrolysis.

    PubMed

    Kolodny, N; Robey, F A

    1990-05-15

    A method described here for conjugating synthetic peptides to carrier proteins provides a convenient method for determining peptide-to-carrier protein ratios. N-Bromoacetyl-containing peptides are reacted in situ with carrier proteins in which the disulfide bonds were reduced with tri-n-butylphosphine. At pH 7-8 and ambient temperature, the newly formed sulfhydryl groups of the carrier protein react exclusively with the bromoacetyl mokiety of the peptide to form conjugates having stable thio ether linkages. Acid hydrolyses of these conjugates release S-carboxymethylcysteine in amounts proportional to the amounts of peptides conjugated and thus allow determination of peptide-to-protein ratios.

  15. Conformation of the phosphate D-alanine zwitterion in bacterial teichoic acid from nuclear magnetic resonance spectroscopy.

    PubMed

    Garimella, Ravindranath; Halye, Jeffrey L; Harrison, William; Klebba, Phillip E; Rice, Charles V

    2009-10-06

    The conformation of d-alanine (d-Ala) groups of bacterial teichoic acid is a central, yet untested, paradigm of microbiology. The d-Ala binds via the C-terminus, thereby allowing the amine to exist as a free cationic NH(3)(+) group with the ability to form a contact ion pair with the nearby anionic phosphate group. This conformation hinders metal chelation by the phosphate because the zwitterion pair is charge neutral. To the contrary, the repulsion of cationic antimicrobial peptides (CAMPs) is attributed to the presence of the d-Ala cation; thus the ion pair does not form in this model. Solid-state nuclear magnetic resonance (NMR) spectroscopy has been used to measure the distance between amine and phosphate groups within cell wall fragments of Bacillus subtilis. The bacteria were grown on media containing (15)N d-Ala and beta-chloroalanine racemase inhibitor. The rotational-echo double-resonance (REDOR) pulse sequence was used to measure the internuclear dipolar coupling, and the results demonstrate (1) the metal-free amine-to-phosphate distance is 4.4 A and (2) the amine-to-phosphate distance increases to 5.4 A in the presence of Mg(2+) ions. As a result, the zwitterion exists in a nitrogen-oxygen ion pair configuration providing teichoic acid with a positive charge to repel CAMPs. Additionally, the amine of d-Ala does not prevent magnesium chelation in contradiction to the prevailing view of teichoic acids in metal binding. Thus, the NMR-based description of teichoic acid structure resolves the contradictory models, advances the basic understanding of cell wall biochemistry, and provides possible insight into the creation of new antibiotic therapies.

  16. Conformation of the Phosphate D-alanine Zwitterion in Bacterial Teichoic Acid from Nuclear Magnetic Resonance Spectroscopy

    PubMed Central

    Garimella, Ravindranath; Halye, Jeffrey L.; Harrison, William; Klebba, Phillip E.; Rice, Charles V.

    2009-01-01

    The conformation of D-alanine (D-Ala) groups of bacterial teichoic acid is a central, yet untested, paradigm of microbiology. The D-Ala binds via the C-terminus, thereby allowing the amine to exist as a free cationic NH3+ group with the ability to form a contact-ion-pair with the nearby anionic phosphate group. This conformation hinders metal chelation by the phosphate because the zwitterion pair is charge neutral. To the contrary, the repulsion of cationic antimicrobial peptides (CAMPs) is attributed to the presence of the D-Ala cation; thus the ion-pair does not form in this model. Solid-state nuclear magnetic resonance (NMR) spectroscopy has been used to measure the distance between amine and phosphate groups within cell wall fragments of Bacillus subtilis. The bacteria were grown on media containing 15N D-Ala and β-chloroalanine racemase inhibitor. The rotational-echo double-resonance (REDOR) pulse sequence was used to measure the internuclear dipolar coupling and the results demonstrate: 1) the metal-free amine-to-phosphate distance is 4.4 Å and 2) the amine-to-phosphate distance increases to 5.4 Å in the presence of Mg2+ ions. As a result, the zwitterion exists in a nitrogen-oxygen ion-pair configuration providing teichoic acid with a positive charge to repel CAMPs. Additionally, the amine of D-Ala does not prevent magnesium chelation in contradiction to the prevailing view of teichoic acids in metal binding. Thus, the NMR-based description of teichoic acid structure resolves the contradictory models, advances the basic understanding of cell wall biochemistry, and provides possible insight into the creation of new antibiotic therapies. PMID:19746945

  17. Conformational preferences of synthetic peptides derived from the immunodominant site of the circumsporozoite protein of Plasmodium falciparum by sup 1 H NMR

    SciTech Connect

    Dyson, H.J.; Satterthwait, A.C.; Lerner, R.A.; Wright, P.E. )

    1990-08-28

    Proton nuclear magnetic resonance and ultraviolet circular dichroism spectroscopy have been used to probe the conformational ensemble of the tandemly repeated tetrapeptide unit of the circumsporozoite coat protein of the malaria parasite Plasmodium falciparum. Peptides based on the Asn-Ala-Asn-Pro and Asn-Pro-Asn-Ala cadences and composed of one to three tetrapeptide units were synthesized and examined using one- and two-dimensional NMR spectroscopy. The chemical shift of the amide protons, the temperature dependence of the amide proton chemical shift, and the patterns of NOE connectivities in the various peptides give evidence for the presence of a substantial population of folded conformers in several of the peptides in water solution at pH 5.0. Correlations between the behavior of the tandemly repeated units in different peptides have been used to infer the structure(s) of the folded conformers. The data are consistent with the presence of turnlike structures stabilized by hydrogen bonding of the backbone amid protons of the alanines and the asparagine residues preceding them. Specific differences in the strengths of NOEs between peptides of different lengths indicate that the folded structure is considerably stabilized by the presence of the asparagine residue following the alanine. Differences between peptides with different cadences of the tandemly repeating unit indicate that a repeating structural motif is formed by the Asn-Pro-Asn-Ala-(Asn) cadence.

  18. Conformational stability and vibrational spectrum of glyoxilic acid oxime predicted from ab initio study

    NASA Astrophysics Data System (ADS)

    Trendafilova, N.; Bauer, G.; Georgieva, I.; Delchev, V.

    2002-02-01

    The conformational stability of glyoxilic acid oxime (HOOC-CH-NOH) (GAO) and its anions has been studied by ab initio calculations at different levels of the theory, HF/6-311G ∗∗, MP2/6-311G ∗∗ and B3LYP/6-311G ∗∗. Geometry optimization was performed for 16 conformations of GAO and five anions in Cs symmetry. The interconversion pathways for the four lowest energy conformers as well as the corresponding transition states have been investigated using QST3 and IRC techniques. The minima and the transition states obtained were estimated by calculations of the vibrational frequencies. The energy barriers of three interconversions, ectt-ecct, ectt-ettt and ectt-zccc, have been estimated. Vibrational spectra and IR intensities of the lowest energy conformers, zccc, ectt, ettt and ecct, have been calculated and discussed at HF/6-311G ∗∗ optimized geometries.

  19. Conformational analysis of a nucleoside of 1,4-dihydro-4-oxoquinoline-3-carboxylic acid analogue

    NASA Astrophysics Data System (ADS)

    Zaccur Leal, Kátia; Rudolf Seidl, Peter; Diniz Yoneda, Julliane; Santos, CarlaV. B.; Marques, Isakelly P.; Souza, Maria Cecília B. V.; Francisco Ferreira, Vitor

    2005-06-01

    The synthesis of new ribonucleosides is an essential research area in the investigation of new therapeutically useful agents, particularly those used in the treatment of HIV infection. The conformation of these nucleosides may have direct implications for their ability to bind to receptor targets. We have prepared the 7-methoxy-1,4-dihydro-4-oxoquinoline-3-carboxylic acid derivatives and used the ensemble of low-energy minima to develop conformational profiles of quinolonic nucleosides and verify their accuracy in different calculations of structural parameters. Results are compared with experimental data obtained by X-ray and NMR analysis. Finally, we intend to test the applicability of these methods to conformational analysis of other nucleosides and verify if the preferential conformation is the one which gives the best anti-HIV or antiviral activity.

  20. Amino acid sequence of atrial natriuretic peptides in human coronary sinus plasma.

    PubMed

    Yandle, T; Crozier, I; Nicholls, G; Espiner, E; Carne, A; Brennan, S

    1987-07-31

    Two atrial natriuretic peptides were purified from pooled human coronary sinus plasma by Sep-Pak extraction, immunoaffinity chromatography and reverse phase HPLC. The amino acid sequences of the two peptides were homologous with 99-126 human atrial natriuretic peptide (hANP) and 106-126 hANP, the latter being most probably linked to 99-105 ANP by the disulphide bond. The molar ratio of the peptides in plasma, as assessed by radioimmunoassay was 10:3.

  1. Diaminopyridine-Based Potent and Selective Mps1 Kinase Inhibitors Binding to an Unusual Flipped-Peptide Conformation

    PubMed Central

    2012-01-01

    Monopolar spindle 1 (Mps1) is an attractive cancer drug target due to the important role that it plays in centrosome duplication, the spindle assembly checkpoint, and the maintenance of chromosomal stability. A design based on JNK inhibitors with an aminopyridine scaffold and subsequent modifications identified diaminopyridine 9 with an IC50 of 37 nM. The X-ray structure of 9 revealed that the Cys604 carbonyl group of the hinge region flips to form a hydrogen bond with the aniline NH group in 9. Further optimization of 9 led to 12 with improved cellular activity, suitable pharmacokinetic profiles, and good in vivo efficacy in the mouse A549 xenograft model. Moreover, 12 displayed excellent selectivity over 95 kinases, indicating the contribution of its unusual flipped-peptide conformation to its selectivity. PMID:24900510

  2. Influence of polar side chains modifications on the dual enkephalinase inhibitory activity and conformation of human opiorphin, a pain perception related peptide.

    PubMed

    Rosa, Mònica; Marcelo, Filipa; Calle, Luis P; Rougeot, Catherine; Jiménez-Barbero, Jesús; Arsequell, Gemma; Valencia, Gregorio

    2015-11-15

    The dual inhibitory action of the pain related peptide opiorphin (H-Gln-Arg-Phe-Ser-Arg-OH) against neutral endopeptidase (NEP) and aminopeptidase N (AP-N) was further investigated by a SAR study involving minor modifications on the polar side chains of Arg residues and glycosylation with monosaccharides at Ser. None of them exerted dual or individual inhibitory potency superior than opiorphin. However, the correlations deduced offer further proof for the key role of these residues upon the binding and bioactive conformational stabilization of opiorphin. NMR conformational studies on the glycopeptides suggest that they are still very flexible compounds that may attain their respective bioactive conformations.

  3. NIR Laser Radiation Induced Conformational Changes and Tunneling Lifetimes of High-Energy Conformers of Amino Acids in Low-Temperature Matrices

    NASA Astrophysics Data System (ADS)

    Bazso, Gabor; Najbauer, Eszter E.; Magyarfalvi, Gabor; Tarczay, Gyorgy

    2013-06-01

    We review our recent results on combined matrix isolation FT-IR and NIR laser irradiation studies on glycine alanine, and cysteine. The OH and the NH stretching overtones of the low-energy conformers of these amino acids deposited in Ar, Kr, Xe, and N_{2} matrices were irradiated. At the expense of the irradiated conformer, other conformers were enriched and new, high-energy, formerly unobserved conformers were formed in the matrices. This enabled the separation and unambiguous assignment of the vibrational transitions of the different conformers. The main conversion paths and their efficiencies are described qualitatively showing that there are significant differences in different matrices. It was shown that the high-energy conformer decays in the matrix by H-atom tunneling. The lifetimes of the high-energy conformers in different matrices were measured. Based on our results we conclude that some theoretically predicted low-energy conformers of amino acids are likely even absent in low-energy matrices due to fast H-atom tunneling. G. Bazso, G. Magyarfalvi, G. Tarczay J. Mol. Struct. 1025 (Light-Induced Processes in Cryogenic Matrices Special Issue) 33-42 (2012). G. Bazso, G. Magyarfalvi, G. Tarczay J. Phys. Chem. A 116 (43) 10539-10547 (2012). G. Bazso, E. E. Najbauer, G. Magyarfalvi, G. Tarczay J. Phys. Chem. A in press, DOI: 10.1021/jp400196b. E. E. Najbauer, G. Bazso, G. Magyarfalvi, G. Tarczay in preparation.

  4. Proton and tritium NMR relaxation studies of peptide inhibitor binding to bacterial collagenase: Conformation and dynamics

    SciTech Connect

    Dive, V.; Lai, A.; Valensin, G.; Saba, G.; Yiotakis, A.; Toma, F. )

    1991-02-15

    The interaction of succinyl-Pro-Ala, a competitive inhibitor of Achromobacter iophagus collagenase, with the enzyme was studied by longitudinal proton and tritium relaxation. Specific deuterium and tritium labeling of the succinyl part at vicinal positions allowed the measurement of the cross-relaxation rates of individual proton or tritium spin pairs in the inhibitor-enzyme complex as well as in the free inhibitor. Overall correlation times, internuclear distances, and qualitative information on the internal mobility in Suc1 (as provided by the generalized order parameter S2) could be deduced by the comparison of proton and tritium cross-relaxation of spin pairs at complementary positions in the -CH2- CH2- moiety as analyzed in terms of the model-free approach by Lipari and Szabo. The conformational and motional parameters of the inhibitor in the free and enzyme-bound state were directly compared by this method. The measurement of proton cross-relaxation in the Ala residue provided additional information on the inhibitor binding. The determination of the order parameter in different parts of the inhibitor molecule in the bound state indicates that the succinyl and alanyl residues are primarily involved in the interaction with the enzyme activity site. The succinyl moiety, characterized in solution by the conformational equilibrium among the three staggered rotamers--i.e., trans: 50%; g+: 20%; g-: 30%--adopted in the bound state the unique trans conformation.

  5. Quantum Corrections to the Free Energy Difference between Peptides and Proteins Conformers.

    PubMed

    Cecchini, Marco

    2015-09-08

    The calculation of the free energy of conformation is key in understanding the function of biomolecules and has attracted significant interest in recent years. Most current computational approaches evaluate the difference in conformational free energy in the classical limit based on the common "dogma" that only the lowest-frequency modes make a significant contribution to it, i.e. they assume that quantum mechanical corrections are negligible. Here, I show for three biomolecular systems described in the rigid-rotor, harmonic-oscillator approximation that the zero-point energy contribution, although small, is not negligible even at room temperature. I find that a quantum correction arises from the intermediate-frequency vibrational modes and that its magnitude is strongly correlated with the number of atoms in the system. A straightforward, though approximate, way to account for this quantum correction in the calculation of conformational free-energy differences by classical molecular dynamics is presented. The relevance of the quantum correction analyzed in this paper is discussed in the context of conventional force fields for proteins.

  6. Conformation of the ATP binding peptide in actin revealed by proton NMR spectroscopy

    SciTech Connect

    Barden, J.A.

    1987-09-22

    The actin peptide 106-124 exists in a completely conserved region of the sequence and binds strongly to both ATP and tripolyphosphate. Binding particularly affects residues 116 and 118 and generally affects the two segments 115-118 and 121-124. One-dimensional nuclear Overhauser enhancement difference spectroscopy was used to detect molecular interactions between both adjacent and nonadjacent residues. The N-terminal segment 106-112 was found to be largely extended. A sharp bend was detected between Pro-112 and Lys-113. The triphosphate moiety binds to the strongly hydrophilic central segment of the peptide. Evidence was obtained for a reverse turn involving residues 121-124. Amide proton temperature coefficients and coupling constants provide evidence for a type I ..beta..-turn. A model of the ATP binding site is proposed together with its relationship to other parts of the actin structure and to the phalloidin binding site.

  7. Conformationally restricted C-terminal peptides of substance P. Synthesis, mass spectral analysis and pharmacological properties.

    PubMed

    Theodoropoulos, D; Poulos, C; Gatos, D; Cordopatis, P; Escher, E; Mizrahi, J; Regoli, D; Dalietos, D; Furst, A; Lee, T D

    1985-10-01

    Four cyclic analogues of the C-terminal hepta- or hexapeptide of substance P were prepared by the solution method. The cyclizations were obtained by substituting with cysteine the residues normally present in positions 5 or 6 or 11 of substance P and by subsequent disulfide bond formation. The final products were identified by ordinary analytical procedures and advanced mass spectroscopy. The biological activities were determined on three bioassays: the guinea pig ileum, the guinea pig trachea and the rabbit mesenteric vein. Results obtained with these assays indicate that all peptides with a disulfide bridgehead in position 11 are inactive and that a cycle between positions 5 and 6 already strongly reduces the biological activity. The acyclic precursors containing thiol protection groups display weak biological activities. These results further underline the importance of the side chain in position 11 of substance P and suggest that optimal biological activities may require a linear peptide sequence.

  8. Peptide backbone folding induced by the C(alpha)-tetrasubstituted cyclic alpha-amino acids 4-amino-1,2-dithiolane-4-carboxylic acid (Adt) and 1-aminocyclopentane-1-carboxylic acid (Ac5c). A joint computational and experimental study.

    PubMed

    Aschi, Massimiliano; Lucente, Gino; Mazza, Fernando; Mollica, Adriano; Morera, Enrico; Nalli, Marianna; Paglialunga Paradisi, Mario

    2003-06-07

    The conformational study of a new group of synthetic peptides containing 4-amino-1,2-dithiolane-4-carboxylic acid (Adt), a cysteine-related achiral residue, has been carried out through a joint application of computational and experimental methodologies. Molecular Dynamics simulations clearly suggest the tendency of this molecule to adopt a gamma-turn conformation in vacuum and help in analyzing the complex and crucial conformational behaviour of the dithiolane ring which appears to preferentially adopt a C(S)-like structure. Electronic structure calculations carried out in solution using the Density Functional Theory also indicate the preservation of the gamma-like folding in apolar solvents and the helix-like one in more polar solvents. A comparison with the achiral 1-aminocycloalkane-1-carboxylic acid (Ac5c) has been carried out using the same computational tools. NMR and IR data on dipeptide derivatives containing the Adt or Ac5c residue show that in chloroform solution all the models prefer a gamma-turn structure, centered at the cyclic residue, stabilized by an intramolecular H-bond, whereas in a more polar solvent, i.e. dimethyl sulfoxide, this folding is not maintained. The experimental conformational studies, extended to N-Boc protected tripeptides, clearly indicate the remarkable tendency of both the five-membered C(alpha)-tetrasubstituted cyclic amino acids Adt and Ac5c to induce the gamma-turn structure also in models able to adopt the beta-bend conformation.

  9. Conformational analysis of a toxic peptide from Trimeresurus wagleri which blocks the nicotinic acetylcholine receptor.

    PubMed Central

    Sellin, L C; Mattila, K; Annila, A; Schmidt, J J; McArdle, J J; Hyvönen, M; Rantala, T T; Kivistö, T

    1996-01-01

    The 22-residue toxic peptide (WTX1) from the venom of the Southeast Asian snake Trimeresurus wagleri has multiple sites of action, but its lethal effect has been attributed to blocking the postsynaptic acetylcholine receptor at the neuromuscular junction. The 3-dimensional structure of WTX1 was studied using 2-dimensional nuclear magnetic resonance spectroscopy, circular dichroism, and computer simulations. In aqueous solution, WTX1 was shown to have extended and flexible "tails" defined by a short, rigid disulfide-bonded loop. The flexible regions can undergo structural rearrangement when moved from an aqueous to a less polar environment and may contribute to its effectiveness at different receptor sites. By substituting Gly or Phe for His at position 10, significant effects on the disulfide bond formation and, thereby, the activity of the peptide were observed. These results suggest that even subtle differences in single residues can have profound effects on the dynamics of folding, disulfide bond formation, and activity of this toxic peptide. Images FIGURE 10 FIGURE 12 PMID:8770182

  10. Di-heterometalation of thiol-functionalized peptide nucleic acids

    PubMed Central

    Joshi, Tanmaya; Patra, Malay; Spiccia, Leone; Gasser, Gilles

    2013-01-01

    As a proof-of-principle, two hetero-bimetallic PNA oligomers containing a ruthenium(II) polypyridyl and a cyclopentadienyl manganese tricarbonyl complex have been prepared by serial combination of solid-phase peptide coupling and in-solution thiol chemistry. Solid-phase N-terminus attachment of Ru(II)-polypyridyl carboxylic acid derivative, C1, onto the thiol-functionalized PNA backbone (H-a-a-g-t-c-t-g-c-linker-cys-NH2) has been performed by standard peptide coupling method. As two parallel approaches, the strong affinity of thiols for maleimide and haloacetyl group has been exploited for subsequent post-SPPS addition of cymantrene-based organometallic cores, C2 and C3. Michael-like addition and thioether ligation of thiol functionalized PNA1 (H-gly-a-a-g-t-c-t-g-c-linker-cys-NH2) and PNA2 (C1-a-a-g-t-c-t-g-c-linker-cys-NH2) to cymantrene maleimide and chloroacetyl derivatives, C2 and C3, respectively, has been performed. The synthesized ruthenium(II)-cymantrenyl PNA oligomers have been characterized by mass spectrometry (ESI-MS) and IR spectroscopy. The distinct Mn-CO vibrational IR stretches, between 1,924–2,074 cm−1, have been used as markers to confirm the presence of cymantrenyl units in the PNA sequences and the purity of the HPLC-purified PNA thioethers assessed using LC-MS. PMID:23422249

  11. Formation of taste-active amino acids, amino acid derivatives and peptides in food fermentations - A review.

    PubMed

    Zhao, Cindy J; Schieber, Andreas; Gänzle, Michael G

    2016-11-01

    Fermented foods are valued for their rich and complex odour and taste. The metabolic activity of food-fermenting microorganisms determines food quality and generates odour and taste compounds. This communication reviews the formation of taste-active amino acids, amino acid derivatives and peptides in food fermentations. Pathways of the generation of taste compounds are presented for soy sauce, cheese, fermented meats, and bread. Proteolysis or autolysis during food fermentations generates taste-active amino acids and peptides; peptides derived from proteolysis particularly impart umami taste (e.g. α-glutamyl peptides) or bitter taste (e.g. hydrophobic peptides containing proline). Taste active peptide derivatives include pyroglutamyl peptides, γ-glutamyl peptides, and succinyl- or lactoyl amino acids. The influence of fermentation microbiota on proteolysis, and peptide hydrolysis, and the metabolism of glutamate and arginine is well understood, however, the understanding of microbial metabolic activities related to the formation of taste-active peptide derivatives is incomplete. Improved knowledge of the interactions between taste-active compounds will enable the development of novel fermentation strategies to develop tastier, less bitter, and low-salt food products, and may provide novel and "clean label" ingredients to improve the taste of other food products. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Synthesis of hybrid hydrazino peptides: protected vs unprotected chiral α-hydrazino acids.

    PubMed

    Suć, Josipa; Jerić, Ivanka

    2015-01-01

    Peptidomimetics based on hydrazino derivatives of α-amino acids represent an important class of peptidic foldamers with promising biological activities, like protease inhibition and antimicrobial activity. However, the lack of straightforward method for the synthesis of optically pure hydrazino acids and efficient incorporation of hydrazino building blocks into peptide sequence hamper wider exploitation of hydrazino peptidomimetics. Here we described the utility of N (α)-benzyl protected and unprotected hydrazino derivatives of natural α-amino acids in synthesis of peptidomimetics. While incorporation of N (α)-benzyl-hydrazino acids into peptide chain and deprotection of benzyl moiety proceeded with difficulties, unprotected hydrazino acids allowed fast and simple construction of hybrid peptidomimetics.

  13. Formic and acetic acids in a nitrogen matrix: Enhanced stability of the higher-energy conformer

    NASA Astrophysics Data System (ADS)

    Lopes, Susy; Domanskaya, Alexandra V.; Fausto, Rui; Räsänen, Markku; Khriachtchev, Leonid

    2010-10-01

    Formic acid (HCOOH, FA) and acetic acid (CH3COOH, AA) are studied in a nitrogen matrix. The infrared (IR) spectra of cis and trans conformers of these carboxylic acids (and also of the HCOOD isotopologue of FA) are reported and analyzed. The higher-energy cis conformer of these molecules is produced by narrowband near-IR excitation of the more stable trans conformer, and the cis-to-trans tunneling decay is evaluated spectroscopically. The tunneling process in both molecules is found to be substantially slower in a nitrogen matrix than in rare-gas matrices, the cis-form decay constants being approximately 55 and 600 times smaller in a nitrogen matrix than in an argon matrix, for FA and AA respectively. The stabilization of the higher-energy cis conformer is discussed in terms of specific interactions with nitrogen molecule binding with the OH group of the carboxylic acid. This model is in agreement with the observed differences in the IR spectra in nitrogen and argon matrices, in particular, the relative frequencies of the νOH and τCOH modes and the relative intensities of the νOH and νCO bands.

  14. The effect of citric acid on the activity, thermodynamics and conformation of mushroom polyphenoloxidase.

    PubMed

    Liu, Wei; Zou, Li-qiang; Liu, Jun-ping; Zhang, Zhao-qin; Liu, Cheng-mei; Liang, Rui-hong

    2013-09-01

    Few reports have focused on the effect of citric acid on thermodynamics and conformation of polyphenoloxidase (PPO). In this study, variations on activity, thermodynamics and conformation of mushroom PPO induced by citric acid (1-60mM) and relationships among these were investigated. It showed that with the increasing concentration of citric acid, the activity of PPO decreased gradually to an inactivity condition; inactivation rate constant (k) of PPO increased and the activation energy (Ea) as well as thermodynamic parameters (ΔG, ΔH, ΔS) decreased, which indicated that the thermosensitivity, stability and number of non-covalent bonds of PPO decreased. The conformation was gradually unfolded, which was reflected in the decrease of α-helix contents, increase of β-sheet and exposure of aromatic amino acid residuals. Moreover, two linear relationships of relative activities, enthalpies (ΔH) against α-helix contents were obtained. It indicated that changes of activity and thermodynamics might correlate to the unfolding of conformation.

  15. Time-measurement-regulating peptide PIN may alter a timer conformation of Time Interval Measuring Enzyme (TIME).

    PubMed

    Ti, Xiaonan; Tani, Naoki; Isobe, Minoru; Kai, Hidenori

    2006-05-01

    The TIME (Time Interval Measuring Enzyme) ATPase measures time intervals in accordance with diapause development, which indispensably requires cold for resumption of embryonic development in the silkworm (Bombyx mori). The PIN (Peptidyl Inhibitory Needle) peptide regulates the time measurement function of TIME. In the present study we investigated the interaction between TIME and PIN in order to address the mechanism of diapause development. When TIME was isolated from eggs later than 12 days after oviposition, transient bursts of ATPase activity occurred 18h after isolation of TIME, and the younger the eggs and pupal ovaries from which TIME was isolated, the earlier the bursts of ATPase activity appeared. However, no interval-timer activation of ATPase occurred in ovaries earlier than 6 days after pupation. Similar patterns of ATPase activity occurred in test tubes after mixing TIME with PIN. The shorter the time PIN was mixed with TIME, the earlier the ATPase activity appeared. The timer may be built into the protein conformation of TIME, and PIN (which is present in ovaries beginning 6 days after pupation) appears able to alter this timer conformation through pupal stages to laid eggs. We discuss the possible mechanism of diapause development in relation to the timer mechanism of TIME.

  16. Glycation induces conformational changes in the amyloid-β peptide and enhances its aggregation propensity: molecular insights.

    PubMed

    Jana, Asis K; Batkulwar, Kedar B; Kulkarni, Mahesh J; Sengupta, Neelanjana

    2016-11-23

    The cytotoxicity of the amyloid beta (Aβ) peptide, implicated in the pathogenesis of Alzheimer's disease (AD), can be enhanced by its post-translational glycation, a series of non-enzymatic reactions with reducing sugars and reactive dicarbonyls. However, little is known about the underlying mechanisms that potentially enhance the cytotoxicity of the advanced glycation modified Aβ. In this work, fully atomistic molecular dynamics (MD) simulations are exploited to obtain direct molecular insights into the process of early Aβ self-assembly in the presence and absence of glycated lysine residues. Analyses of data exceeding cumulative timescales of 1 microsecond for each system reveal that glycation results in a stronger enthalpy of association between Aβ monomers and lower conformational entropy, in addition to a sharp overall increase in the beta-sheet content. Further analyses reveal that the enhanced interactions originate, in large part, due to markedly stronger, as well as new, inter-monomer salt bridging propensities in the glycated variety. Interestingly, these conformational and energetic effects are broadly reflected in preformed protofibrillar forms of Aβ small oligomers modified with glycation. Our combined results imply that glycation consolidates Aβ self-assembly regardless of its point of occurrence in the pathway. They provide a basis for further mechanistic studies and therapeutic endeavors that could potentially result in novel ways of combating AGE related AD progression.

  17. ArrayPitope: Automated Analysis of Amino Acid Substitutions for Peptide Microarray-Based Antibody Epitope Mapping

    PubMed Central

    Hansen, Christian Skjødt; Østerbye, Thomas; Marcatili, Paolo; Lund, Ole; Buus, Søren

    2017-01-01

    Identification of epitopes targeted by antibodies (B cell epitopes) is of critical importance for the development of many diagnostic and therapeutic tools. For clinical usage, such epitopes must be extensively characterized in order to validate specificity and to document potential cross-reactivity. B cell epitopes are typically classified as either linear epitopes, i.e. short consecutive segments from the protein sequence or conformational epitopes adapted through native protein folding. Recent advances in high-density peptide microarrays enable high-throughput, high-resolution identification and characterization of linear B cell epitopes. Using exhaustive amino acid substitution analysis of peptides originating from target antigens, these microarrays can be used to address the specificity of polyclonal antibodies raised against such antigens containing hundreds of epitopes. However, the interpretation of the data provided in such large-scale screenings is far from trivial and in most cases it requires advanced computational and statistical skills. Here, we present an online application for automated identification of linear B cell epitopes, allowing the non-expert user to analyse peptide microarray data. The application takes as input quantitative peptide data of fully or partially substituted overlapping peptides from a given antigen sequence and identifies epitope residues (residues that are significantly affected by substitutions) and visualize the selectivity towards each residue by sequence logo plots. Demonstrating utility, the application was used to identify and address the antibody specificity of 18 linear epitope regions in Human Serum Albumin (HSA), using peptide microarray data consisting of fully substituted peptides spanning the entire sequence of HSA and incubated with polyclonal rabbit anti-HSA (and mouse anti-rabbit-Cy3). The application is made available at: www.cbs.dtu.dk/services/ArrayPitope. PMID:28095436

  18. Peptide Fragments of Odin-Sam1: Conformational Analysis and Interaction Studies with EphA2-Sam.

    PubMed

    Mercurio, Flavia A; Di Natale, Concetta; Pirone, Luciano; Scognamiglio, Pasqualina L; Marasco, Daniela; Pedone, Emilia M; Saviano, Michele; Leone, Marilisa

    2015-07-27

    Odin is a protein belonging to the ANKS family, and has two tandem Sam domains. The first, Odin-Sam1, binds to the Sam domain of the EphA2 receptor (EphA2-Sam); this interaction could be crucial for the regulation of receptor endocytosis and might have an impact on cancer. Odin-Sam1 associates with EphA2-Sam by adopting a "mid-loop/end-helix" model. In this study three peptide sequences, encompassing the mid-loop interacting portion of Odin-Sam1 and its C-terminal α5 helix, were designed. Their conformational properties were analyzed by CD and NMR. In addition, their abilities to interact with EphA2-Sam were investigated by SPR studies. The peptides adopt a predominantly disordered state in aqueous buffer, but a higher helical content is evident in the presence of the cosolvent trifluoroethanol. Dissociation constants towards EphA2-Sam were in the high micromolar range. The structural findings suggest further routes for the design of potential anti-cancer therapeutics as inhibitors of EphA2-Sam heterotypic interactions.

  19. Pyrrolidinyl peptide nucleic acid homologues: effect of ring size on hybridization properties.

    PubMed

    Mansawat, Woraluk; Vilaivan, Chotima; Balázs, Árpád; Aitken, David J; Vilaivan, Tirayut

    2012-03-16

    The effect of ring size of four- to six-membered cyclic β-amino acid on the hybridization properties of pyrrolidinyl peptide nucleic acid with an alternating α/β peptide backbone is reported. The cyclobutane derivatives (acbcPNA) show the highest T(m) and excellent specificity with cDNA and RNA.

  20. Templated Self-Assembly of Dynamic Peptide Nucleic Acids.

    PubMed

    Beierle, John M; Ura, Yasuyuki; Ghadiri, M Reza; Leman, Luke J

    2017-09-06

    Template-directed macromolecule synthesis is a hallmark of living systems. Inspired by this natural process, several fundamentally novel mechanisms for template-directed assembly of nucleic acid analogues have been developed. Although these approaches have broad significance, including potential applications in biotechnology and implications for the origins of life, there are unresolved challenges in how to characterize in detail the complex assembly equilibria associated with dynamic templated reactions. Here we describe mechanistic studies of template-directed dynamic assembly for thioester peptide nucleic acid (tPNA), an informational polymer that responds to selection pressures under enzyme-free conditions. To overcome some of the inherent challenges of mechanistic studies of dynamic oligomers, we designed, synthesized, and implemented tPNA-DNA conjugates. The DNA primer region affords a high level of control over the location and register of the tPNA backbone in relation to the template strand. We characterized the degree and kinetics of dynamic nucleobase mismatch correction at defined backbone positions. Furthermore, we report the fidelity of dynamic assembly in tPNA as a function of position along the peptide backbone. Finally, we present theoretical studies that explore the level of fidelity that can be expected for an oligomer having a given hybridization affinity in dynamic templated reactions and provide guidance for the future development of sequence self-editing polymers and materials. As our results demonstrate, the use of molecular conjugates of constitutionally static and dynamic polymers establishes a new methodology for expediting the characterization of the complex chemical equilibria that underlie the assembly of dynamic informational polymers.

  1. Characterization of desmoglein-3 epitope region peptides as synthetic antigens: analysis of their in vitro T cell stimulating efficacy, cytotoxicity, stability, and their conformational features.

    PubMed

    Szabados, Hajnalka; Uray, Katalin; Majer, Zsuzsa; Silló, Pálma; Kárpáti, Sarolta; Hudecz, Ferenc; Bősze, Szilvia

    2015-09-01

    Desmoglein-3 (Dsg3) adhesion protein is the main target of autoantibodies and autoreactive T cells in Pemphigus vulgaris (PV) autoimmune skin disorder. Several mapping studies of Dsg3 T cell epitope regions were performed, and based on those data, we designed and synthesized four peptide series corresponding to Dsg3 T cell epitope regions. Each peptide series consists of a 17mer full-length peptide (Dsg3/189-205, Dsg3/206-222, Dsg3/342-358, and Dsg3/761-777) and its N-terminally truncated derivatives, resulting in 15 peptides altogether. The peptides were prepared on solid phase and were chemically characterized. In order to establish a structure-activity relationship, the solution conformation of the synthetic peptides has been investigated using electronic circular dichroism spectroscopy. The in vitro T cell stimulating efficacy of the peptides has been determined on peripheral blood mononuclear cells isolated from whole blood of PV patients and also from healthy donors. After 20 h of stimulation, the interferon (IFN)-γ content of the supernatants was measured by enzyme-linked immunosorbent assay. In the in vitro conditions, peptides were stable and non-cytotoxic. The in vitro IFN-γ production profile of healthy donors and PV patients, induced by peptides as synthetic antigens, was markedly different. The most unambiguous differences were observed after stimulation with 17mer peptide Dsg3/342-358, and three truncated derivatives from two other peptide series, namely, peptides Dsg3/192-205, Dsg3/763-777, and Dsg3/764-777. Comparative analysis of in vitro activity and the capability of oligopeptides to form ordered or unordered secondary structure showed that peptides bearing high solvent sensibility and backbone flexibility were the most capable to distinguish between healthy and PV donors.

  2. Intrinsic propensities of amino acid residues in GxG peptides inferred from amide I' band profiles and NMR scalar coupling constants.

    PubMed

    Hagarman, Andrew; Measey, Thomas J; Mathieu, Daniel; Schwalbe, Harald; Schweitzer-Stenner, Reinhard

    2010-01-20

    A reliable intrinsic propensity scale of amino acid residues is indispensable for an assessment of how local conformational distributions in the unfolded state can affect the folding of peptides and proteins. Short host-guest peptides, such as GxG tripeptides, are suitable tools for probing such propensities. To explore the conformational distributions sampled by the central amino acid residue in these motifs, we combined vibrational (IR, Raman, and VCD) with NMR spectroscopy. The data were analyzed in terms of a superposition of two-dimensional Gaussian distribution functions in the Ramachandran space pertaining to subensembles of polyproline II, beta-strand, right- and left-handed helical, and gamma-turn-like conformations. The intrinsic propensities of eight amino acid residues (x = A, V, F, L, S, E, K, and M) in GxG peptides were determined as mole fractions of these subensembles. Our results show that alanine adopts primarily (approximately 80%) a PPII-like conformation, while valine and phenylalanine were found to sample PPII and beta-strand-like conformations equally. The centers of the respective beta-strand distributions generally do not coincide with canonical values of dihedral angles of residues in parallel or antiparallel beta-strands. In fact, the distributions for most residues found in the beta-region significantly overlap the PPII-region. A comparison with earlier reported results for trivaline reveals that the terminal valines increase the beta-strand propensity of the central valine residue even further. Of the remaining investigated amino acids, methionine preferred PPII the most (0.64), and E, S, L, and K exhibit moderate (0.56-0.45) PPII propensities. Residues V, F, S, E, and L sample, to a significant extent, a region between the canonical PPII and (antiparallel) beta-strand conformations. This region coincides with the sampling reported for L and V using theoretical predictions (Tran et al. Biochemistry 2005, 44, 11369). The distributions of

  3. A dominant conformational role for amino acid diversity in minimalist protein–protein interfaces

    SciTech Connect

    Gilbreth, Ryan N.; Esaki, Kaori; Koide, Akiko; Sidhu, Sachdev S.; Koide, Shohei

    2008-08-01

    Recent studies have shown that highly simplified interaction surfaces consisting of combinations of just two amino acids, Tyr and Ser, exhibit high affinity and specificity. The high functional levels of such minimalist interfaces might thus indicate small contributions of greater amino acid diversity seen in natural interfaces. Toward addressing this issue, we have produced a pair of binding proteins built on the fibronectin type III scaffold, termed “monobodies.” One monobody contains the Tyr/Ser binary-code interface (termed YS) and the other contains an expanded amino acid diversity interface (YSX), but both bind to an identical target, maltose-binding protein. The YSX monobody bound with higher affinity, a slower off rate and a more favorable enthalpic contribution than the YS monobody. High-resolution X-ray crystal structures revealed that both proteins bound to an essentially identical epitope, providing a unique opportunity to directly investigate the role of amino acid diversity in a protein interaction interface. Surprisingly, Tyr still dominates the YSX paratope and the additional amino acid types are primarily used to conformationally optimize contacts made by tyrosines. Scanning mutagenesis showed that while all contacting Tyr side chains are essential in the YS monobody, the YSX interface was more tolerant to mutations. These results suggest that the conformational, not chemical, diversity of additional types of amino acids provided higher functionality and evolutionary robustness, supporting the dominant role of Tyr and the importance of conformational diversity in forming protein interaction interfaces.

  4. Regulation of sporulation initiation by NprR and its signaling peptide NprRB: molecular recognition and conformational changes.

    PubMed

    Cabrera, Rosina; Rocha, Jorge; Flores, Víctor; Vázquez-Moreno, Luz; Guarneros, Gabriel; Olmedo, Gabriela; Rodríguez-Romero, Adela; de la Torre, Mayra

    2014-11-01

    NprR belongs to the RNPP family of quorum-sensing receptors, a group of intracellular regulators activated directly by signaling oligopeptides in Gram-positive bacteria. In Bacillus thuringiensis (Bt), nprR is located in a transcriptional cassette with nprRB that codes for the precursor of the signaling peptide NprRB. NprR is a transcriptional regulator activated by binding of reimported NprRB; however, several reports suggest that NprR also participates in sporulation but the mechanism is unknown. Our in silico results, based on the structural similarity between NprR from Bt and Spo0F-binding Rap proteins from Bacillus subtilis, suggested that NprR could bind Spo0F to modulate the sporulation phosphorelay in Bt. Deletion of nprR-nprRB cassette from Bt caused a delay in sporulation and defective trigger of the Spo0A∼P-activated genes spoIIA and spoIIIG. The DNA-binding domain of NprR was not necessary for this second function, since truncated NprRΔHTH together with nprRB gene was able to restore the sporulation wild type phenotype in the ΔnprR-nprRB mutant. Fluorescence assays showed direct binding between NprR and Spo0F, supporting that NprR is a bifunctional protein. To understand how the NprR activation by NprRB could result in two different functions, we studied the molecular recognition mechanism between the signaling peptide and the receptor. Using synthetic variants of NprRB, we found that SSKPDIVG displayed the highest affinity (Kd = 7.19 nM) toward the recombinant NprR and demonstrated that recognition involves conformational selection. We propose that the peptide concentration in the cell controls the oligomerization state of the NprR-NprRB complex for switching between its two functions.

  5. Effects of vector fusion peptides on the conformation and immune reactivity of epitope-shuffled, recombinant multi-epitope antigens.

    PubMed

    Wang, Jian; Lin, Yahui; Cai, Pengfei; Wang, Heng

    2011-01-01

    The use of multi-epitopes has been considered as a promising strategy to overcome the obstacle of antigenic variation in malarial vaccine development. Previously, we constructed a multi-epitope artificial antigen, Malaria Random Constructed Antigen-1(M.RCAg-1), to optimize expression of the antigen, and we subcloned the gene into three prokaryotic expression vectors that contain different fusion tags at the N-terminus. Three recombinant proteins expressed by these vectors, named M.RCAg-1/Exp.V-1, V-2, and V-3, were purified after the cleavage of the fusion tag. All three recombinant proteins were able to induce similar levels of antigenicity in BALB/c murine models. However, the antibody responses against the individual epitope peptides of the recombinant products were dramatically different. Additionally, the different epitopes elicited various CD4(+) T-cell responses, as shown by the resulting lymphocyte proliferation and varied IFN-γ and IL-4 levels determined by EILSPOT; however, each could be distinctly recognized by sera derived from malaria patients. Additionally, the rabbit antibody induced by these proteins showed diverse efficacy in malaria parasite growth inhibition assays in vitro. Furthermore, analysis via circular dichroism spectroscopy confirmed that the secondary structure was different among these recombinant proteins. These results suggest that the expressed multi-epitope artificial antigens originating from the different vector fusion peptides indeed affect the protein folding and, subsequently, the epitope exposure. Thus, these proteins are able to induce both distinct humoral and cellular immune responses in animal models, and they affect the efficacy of immune inhibition against the parasite. This work should lead to a further understanding of the impact of vector fusion peptides on the conformation and immune reactivity of recombinant proteins and could provide a useful reference for the development of artificial multi-epitope vaccines.

  6. Uncovering the Relationship between Sulphation Patterns and Conformation of Iduronic Acid in Heparan Sulphate

    NASA Astrophysics Data System (ADS)

    Hsieh, Po-Hung; Thieker, David F.; Guerrini, Marco; Woods, Robert J.; Liu, Jian

    2016-07-01

    The L-iduronic acid (IdoA) residue is a critically important structural component in heparan sulphate polysaccharide for the biological functions. The pyranose ring of IdoA is present in 1C4-chair, 2SO-skew boat, and less frequently, in 4C1-chair conformations. Here, we analyzed the conformation of IdoA residue in eight hexasaccharides by NMR. The data demonstrate a correlation between the conformation of IdoA and sulphations in the surrounding saccharide residues. For the 2-O-sulpho IdoA residue, a high degree of sulphation on neighboring residues drives ring dynamics towards the 2SO-skew boat conformer. In contrast, the nonsulphated IdoA residue is pushed towards the 1C4-chair conformer when the neighboring residues are highly sulphated. Our data suggest that the conformation of IdoA is regulated by the sulphation pattern of nearby saccharides that is genetically controlled by the heparan sulphate biosynthetic pathway.

  7. Uncovering the Relationship between Sulphation Patterns and Conformation of Iduronic Acid in Heparan Sulphate.

    PubMed

    Hsieh, Po-Hung; Thieker, David F; Guerrini, Marco; Woods, Robert J; Liu, Jian

    2016-07-14

    The L-iduronic acid (IdoA) residue is a critically important structural component in heparan sulphate polysaccharide for the biological functions. The pyranose ring of IdoA is present in (1)C4-chair, (2)SO-skew boat, and less frequently, in (4)C1-chair conformations. Here, we analyzed the conformation of IdoA residue in eight hexasaccharides by NMR. The data demonstrate a correlation between the conformation of IdoA and sulphations in the surrounding saccharide residues. For the 2-O-sulpho IdoA residue, a high degree of sulphation on neighboring residues drives ring dynamics towards the (2)SO-skew boat conformer. In contrast, the nonsulphated IdoA residue is pushed towards the (1)C4-chair conformer when the neighboring residues are highly sulphated. Our data suggest that the conformation of IdoA is regulated by the sulphation pattern of nearby saccharides that is genetically controlled by the heparan sulphate biosynthetic pathway.

  8. Prolonged signaling at the parathyroid hormone receptor by peptide ligands targeted to a specific receptor conformation

    PubMed Central

    Okazaki, Makoto; Ferrandon, Sebastien; Vilardaga, Jean-Pierre; Bouxsein, Mary L.; Potts, John T.; Gardella, Thomas J.

    2008-01-01

    The parathyroid hormone receptor (PTHR) is a class B G protein-coupled receptor that plays critical roles in bone and mineral ion metabolism. Ligand binding to the PTHR involves interactions to both the amino-terminal extracellular (N) domain, and transmembrane/extracellular loop, or juxtamembrane (J) regions of the receptor. Recently, we found that PTH(1–34), but not PTH-related protein, PTHrP(1–36), or M-PTH(1–14) (M = Ala/Aib1,Aib3,Gln10,Har11,Ala12,Trp14,Arg19), binds to the PTHR in a largely GTPγS-resistant fashion, suggesting selective binding to a novel, high-affinity conformation (R0), distinct from the GTPγS-sensitive conformation (RG). We examined the effects in vitro and in vivo of introducing the M substitutions, which enhance interaction to the J domain, into PTH analogs extended C-terminally to incorporate residues involved in the N domain interaction. As compared with PTH(1–34), M-PTH(1–28) and M-PTH(1–34) bound to R0 with higher affinity, produced more sustained cAMP responses in cells, formed more stable complexes with the PTHR in FRET and subcellular localization assays, and induced more prolonged calcemic and phosphate responses in mice. Moreover, after 2 weeks of daily injection in mice, M-PTH(1–34) induced larger increases in trabecular bone volume and greater increases in cortical bone turnover, than did PTH(1–34). Thus, the putative R0 PTHR conformation can form highly stable complexes with certain PTH ligand analogs and thereby mediate surprisingly prolonged signaling responses in bone and/or kidney PTH target cells. Controlling, via ligand analog design, the selectivity with which a PTH ligand binds to R0, versus RG, may be a strategy for optimizing signaling duration time, and hence therapeutic efficacy, of PTHR agonist ligands. PMID:18946036

  9. The first experimental observation of the higher-energy trans conformer of trifluoroacetic acid

    NASA Astrophysics Data System (ADS)

    Apóstolo, R. F. G.; Bazsó, Gábor; Bento, R. R. F.; Tarczay, G.; Fausto, R.

    2016-12-01

    We report here the first experimental observation of the higher-energy conformer of trifluoroacetic acid (trans-TFA). The new conformer was generated by selective narrowband near-infrared vibrational excitation of the lower-energy cis-TFA conformer isolated in cryogenic matrices (Ar, Kr, N2) and shown to spontaneously decay to this latter form in the various matrix media, by tunneling. The decay rates in the different matrices were measured and compared with those of the trans conformers of other carboxylic acids in similar experimental conditions. The experimental studies received support from quantum chemistry calculations undertaken at various levels of approximation, which allowed a detailed characterization of the relevant regions of the potential energy surface of the molecule and the detailed assignment of the infrared spectra of the two conformers in the various matrices. Noteworthly, in contrast to cis-TFA that has its trifluoromethyl group eclipsed with the Cdbnd O bond of the carboxylic moiety, trans-TFA has the trifluoromethyl group eclipsed with the Csbnd O bond. This unusual structure of trans-TFA results from the fact that the relative orientation of the CF3 and COOH groups in this geometry facilitates the establishment of an intramolecular hydrogen-bond-like interaction between the OH group and the closely located in-plane fluorine atom of the CF3 moiety.

  10. Helix formation in preorganized beta/gamma-peptide foldamers: hydrogen-bond analogy to the alpha-helix without alpha-amino acid residues.

    PubMed

    Guo, Li; Almeida, Aaron M; Zhang, Weicheng; Reidenbach, Andrew G; Choi, Soo Hyuk; Guzei, Ilia A; Gellman, Samuel H

    2010-06-16

    We report the first high-resolution structural data for the beta/gamma-peptide 13-helix (i,i+3 C=O...H-N H-bonds), a secondary structure that is formed by oligomers with a 1:1 alternation of beta- and gamma-amino acid residues. Our characterization includes both crystallographic and 2D NMR data. Previous studies suggested that beta/gamma-peptides constructed from conformationally flexible residues adopt a different helical secondary structure in solution. Our design features preorganized beta- and gamma-residues, which strongly promote 13-helical folding by the 1:1 beta/gamma backbone.

  11. Apela exhibits isoform- and headgroup-dependent modulation of micelle binding, peptide conformation and dynamics.

    PubMed

    Huang, Shuya K; Shin, Kyungsoo; Sarker, Muzaddid; Rainey, Jan K

    2017-05-01

    Apela (also referred to as ELABELA and toddler) is a peptide hormone that activates the apelin receptor (AR or APJ) to regulate cardiovascular system development and function. Here, we report the first biophysical characterization of three apela isoforms, apela-54, -32, and -11, alongside a monomeric C1S-apela-11 mutant, using circular dichroism (CD) spectropolarimetry and nuclear magnetic resonance (NMR) spectroscopy. The behaviour of apela-54 is consistent with a preprotein containing a hydrophobic, N-terminal signal peptide. The potential for apela-membrane binding, leading to membrane catalyzed interactions with AR, was tested comprehensively for apela-32 and -11 in the presence of membrane-mimetic dodecylphosphocholine (DPC), sodium dodecyl sulfate (SDS), and 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-rac-(1-glycerol)] (LPPG) micelles. According to pulsed-field gradient diffusion NMR experiments, apela-32 interacts with all three micelles. Chemical shift perturbations indicate widespread interactions along apela, with DPC and LPPG micelles inducing short segments with α-helical character at distinct regions. Consistent with these data, ps-ns dynamics along the peptide backbone appear decreased in the presence of micelles. Apela-11 and C1S-apela-11, alternatively, interact preferentially with SDS and LPPG micelles, promoting β-turn character observable by CD. Distinct differences in membrane-interaction propensity are therefore apparent both as a function of apela isoform and of detergent headgroup. These results imply the potential for cell membrane involvement in apela-AR recognition and binding, with the implication that membrane catalysis has distinct functional and regulatory roles throughout the apelinergic system. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Conformational transitions in peptides containing two putative alpha-helices of the prion protein.

    PubMed

    Zhang, H; Kaneko, K; Nguyen, J T; Livshits, T L; Baldwin, M A; Cohen, F E; James, T L; Prusiner, S B

    1995-07-21

    Prions are composed largely, if not entirely, of the scrapie isoform of the prion protein (PrPSc). Conversion of the cellular isoform (PrPC) to PrPSc is accompanied by a diminution in the alpha-helical content and an increase in the beta-sheet structure. To investigate the structural basis of this transition, peptide fragments corresponding to Syrian hamster PrP residues 90 to 145 and 109 to 141, which contain the most conserved residues of the prion protein and the first two putative alpha-helical regions in a PrPC model, were studied using infrared spectroscopy and circular dichroism. The peptides could be induced to form alpha-helical structures in aqueous solutions in the presence of organic solvents, such as trifluoroethanol and hexafluoroisopropanol, or detergents, such as sodium dodecyl sulfate and dodecyl phosphocholine. NaCl at physiological concentration or acetonitrile induced the peptides to acquire substantial beta-sheet. The intermolecular nature of the beta-sheet was evident in the formation of rod-shaped polymers as detected by electron microscopy. Resistance to hydrolysis by proteinase K and epitope mapping argue that the beta-sheet structures were formed by the interaction of residues lying between 109 and 141. A similar range of residues was shown by nuclear magnetic resonance spectroscopy to be capable of forming alpha-helices. The alpha-helical structures seem to require a hydrophobic support from either intermolecular interactions or the hydrophobic environment provided by micelles, in agreement with the predicted hydrophobic nature of the packing surface among the four putative helices of PrPC and the outer surfaces of the first two helices. Our results suggest that perturbation of the packing environment of the highly conserved residues is a possible mechanism for triggering the conversion of PrPC to PrPSc where alpha-helices appear to be converted into beta-sheets.

  13. Magic Angle Spinning NMR Reveals Sequence-Dependent Structural Plasticity, Dynamics, and the Spacer Peptide 1 Conformation in HIV-1 Capsid Protein Assemblies

    SciTech Connect

    Han, Yun; Hou, Guangjin; Suiter, Christopher L.; Ahn, Jinwoo; Byeon, In-Ja L.; Lipton, Andrew S.; Burton, Sarah D.; Hung, Ivan; Gorkov, Peter L.; Gan, Zhehong; Brey, William W.; Rice, David M.; Gronenborn, Angela M.; Polenova, Tatyana E.

    2013-11-27

    Maturation of HIV-1 virus into an infectious virion requires cleavage of the Gag polyprotein into its constituent domains and formation of a conical capsid core that encloses viral RNA and a small complement of proteins for replication. The final step of this process is the cleavage of the SP1 peptide from the CA-SP1 maturation intermediate, which triggers the condensation of the CA protein into a conical capsid. The mechanism of this step, including the conformation of the SP1 peptide in CA-SP1, is under intense debate. In this report, we examine the tubular assemblies of CA and the CA-SP1 maturation intermediate using Magic Angle Spinning NMR spectroscopy. At the magnetic fields of 19.9 T and above, tubular CA and CA-SP1 assemblies yield outstanding-quality 2D and 3D MAS NMR spectra, which are amenable to resonance assignments and detailed structural characterization. Dipolar- and scalar-based correlation experiments unequivocally indicate that SP1 peptide is in a random coil conformation and mobile in the assembled CA-SP1. Analysis of two sequence variants reveals that remarkably, the conformation of SP1 tail, of the functionally important CypA loop, and of the loop preceding helix 8 are sequence dependent and modulated by the residue variations at distal sites. These findings challenge the role of SP1 as a conformational switch in the maturation process and establish sequence-dependent conformational plasticity in CA.

  14. New mechanisms that regulate Saccharomyces cerevisiae short peptide transporter achieve balanced intracellular amino acid concentrations.

    PubMed

    Melnykov, Artem V

    2016-01-01

    The budding yeast Saccharomyces cerevisiae is able to take up large quantities of amino acids in the form of di- and tripeptides via a short peptide transporter, Ptr2p. It is known that PTR2 can be induced by certain peptides and amino acids, and the mechanisms governing this upregulation are understood at the molecular level. We describe two new opposing mechanisms of regulation that emphasize potential toxicity of amino acids: the first is upregulation of PTR2 in a population of cells, caused by amino acid secretion that accompanies peptide uptake; the second is loss of Ptr2p activity, due to transporter internalization following peptide uptake. Our findings emphasize the importance of proper amino acid balance in the cell and extend understanding of peptide import regulation in yeast.

  15. Desalted duck egg white peptides promote calcium uptake by counteracting the adverse effects of phytic acid.

    PubMed

    Hou, Tao; Liu, Weiwei; Shi, Wen; Ma, Zhili; He, Hui

    2017-03-15

    The structure of the desalted duck egg white peptides-calcium chelate was characterized by fluorescence spectroscopy, fourier transform infrared spectroscopy, and dynamic light scattering. Characterization results showed structural folding and aggregation of amino acids or oligopeptides during the chelation process. Desalted duck egg white peptides enhanced the calcium uptake in the presence of oxalate, phosphate and zinc ions in Caco-2 monolayers. Animal model indicated that desalted duck egg white peptides effectively enhanced the mineral absorption and counteracted the deleterious effects of phytic acid. These findings suggested that desalted duck egg white peptides might promote calcium uptake in three pathways: 1) desalted duck egg white peptides bind with calcium to form soluble chelate and avoid precipitate; 2) the chelate is absorbed as small peptides by enterocyte; and 3) desalted duck egg white peptides regulate the proliferation and differentiation of enterocytes through the interaction with transient receptor potential vanilloid 6 calcium channel.

  16. Three conformers of 2-furoic acid: structure changes induced with near-IR laser light.

    PubMed

    Halasa, Anna; Lapinski, Leszek; Reva, Igor; Rostkowska, Hanna; Fausto, Rui; Nowak, Maciej J

    2015-02-12

    Conformers of 2-furoic acid were studied using the matrix-isolation technique combined with narrow-band near-IR excitations with tunable laser light. Two conformers of the compound were trapped from the gas phase into low-temperature Ar or Ne matrixes with the population ratio of nearly 1:1. The two forms differ from each other by 180° rotation of the carboxylic group with respect to the furan ring. In both structures, the OH group adopts the cis orientation, with its H atom directed toward the C═O bond of the O═C-O-H group. Narrow-band near-IR excitations of the OH stretching overtone vibrations resulted in transformation of one of the initially observed conformers into a third conformational structure. This near-IR-induced isomerization concerned rotation of the OH group from the initial cis orientation to the trans conformation with the hydrogen atom directed toward the oxygen atom of the furan ring. In the photoproduced conformer, the hydrogen-bond-like O-H···O interaction (between O-H and the oxygen atom of the furan ring) is rather weak. Nevertheless, this interaction stabilized the structure so that it was present in the matrix for several hours after the near-IR-induced generation. The spontaneous conversion of the photogenerated, higher-energy form back into the more stable conformer with the carboxylic group in cis orientation was monitored for 2-furoic acid isolated in Ar and Ne matrixes. The speed of this process was found to be dependent on temperature and on the matrix material. The experimentally determined half-life times of this conformational conversion occurring in the dark are t1/2 = 1390 min (Ar, 5.5 K); t1/2 = 630 min (Ar, 15 K); t1/2 = 240 min (Ne, 5.5 K). The three conformers of 2-furoic acid observed in the present work were identified by comparison of their infrared spectra with the spectra theoretically calculated for the candidate structures.

  17. Conformational Dynamics of RNA-Peptide Binding: A Molecular Dynamics Simulation Study

    PubMed Central

    Mu, Yuguang; Stock, Gerhard

    2006-01-01

    Molecular dynamics simulations of the binding of the heterochiral tripeptide KkN to the transactivation responsive (TAR) RNA of HIV-1 is presented, using an all-atom force field with explicit water. To obtain starting structures for the TAR-KkN complex, semirigid docking calculations were performed that employ an NMR structure of free TAR RNA. The molecular dynamics simulations show that the starting structures in which KkN binds to the major groove of TAR (as it is the case for the Tat-TAR complex of HIV-1) are unstable. On the other hand, the minor-groove starting structures are found to lead to several binding modes, which are stabilized by a complex interplay of stacking, hydrogen bonding, and electrostatic interactions. Although the ligand does not occupy the binding position of Tat protein, it is shown to hinder the interhelical motion of free TAR RNA. The latter is presumably necessary to achieve the conformational change of TAR RNA to bind Tat protein. Considering the time evolution of the trajectories, the binding process is found to be ligand-induced and cooperative. That is, the conformational rearrangement only occurs in the presence of the ligand and the concerted motion of the ligand and a large part of the RNA binding site is necessary to achieve the final low-energy binding state. PMID:16239331

  18. Applications of peptide nucleic acids (PNAs) and locked nucleic acids (LNAs) in biosensor development.

    PubMed

    Briones, Carlos; Moreno, Miguel

    2012-04-01

    Nucleic acid biosensors have a growing number of applications in genetics and biomedicine. This contribution is a critical review of the current state of the art concerning the use of nucleic acid analogues, in particular peptide nucleic acids (PNA) and locked nucleic acids (LNA), for the development of high-performance affinity biosensors. Both PNA and LNA have outstanding affinity for natural nucleic acids, and the destabilizing effect of base mismatches in PNA- or LNA-containing heterodimers is much higher than in double-stranded DNA or RNA. Therefore, PNA- and LNA-based biosensors have unprecedented sensitivity and specificity, with special applicability in DNA genotyping. Herein, the most relevant PNA- and LNA-based biosensors are presented, and their advantages and their current limitations are discussed. Some of the reviewed technology, while promising, still needs to bridge the gap between experimental status and the harder reality of biotechnological or biomedical applications.

  19. Gamma Peptide Nucleic Acids: As Orthogonal Nucleic Acid Recognition Codes for Organizing Molecular Self-Assembly.

    PubMed

    Sacui, Iulia; Hsieh, Wei-Che; Manna, Arunava; Sahu, Bichismita; Ly, Danith H

    2015-07-08

    Nucleic acids are an attractive platform for organizing molecular self-assembly because of their specific nucleobase interactions and defined length scale. Routinely employed in the organization and assembly of materials in vitro, however, they have rarely been exploited in vivo, due to the concerns for enzymatic degradation and cross-hybridization with the host's genetic materials. Herein we report the development of a tight-binding, orthogonal, synthetically versatile, and informationally interfaced nucleic acid platform for programming molecular interactions, with implications for in vivo molecular assembly and computing. The system consists of three molecular entities: the right-handed and left-handed conformers and a nonhelical domain. The first two are orthogonal to each other in recognition, while the third is capable of binding to both, providing a means for interfacing the two conformers as well as the natural nucleic acid biopolymers (i.e., DNA and RNA). The three molecular entities are prepared from the same monomeric chemical scaffold, with the exception of the stereochemistry or lack thereof at the γ-backbone that determines if the corresponding oligo adopts a right-handed or left-handed helix, or a nonhelical motif. These conformers hybridize to each other with exquisite affinity, sequence selectivity, and level of orthogonality. Recognition modules as short as five nucleotides in length are capable of organizing molecular assembly.

  20. Comparative studies of adhesion peptides based on l- or d-amino acids.

    PubMed

    Nikitin, Sergey; Palmer, Daniel; Meldal, Morten; Diness, Frederik

    2016-10-01

    Detailed studies comparing solid-supported l- or d-amino acid adhesion peptides based on the sequence KLHRIRA were performed. Stability towards proteases and levels of cellular adhesion to the otherwise inert surface of PEGA resin were compared by using fluorescently labelled peptides. A clear difference in the peptide stability towards cleavage by subtilisin, trypsin, or papain was observed. However, all of the on-bead peptides provided an optimal surface for cell adhesion and proliferation. In long-term experiments, these properties were still found to be similar on the resins modified either with l- or with d-amino acids and unaffected by the nature of their fluorescence labelling at either terminus. These results support that the more accessible l-amino acids can be utilized for cell adhesion experiments and confirm the nonspecific interaction mechanism of cell binding to these peptides on the bead surface. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

  1. Alkylated dihydroxybenzoic acid as a MALDI matrix additive for hydrophobic peptide analysis.

    PubMed

    Fukuyama, Yuko; Tanimura, Ritsuko; Maeda, Kazuki; Watanabe, Makoto; Kawabata, Shin-Ichirou; Iwamoto, Shinichi; Izumi, Shunsuke; Tanaka, Koichi

    2012-05-01

    Hydrophobic peptides are generally difficult to detect using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) because the majority of MALDI matrixes are hydrophilic and therefore have a low affinity for hydrophobic peptides. Here, we report on a novel matrix additive, o-alkylated dihydroxybenzoic acid (ADHB), which is a 2,5-dihydroxybenzoic acid (DHB) derivative incorporating a hydrophobic alkyl chain on a hydroxyl group to improve its affinity for hydrophobic peptides, thereby improving MALDI-MS sensitivity. The addition of ADHB to the conventional matrix α-cyano-4-hydroxycinnamic acid (CHCA) improved the sensitivity of hydrophobic peptides 10- to 100-fold. The sequence coverage of phosphorylase b digest was increased using ADHB. MS imaging indicated that hydrophobic peptides were enriched in the rim of a matrix/analyte dried spot when ADHB was used. In conclusion, the addition of ADHB to the standard matrix led to improved sensitivity of hydrophobic peptides by MALDI-MS.

  2. The CLN025 Deca-Peptide Retains a β-Hairpin Conformation in Urea and GdmCl

    PubMed Central

    Hatfield, Marcus P. D.; Murphy, Richard F.

    2011-01-01

    The conformational stability of the β-hairpin miniprotein, CLN025, a variant of chignolin in which the N- and C-terminal Gly are replaced by Tyr, in various concentrations of GdmCl and urea was examined by Molecular Dynamics simulations and Electronic Circular Dichroism (ECD) spectropolarimetry. The peptide maintains its β-hairpin conformation in GdmCl and urea solutions. In GdmCl, Gly7 influences the turn to reduce the number of Asp3-Gly7 H-bonds and the Tyr1-Trp9 H-bond is lost. The structure of the peptide is less stable in 3 M GdmCl than in water or 6 M GdmCl, because the number of Asp3-Thr8 and Tyr1-Tyr10 H-bonds are reduced and the Tyr2 side-chain moves away from the Pro4 and Trp9 side-chains and towards the Tyr10 side-chain. This reduces the number of Tyr2-Pro4 CH-π interactions and Tyr2-Trp9 and Tyr1-Tyr10 aromatic-aromatic (Ar-Ar) interactions and increases the number of Tyr2-Tyr10 Ar-Ar interactions. In 6 M GdmCl at 300 and 333 K, the number of Tyr1-Tyr10 and Asp3-Thr8 H-bonds increases, but fewer structures have Tyr2-Pro4 CH-π and Tyr1-Tyr10 and Tyr2-Trp9 Ar-Ar interactions. In urea, Gly7 is in a mixture of β-turn and random meander structures and the number of Asp3-Thr6 and Tyr1-Tyr10 H-bonds are reduced as are the number of Tyr2-Pro4 CH-π interactions and Tyr1-Tyr10 and Tyr2-Trp9 Ar-Ar interactions. In 4 M urea, a shorter turn places Gly7 in to the β-sheet region and Tyr10 is pushed out into the solvent. In 8 M urea, the number of Asp3-Glu5 H-bonds is increased and the β-sheet is lost, but the electrostatic interaction between the charged termini is restored and a cation-π interaction between the indolyl ring of Trp9 and the positively charged N-terminus is formed. In 8 M urea at 333 K, the β-hairpin conformation is almost lost. The structure of CLN025 is stable, because the weakly polar interactions and H-bonds maintain the β-hairpin conformation in the various environments. CLN025 should not be considered a miniprotein, because it lacks a

  3. Predicting the conformations of peptides and proteins in early evolution. A review article submitted to Biology Direct

    PubMed Central

    Milner-White, E James; Russell, Michael J

    2008-01-01

    Considering that short, mainly heterochiral, polypeptides with a high glycine content are expected to have played a prominent role in evolution at the earliest stage of life before nucleic acids were available, we review recent knowledge about polypeptide three-dimensional structure to predict the types of conformations they would have adopted. The possible existence of such structures at this time leads to a consideration of their functional significance, and the consequences for the course of evolution. This article was reviewed by Bill Martin, Eugene Koonin and Nick Grishin. PMID:18226248

  4. Tri-peptide reference structures for the calculation of relative solvent accessible surface area in protein amino acid residues.

    PubMed

    Topham, Christopher M; Smith, Jeremy C

    2015-02-01

    Relative amino acid residue solvent accessibility values allow the quantitative comparison of atomic solvent-accessible surface areas in different residue types and physical environments in proteins and in protein structural alignments. Geometry-optimised tri-peptide structures in extended solvent-exposed reference conformations have been obtained for 43 amino acid residue types at a high level of quantum chemical theory. Significant increases in side-chain solvent accessibility, offset by reductions in main-chain atom solvent exposure, were observed for standard residue types in partially geometry-optimised structures when compared to non-minimised models built from identical sets of proper dihedral angles abstracted from the literature. Optimisation of proper dihedral angles led most notably to marked increases of up to 54% in proline main-chain atom solvent accessibility compared to literature values. Similar effects were observed for fully-optimised tri-peptides in implicit solvent. The relief of internal strain energy was associated with systematic variation in N, C(α) and C(β) atom solvent accessibility across all standard residue types. The results underline the importance of optimisation of 'hard' degrees of freedom (bond lengths and valence bond angles) and improper dihedral angle values from force field or other context-independent reference values, and impact on the use of standardised fixed internal co-ordinate geometry in sampling approaches to the determination of absolute values of protein amino acid residue solvent accessibility. Quantum chemical methods provide a useful and accurate alternative to molecular mechanics methods to perform energy minimisation of peptides containing non-standard (chemically modified) amino acid residues frequently present in experimental protein structure data sets, for which force field parameters may not be available. Reference tri-peptide atomic co-ordinate sets including hydrogen atoms are made freely available.

  5. Fecal Excretion of Orally Administered Collagen-Like Peptides in Rats: Contribution of the Triple-Helical Conformation to Their Stability.

    PubMed

    Koide, Takaki; Yamamoto, Naoyuki; Taira, Kazuma B; Yasui, Hiroyuki

    2016-01-01

    Orally ingested peptides are generally digested in the gastrointestinal (GI) tract and absorbed in the form of oligopeptides. We previously reported that intravenously administered collagen-like triple-helical peptides circulated in the bloodstream and were excreted in their intact forms in urine nearly quantitatively. In the present study, we investigated the fates of orally administered collagen-like peptides in rats. (Pro-Hyp-Gly)10 (Hyp: 4-hydroxyproline), which formed a stable triple-helical structure, was stable in the GI tract, and 72.3±13.0% of the peptide was excreted in the feces. Its recovery ratio was similar to that of all-D-(Pro-Pro-Gly)10 (75.1±15.7%), the indigestible control. In contrast, (Pro-Hyp-Gly)5 and (Pro-Pro-Gly)10, the random coil conformations of which were dominant at body temperature, were not detected in fecal samples, indicating that they were digested by proteases. The high stability of the triple-helical conformation in mammalian bodies suggests the potential use of collagen-like peptides as novel scaffolds of peptide drugs.

  6. Contemporary strategies for the stabilization of peptides in the alpha-helical conformation.

    PubMed

    Henchey, Laura K; Jochim, Andrea L; Arora, Paramjit S

    2008-12-01

    Herein we review contemporary synthetic and protein design strategies to stabilize the alpha-helical motif in short peptides and miniature proteins. Advances in organometallic catalyst design, specifically for the olefin metathesis reaction, enable the use of hydrocarbon bridges to either crosslink side chains of specific residues or mimic intramolecular hydrogen bonds with carbon-carbon bonds. The resulting hydrocarbon-stapled and hydrogen bond surrogate alpha-helices provide unique synthetic ligands for targeting biomolecules. In the protein design realm, several classes of miniature proteins that display stable helical domains have been engineered and manipulated with powerful in vitro selection technologies to yield libraries of sequences that retain their helical folds. Rational re-design of these scaffolds provide distinctive reagents for the modulation of protein-protein interactions.

  7. Effect of specific amino acid substitutions in the putative fusion peptide of structural glycoprotein E2 on Classical Swine Fever Virus replication

    SciTech Connect

    Fernández-Sainz, I.J.; Largo, E.; Gladue, D.P.; Fletcher, P.; O’Donnell, V.; Holinka, L.G.; Carey, L.B.; Lu, X.; Nieva, J.L.; Borca, M.V.

    2014-05-15

    E2, along with E{sup rns} and E1, is an envelope glycoprotein of Classical Swine Fever Virus (CSFV). E2 is involved in several virus functions: cell attachment, host range susceptibility and virulence in natural hosts. Here we evaluate the role of a specific E2 region, {sup 818}CPIGWTGVIEC{sup 828}, containing a putative fusion peptide (FP) sequence. Reverse genetics utilizing a full-length infectious clone of the highly virulent CSFV strain Brescia (BICv) was used to evaluate how individual amino acid substitutions within this region of E2 may affect replication of BICv. A synthetic peptide representing the complete E2 FP amino acid sequence adopted a β-type extended conformation in membrane mimetics, penetrated into model membranes, and perturbed lipid bilayer integrity in vitro. Similar peptides harboring amino acid substitutions adopted comparable conformations but exhibited different membrane activities. Therefore, a preliminary characterization of the putative FP {sup 818}CPIGWTGVIEC{sup 828} indicates a membrane fusion activity and a critical role in virus replication. - Highlights: • A putative fusion peptide (FP) region in CSFV E2 protein was shown to be critical for virus growth. • Synthetic FPs were shown to efficiently penetrate into lipid membranes using an in vitro model. • Individual residues in the FP affecting virus replication were identified by reverse genetics. • The same FP residues are also responsible for mediating membrane fusion.

  8. Minor influence of sialic acid on conformation of a membrane-bound oligosaccharide recognition site.

    PubMed

    Jones, D H; Barber, K R; Grant, C W

    1996-04-16

    Wideline 2H NMR spectroscopy was used to assess the conformational and orientational effects of N-acetylneuraminic acid (NeuAc) (sialic acid) as a component of a particular oligosaccharide chain at a bilayer membrane surface. For this purpose, three glycosphingolipids, sharing a neutral core tetrasaccharide and differing only in the number of sialic acid residues, were compared. The starting compound was GD1A, which has terminal sialic acid attached to the second and fourth sugars of its neutral tetrasaccharide core. GD1A was probe-labeled in a non-perturbing fashion on both of these sialic acid residues and on its single GalNAc residue by replacement of -COCH3 with -COCD3 giving [(d3NeuAc)2,d3-GalNAc]GA1a. This represents the most complex glycolipid to have been studied by 2H NMR spectroscopy at a bilayer membrane surface. The sialic acid residue on the fourth sugar from the membrane was subsequently removed to produce the glycolipid [d3NeuAc,d3GalNAc]GM1, deuterated at the two remaining amino sugars. The neutral glycolipid [d3GalNAc]asialo-GM1 was then generated by removal of the second sialic acid residue, leaving an uncharged species deuterated at one (internal) oligosaccharide chain site (GalNAc). The effect of sialic acid was futher examined by selective deuteration of GM1 and asialo-GM1 at C6 of the terminal Gal residue, giving [d2Gal]GM1 and [d2Gal]asialo-GM1. Spectra of the three glycosphingolipids were compared at 7.7 mol % in unsoncicated fluid bilayers of 1-palmitoyl-2-oleoylphosphatidylcholine containing 23 mol % cholesterol. For liposomes suspend in buffered salt solutions with 2 mM Ca2+, 2H NMR spectra demonstrated the presence of well defined average conformation for each oligosaccharide chain. This preferred average conformation persisted over a wide temperature range, consistent with there being a single major oligosaccharide conformer in each case. Spectral features arising from both deuterated amino sugar (GalNAc) of asialo-GM1 could be

  9. Synthesis and pharmacological evaluation of conformationally constrained glutamic acid higher homologues.

    PubMed

    Tamborini, Lucia; Cullia, Gregorio; Nielsen, Birgitte; De Micheli, Carlo; Conti, Paola; Pinto, Andrea

    2016-11-15

    Homologation of glutamic acid chain together with conformational constraint is a commonly used strategy to achieve selectivity towards different types of glutamate receptors. In the present work, starting from two potent and selective unnatural amino acids previously developed by us, we investigated the effects on the activity/selectivity profile produced by a further increase in the distance between the amino acidic moiety and the distal carboxylate group. Interestingly, the insertion of an aromatic ring as a spacer produced a low micromolar affinity NMDA ligand that might represent a lead for the development of a new class of NMDA antagonists. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Conformational mapping of the N-terminal peptide of HIV-1 gp41 in lipid detergent and aqueous environments using 13C-enhanced Fourier transform infrared spectroscopy

    PubMed Central

    Gordon, Larry M.; Mobley, Patrick W.; Lee, William; Eskandari, Sepehr; Kaznessis, Yiannis N.; Sherman, Mark A.; Waring, Alan J.

    2004-01-01

    The N-terminal domain of HIV-1 glycoprotein 41,000 (gp41) participates in viral fusion processes. Here, we use physical and computational methodologies to examine the secondary structure of a peptide based on the N terminus (FP; residues 1–23) in aqueous and detergent environments. 12C-Fourier transform infrared (FTIR) spectroscopy indicated greater α-helix for FP in lipid-detergent sodium dodecyl sulfate (SDS) and aqueous phosphate-buffered saline (PBS) than in only PBS. 12C-FTIR spectra also showed disordered FP conformations in these two environments, along with substantial β-structure for FP alone in PBS. In experiments that map conformations to specific residues, isotope-enhanced FTIR spectroscopy was performed using FP peptides labeled with 13C-carbonyl. 13C-FTIR results on FP in SDS at low peptide loading indicated α-helix (residues 5 to 16) and disordered conformations (residues 1–4). Because earlier 13C-FTIR analysis of FP in lipid bilayers demonstrated α-helix for residues 1–16 at low peptide loading, the FP structure in SDS micelles only approximates that found for FP with membranes. Molecular dynamics simulations of FP in an explicit SDS micelle indicate that the fraying of the first three to four residues may be due to the FP helix moving to one end of the micelle. In PBS alone, however, electron microscopy of FP showed large fibrils, while 13C-FTIR spectra demonstrated antiparallel β-sheet for FP (residues 1–12), analogous to that reported for amyloid peptides. Because FP and amyloid peptides each exhibit plaque formation, α-helix to β-sheet interconversion, and membrane fusion activity, amyloid and N-terminal gp41 peptides may belong to the same superfamily of proteins. PMID:15044732

  11. Analysis of local conformation of membrane-bound and polycrystalline peptides by two-dimensional slow-spinning rotor-synchronized MAS exchange spectroscopy.

    PubMed

    Gabrys, Charles M; Yang, Jun; Weliky, David P

    2003-05-01

    2D slow-spinning, rotor-synchronized MAS exchange spectroscopy (SSRS-MASE) was applied to study local secondary structure of three structurally different peptides, two of which were membrane-bound. Each peptide was (13)C carbonyl labeled at two adjacent residues in the peptide backbone. In general, this methodology is attractive for membrane-bound peptides because of its lenient spinning, decoupling, and RF homogeneity requirements. For a single set of raw SSRS-MASE data, two linearly independent methods exist for obtaining a 2D spectrum and each spectrum can be fit to obtain conformational constraints. An approach is described for combining the results of these two fits and this method is shown to work for spectra with both resolved and unresolved labeled site resonances. A spectrum is often fit well to a few different conformations which have somewhat different values of the fitting parameter chi(2). A simple statistical theory is developed which relates the deltachi(2) difference between a local minimum and the global minimum chi(2) to the likelihood that the local minimum conformation is the correct structure. Because uncertainty in the simulated data can also contribute to the overall fitting uncertainty, an empirical method is described for incorporating the simulation uncertainty into the deltachi(2) analysis. These data analysis methods were tested on polycrystalline Ala-Gly-Gly and then applied to the membrane-bound melittin and HIV-1 fusion peptides. Melittin gave a best-fit alpha helical structure at Ala-4 while the fusion peptide gave a good-fit beta strand structure at Phe-8. The melittin analysis is in agreement with the known overall structure of this peptide.

  12. Conformational Analysis of Retinoic Acids: Effects of Steric Interactions on Nonplanar Conjugated Polyenes

    PubMed Central

    Cox, Bryan D.; Muccio, Donald D.; Hamilton, Tracy P.

    2013-01-01

    Retinoic acids and other vitamin A analogs contain a trimethylcyclohexenyl ring in conjugation with a polyene chain joined at carbon-6 (C6) and carbon-7 (C7). A MP2-SCS/cc-pVDZ// B3LYP/6-31G(d) 2-D potential energy surface was computed for all-trans retinoic acid, which had 6 minima (3 enantiomeric pairs). The global minima were distorted s-gauche enantiomers (6–7 = 53°) with half-chair conformations of the ring. Distorted s-gauche enantiomers (6–7 = 55°) with inverted half-chair ring conformations were 1.7 kJ/mol above the global minima. The s-trans enantiomers (6–7 = 164°) were 11.3 kJ/mol above the global minima. Steric energies were computed by the method of Guo and Karplus to identify key structural elements in retinoic acids which determines their conformation. Small molecule crystal structures in the CCDC database with trimethylcyclohexenyl ring and exocyclic double bonds have ring-chain geometries near to one of the 6 energy minima of retinoic acids, except for retinaldehyde iminium cations. PMID:25798372

  13. Conformational Analysis of Retinoic Acids: Effects of Steric Interactions on Nonplanar Conjugated Polyenes.

    PubMed

    Cox, Bryan D; Muccio, Donald D; Hamilton, Tracy P

    2013-05-01

    Retinoic acids and other vitamin A analogs contain a trimethylcyclohexenyl ring in conjugation with a polyene chain joined at carbon-6 (C6) and carbon-7 (C7). A MP2-SCS/cc-pVDZ// B3LYP/6-31G(d) 2-D potential energy surface was computed for all-trans retinoic acid, which had 6 minima (3 enantiomeric pairs). The global minima were distorted s-gauche enantiomers (6-7 = 53°) with half-chair conformations of the ring. Distorted s-gauche enantiomers (6-7 = 55°) with inverted half-chair ring conformations were 1.7 kJ/mol above the global minima. The s-trans enantiomers (6-7 = 164°) were 11.3 kJ/mol above the global minima. Steric energies were computed by the method of Guo and Karplus to identify key structural elements in retinoic acids which determines their conformation. Small molecule crystal structures in the CCDC database with trimethylcyclohexenyl ring and exocyclic double bonds have ring-chain geometries near to one of the 6 energy minima of retinoic acids, except for retinaldehyde iminium cations.

  14. Conjugates of amino acids and peptides with 5-o-mycaminosyltylonolide and their interaction with the ribosomal exit tunnel.

    PubMed

    Shishkina, Anna; Makarov, Gennady; Tereshchenkov, Andrey; Korshunova, Galina; Sumbatyan, Nataliya; Golovin, Andrey; Svetlov, Maxim; Bogdanov, Alexey

    2013-11-20

    During protein synthesis the nascent polypeptide chain (NC) extends through the ribosomal exit tunnel (NPET). Also, the large group of macrolide antibiotics binds in the nascent peptide exit tunnel. In some cases interaction of NC with NPET leads to the ribosome stalling, a significant event in regulation of translation. In other cases NC-ribosome interactions lead to pauses in translation that play an important role in cotranslational folding of polypeptides emerging from the ribosome. The precise mechanism of NC recognition in NPET as well as factors that determine NC conformation in the ribosomal tunnel are unknown. A number of derivatives of the macrolide antibiotic 5-O-mycaminosyltylonolide (OMT) containing N-acylated amino acid or peptide residues were synthesized in order to study potential sites of NC-NPET interactions. The target compounds were prepared by conjugation of protected amino acids and peptides with the C23 hydroxyl group of the macrolide. These OMT derivatives showed high although varying abilities to inhibit the firefly luciferase synthesis in vitro. Three glycil-containing derivatives appeared to be strong inhibitors of translation, more potent than parental OMT. Molecular dynamics (MD) simulation of complexes of tylosin, OMT, and some of OMT derivatives with the large ribosomal subunit of E. coli illuminated a plausible reason for the high inhibitory activity of Boc-Gly-OMT. In addition, the MD study detected a new putative site of interaction of the nascent polypeptide chain with the NPET walls.

  15. Hydration studies of electrospray ions from amino acids and small peptides

    NASA Astrophysics Data System (ADS)

    Nguyen, Chuong (Steve)

    This project was undertaken to gain a better understanding of the hydration behaviors of gas phase ions from solutions containing amino acids and peptides. In order to characterize their hydration behavior, the molecules of interest in solutions were first converted into gas phase ions by electrospray ionization (ESI). The completely desolvated ions were then deliberately dispersed into an inert bath gas, usually nitrogen, containing accurately known concentrations of solvent vapor. The resulting mixtures of ions and bath gas were subsequently passed into a vacuum chamber by way of an adiabatic supersonic free jet expansion. The cooling during that expansion caused solvation of the ions, the extent of which was determined by a quadrupole mass analyzer. Mass analysis of the solute ions in the absence of vapor showed peaks with the mass to charge ratios corresponding to the desolvated ions. On the other hand, mass spectrometric analyses of ions in the presence of solvent vapor showed sequences of peaks corresponding to the solvated ions with varying numbers of water molecules. The extent of the ion solvation was controlled by varying the concentration of solvent vapor in the bath gas. Two different scales were proposed for the evaluation of the relative affinities of amino acids for water molecules. One was based primarily on the assumption that the affinities of amino acids for water molecules are directly proportional to their gas phase solvation rate constants ( k). An alternative approach produced an affinity scale based on the extent of ion hydration occurred during the free jet expansion. It was found that the addition of a polar solvent vapor to the bath gas at low concentrations substantially enhanced the production of the bare solute ions from the evaporating charged droplets. This remarkable result not only provided a means to increase the ion production and thus detection sensitivity of mass spectrometric analyses, but also yielded important information

  16. Low energy conformations and gas-phase acidity and basicity of pyrrolysine.

    PubMed

    Meng, Lingbiao; Wang, Zhuo; Zhang, Jicheng; Zhou, Minjie; Wu, Weidong

    2014-08-28

    The gas-phase conformational potential energy surfaces (PES) of the last, 22nd amino acid pyrrolysine and related derivatives (neutral, deprotonated, and protonated) were extensively searched for the first time. By considering all possible combinations of the single-bond rotational degrees of freedom with a semiempirical and ab initio combined computational approach, a large set of unique low-energy conformers was identified for each pyrrolysine species, and essential properties such as vibrational frequencies, dipole moments, rotational constants, and intramolecular hydrogen bonding configurations were presented and characterized. The conformational electronic energies and thermochemical properties of proton affinity/dissociation energy (PA/PDE) and gas-phase acidity/basicity (GA/GB) were determined by the density functional BHandHLYP, B3LYP, and M062X, and Møller-Plesset MP2 methods. The MP2 and DFT methods are found to predict disparate PES for neutral and protonated conformations and sufficiently different thermochemical data. The measurements of dipole moments and characteristic IR modes at low temperature as well as GA/GB are demonstrated to be feasible approaches to verify the theoretical predictions.

  17. Conformation of di-n-propylglycine residues (Dpg) in peptides: crystal structures of a type I' beta-turn forming tetrapeptide and an alpha-helical tetradecapeptide.

    PubMed

    Hegde, Raghurama P; Aravinda, Subrayashastry; Rai, Rajkishor; Kaul, Ramesh; Vijayalakshmi, Sarojini; Rao, R Balaji; Shamala, Narayanaswamy; Balaram, Padmanabhan

    2008-05-01

    The crystal structures of two oligopeptides containing di-n-propylglycine (Dpg) residues, Boc-Gly-Dpg-Gly-Leu-OMe (1) and Boc-Val-Ala-Leu-Dpg-Val-Ala-Leu-Val-Ala-Leu-Dpg-Val-Ala-Leu-OMe (2) are presented. Peptide 1 adopts a type I'beta-turn conformation with Dpg(2)-Gly(3) at the corner positions. The 14-residue peptide 2 crystallizes with two molecules in the asymmetric unit, both of which adopt alpha-helical conformations stabilized by 11 successive 5 --> 1 hydrogen bonds. In addition, a single 4 --> 1 hydrogen bond is also observed at the N-terminus. All five Dpg residues adopt backbone torsion angles (phi, psi) in the helical region of conformational space. Evaluation of the available structural data on Dpg peptides confirm the correlation between backbone bond angle N-C(alpha)-C' (tau) and the observed backbone phi,psi values. For tau > 106 degrees, helices are observed, while fully extended structures are characterized by tau < 106 degrees. The mean tau values for extended and folded conformations for the Dpg residue are 103.6 degrees +/- 1.7 degrees and 109.9 degrees +/- 2.6 degrees, respectively.

  18. Mutual Amino Acid Catalysis in Salt-Induced Peptide Formation Supports this Mechanism's Role in Prebiotic Peptide Evolution

    NASA Astrophysics Data System (ADS)

    Suwannachot, Yuttana; Rode, Bernd M.

    1999-10-01

    The presence of some amino acids and dipeptides under the conditions of the salt-induced peptide formation reaction (aqueous solution at 85 °C, Cu(II) and NaCl) has been found to catalyze the formation of homopeptides of other amino acids, which are otherwise produced only in traces or not at all by this reaction. The condensation of Val, Leu and Lys to form their homodipeptides can occur to a considerable extent due to catalytic effects of other amino acids and related compounds, among which glycine, histidine, diglycine and diketopiperazine exhibit the most remarkable activity. These findings also lead to a modification of the table of amino acid sequences preferentially formed by the salt-induced peptide formation (SIPF) reaction, previously used for a comparison with the sequence preferences in membrane proteins of primitive organisms

  19. Peptide-Conjugation Induced Conformational Changes in Human IgG1 Observed by Optimized Negative-Staining and Individual-Particle Electron Tomography

    PubMed Central

    Tong, Huimin; Zhang, Lei; Kaspar, Allan; Rames, Matthew J.; Huang, Liqing; Woodnutt, Gary; Ren, Gang

    2013-01-01

    Peptides show much promise as potent and selective drug candidates. Fusing peptides to a scaffold monoclonal antibody produces a conjugated antibody which has the advantages of peptide activity yet also has the pharmacokinetics determined by the scaffold antibody. However, the conjugated antibody often has poor binding affinity to antigens that may be related to unknown structural changes. The study of the conformational change is difficult by conventional techniques because structural fluctuation under equilibrium results in multiple structures co-existing. Here, we employed our two recently developed electron microscopy (EM) techniques: optimized negative-staining (OpNS) EM and individual-particle electron tomography (IPET). Two-dimensional (2D) image analyses and three-dimensional (3D) maps have shown that the domains of antibodies present an elongated peptide-conjugated conformational change, suggesting that our EM techniques may be novel tools to monitor the structural conformation changes in heterogeneous and dynamic macromolecules, such as drug delivery vehicles after pharmacological synthesis and development. PMID:23346347

  20. Amino acid polymorphisms in the fibronectin-binding repeats of fibronectin-binding protein A affect bond strength and fibronectin conformation.

    PubMed

    Casillas-Ituarte, Nadia N; Cruz, Carlos H B; Lins, Roberto D; DiBartola, Alex C; Howard, Jessica; Liang, Xiaowen; Höök, Magnus; Viana, Isabelle F T; Sierra-Hernández, M Roxana; Lower, Steven K

    2017-04-11

    The Staphylococcus aureus cell surface contains cell wall-anchored proteins such as fibronectin-binding protein A (FnBPA) that bind to host ligands (e.g. fibronectin; Fn) present in the extracellular matrix of tissue or coatings on cardiac implants. Recent clinical studies have found a correlation between cardiovascular infections caused by S. aureus and nonsynonymous single nucleotide polymorphisms (SNPs) in FnBPA. Atomic force microscopy (AFM), surface plasmon resonance (SPR), and molecular simulations were used to investigate interactions between Fn and each of eight, 20-mer peptide variants containing amino acids A, H, I, K, N, and Q at positions equivalent to 782 and/or 786 in Fn-binding repeat-9 (FnBR-9) of FnBPA. Experimentally measured bond lifetimes (1/koff ) and dissociation constants (Kd = koff / kon ), determined by mechanically dissociating the Fn-peptide complex at loading rates relevant to the cardiovascular system varied from the lowest-affinity H782A+K786A peptide (0.011 sec, 747 µM) to the highest-affinity H782Q+K786N peptide (0.192 sec, 15.7 µM). These AFM results tracked remarkably well to metadynamics simulations in which peptide detachment was defined solely by the free-energy landscape. Simulations and SPR experiments suggested that an Fn conformational change may enhance the stability of the binding complex for peptides with K786I or H782Q+K786I (Kd(app) = 0.2 to 0.5 µM. as determined by SPR) compared with the lowest-affinity double alanine peptide (Kd(app) = 3.8 µM). Together, these findings demonstrate that amino acid substitutions in FnBR-9 can significantly affect bond strength and influence the conformation of Fn upon binding. They provide a mechanistic explanation for the observation of nonsynonymous SNPs in fnbA) among clinical isolates of S. aureus that cause endovascular infections.

  1. Synthesis and pharmacological evaluation of novel conformationally constrained homologues of glutamic acid.

    PubMed

    Conti, Paola; Caligiuri, Antonio; Pinto, Andrea; Roda, Gabriella; Tamborini, Lucia; Nielsen, Birgitte; Madsen, Ulf; Frydenvang, Karla; Colombo, Alessio; De Micheli, Carlo

    2007-08-01

    Twelve novel conformationally constrained homologues of glutamic acid have been synthesized and pharmacologically characterized at ionotropic glutamate receptors (iGluRs). Synthesis of the target compounds involved 1,3-dipolar cycloaddition of nitrile oxides to suitable dipolarophiles. The structure to the compounds has been assigned by (1)H NMR and, in the case of derivatives (+/-)-4a, (+/-)-4b, (+/-)-5a, and (+/-)-5b, by means of an X-ray crystallographic analysis carried out on intermediate (+/-)-12a. The synthesized amino acids were found to be without affinity (K(i)/IC(50)>100microM) for iGluRs with the exception of compounds (+/-)-4b and (+/-)-5b, which showed a modest affinity for NMDA receptors (K(i)=34 and 13microM, respectively). The results indicate that the increased conformational constraints introduced by the cyclopropane ring and the spiro-attached proline ring are both detrimental to the pharmacological activity.

  2. New proctolin analogues modified by D-amino acids in the peptide chain and their high cardioexcitatory effect on Tenebrio molitor.

    PubMed

    Kuczer, M; Rosiński, G; Lisowski, M; Picur, B; Konopiñska, D

    1996-09-01

    The object of our studies was the synthesis and conformational and biological evaluation of the series of 14 analogues of the insect neuropeptide, proctolin. The analogues were obtained by replacement of the native L-amino acids by their D-isomers in one, two, and all positions. Biological effects of the peptides were examined by cardioexcitatory test on the heart of yellow mealworm, Tenebrio molitor, in vitro. In biotest performed on insects, D-Arg-D-Tyr-D-Leu-D-Pro-D-Thr, [D-Arg(N-G-nitro)1,D-Leu3]-, [D-Arg1,D-Leu3]-, [D-Tyr2,D-Thr5]- and [D-Arg1,D-Pro4]-proctolin exert high agonistic activity of proctolin on the heart of insects at 10(-11) - 10(-10) M concentrations. The proctolin analogue containing only D-amino acid residues in the peptide chain unexpectedly shows a much higher cardioexcitatory effect than the native peptide. Moreover, preliminary CD and NMR conformational studies show that proctolin analogues investigated here seem to prefer rather ordered structures, although their conformations differ in some cases.

  3. Assessing the Chemical Accuracy of Protein Structures via Peptide Acidity

    PubMed Central

    Anderson, Janet S.; Hernández, Griselda; LeMaster, David M.

    2012-01-01

    Although the protein native state is a Boltzmann conformational ensemble, practical applications often require a representative model from the most populated region of that distribution. The acidity of the backbone amides, as reflected in hydrogen exchange rates, is exquisitely sensitive to the surrounding charge and dielectric volume distribution. For each of four proteins, three independently determined X-ray structures of differing crystallographic resolution were used to predict exchange for the static solvent-exposed amide hydrogens. The average correlation coefficients range from 0.74 for ubiquitin to 0.93 for Pyrococcus furiosus rubredoxin, reflecting the larger range of experimental exchange rates exhibited by the latter protein. The exchange prediction errors modestly correlate with the crystallographic resolution. MODELLER 9v6-derived homology models at ~60% sequence identity (36% identity for chymotrypsin inhibitor CI2) yielded correlation coefficients that are ~0.1 smaller than for the cognate X-ray structures. The most recently deposited NOE-based ubiquitin structure and the original NMR structure of CI2 fail to provide statistically significant predictions of hydrogen exchange. However, the more recent RECOORD refinement study of CI2 yielded predictions comparable to the X-ray and homology model-based analyses. PMID:23182463

  4. Ab initio studies of aspartic acid conformers in gas phase and in solution

    NASA Astrophysics Data System (ADS)

    Chen, Mingliang; Lin, Zijing

    2007-10-01

    Systematic and extensive conformational searches of aspartic acid in gas phase and in solution have been performed. For the gaseous aspartic acid, a total of 1296 trial canonical structures and 216 trial zwitterionic structures were generated by allowing for all combinations of internal single-bond rotamers. All the trial structures were optimized at the B3LYP /6-311G* level and then subjected to further optimization at the B3LYP /6-311++G** level. A total of 139 canonical conformers were found, but no stable zwitterionic structure was found. The rotational constants, dipole moments, zero-point vibrational energies, harmonic frequencies, and vertical ionization energies of the canonical conformers were determined. Single-point energies were also calculated at the MP2/6-311++G** and CCSD /6-311++G** levels. The equilibrium distributions of the gaseous conformers at various temperatures were calculated. The proton affinity and gas phase basicity were calculated and the results are in excellent agreement with the experiments. The conformations in the solution were studied with different solvation models. The 216 trial zwitterionic structures were first optimized at the B3LYP /6-311G* level using the Onsager self-consistent reaction field model (SCRF) and then optimized at the B3LYP /6-311++G** level using the conductorlike polarized continuum model (CPCM) SCRF theory. A total of 22 zwitterions conformers were found. The gaseous canonical conformers were combined with the CPCM model and optimized at the B3LYP /6-311++G** level. The solvated zwitterionic and canonical structures were further examined by the discrete/SCRF model with one and two water molecules. The incremental solvation of the canonical and zwitterionic structures with up to six water molecules in gas phase was systematically examined. The studies show that combining aspartic acid with at least six water molecules in the gas phase or two water molecules and a SCRF solution model is required to provide

  5. Conformation of the umifenovir cation in the molecular and crystal structures of four carboxylic acid salts

    NASA Astrophysics Data System (ADS)

    Orola, Liana; Sarcevica, Inese; Kons, Artis; Actins, Andris; Veidis, Mikelis V.

    2014-01-01

    The umifenovir salts of maleic, salicylic, glutaric, and gentisic acid as well as the chloroform solvate of the salicylate were prepared. Single crystals of the five compounds were obtained and their molecular and crystal structures determined by X-ray diffraction. In each structure the conformation of phenyl ring with respect to the indole group of the umifenovir moiety is different. The water solubility and melting points of the studied umifenovir salts have been determined.

  6. Discrepancies between conformational distributions of a polyalanine peptide in solution obtained from molecular dynamics force fields and amide I' band profiles.

    PubMed

    Verbaro, Daniel; Ghosh, Indrajit; Nau, Werner M; Schweitzer-Stenner, Reinhard

    2010-12-30

    Structural preferences in the unfolded state of peptides determined by molecular dynamics still contradict experimental data. A remedy in this regard has been suggested by MD simulations with an optimized Amber force field ff03* ( Best, R. Hummer, G. J. Phys. Chem. B 2009 , 113 , 9004 - 9015 ). The simulations yielded a statistical coil distribution for alanine which is at variance with recent experimental results. To check the validity of this distribution, we investigated the peptide H-A(5)W-OH, which with the exception of the additional terminal tryptophan is analogous to the peptide used to optimize the force fields ff03*. Electronic circular dichroism, vibrational circular dichroism, and infrared spectroscopy as well as J-coupling constants obtained from NMR experiments were used to derive the peptide's conformational ensemble. Additionally, Förster resonance energy transfer between the terminal chromophores of the fluorescently labeled peptide analogue H-Dbo-A(5)W-OH was used to determine its average length, from which the end-to-end distance of the unlabeled peptide was estimated. Qualitatively, the experimental (3)J(H(N),C(α)), VCD, and ECD indicated a preference of alanine for polyproline II-like conformations. The experimental (3)J(H(N),C(α)) for A(5)W closely resembles the constants obtained for A(5). In order to quantitatively relate the conformational distribution of A(5) obtained with the optimized AMBER ff03* force field to experimental data, the former was used to derive a distribution function which expressed the conformational ensemble as a mixture of polyproline II, β-strand, helical, and turn conformations. This model was found to satisfactorily reproduce all experimental J-coupling constants. We employed the model to calculate the amide I' profiles of the IR and vibrational circular dichroism spectrum of A(5)W, as well as the distance between the two terminal peptide carbonyls. This led to an underestimated negative VCD couplet and an

  7. Phospholipid conjugate for intracellular delivery of peptide nucleic acids

    PubMed Central

    Shen, Gang; Fang, Huafeng; Song, Yinyin; Bielska, Agata A.; Wang, Zhenghui; Taylor, John-Stephen A.

    2009-01-01

    Peptide nucleic acids (PNAs) have a number of attractive features that have made them an ideal choice for antisense and antigene-based tools, probes and drugs, but their poor membrane permeability has limited their application as therapeutic or diagnostic agents. Herein we report a general method for the synthesis of phospholipid-PNAs (LP-PNAs), and compare the effect of non-cleavable lipids and bioreductively cleavable lipids (L and LSS) and phospholipid (LP) on the splice-correcting bioactivity of a PNA bearing the cell penetrating Arg9 group (PNA-R9). While the three constructs show similar and increasing bioactivity at 1–3 μM, the activity of LP-PNA-R9 continues to increase from 4–6 μM while the activity of L-PNA-R9 remains constant and LSS-PNA-R9 decreases rapidly in parallel with their relative cytotoxicity. The activity of both LP-PNA-R9 and L-PNA-R9 were found to dramatically increase with chloroquine, as expected for an endocytotic entry mechanism. Both constructs were also found to have CMC values of 1.0 and 4.5 μM in 150 mM NaCl, pH 7 water, suggesting that micelle formation may play a hitherto unrecognized role in modulating toxicity and/or facilitating endocytosis. PMID:19678628

  8. Conformational states of syntaxin-1 govern the necessity of N-peptide binding in exocytosis of PC12 cells and Caenorhabditis elegans

    PubMed Central

    Park, Seungmee; Bin, Na-Ryum; Michael Rajah, Maaran; Kim, Byungjin; Chou, Ting-Chieh; Kang, Soo-young Ann; Sugita, Kyoko; Parsaud, Leon; Smith, Matthew; Monnier, Philippe P.; Ikura, Mitsuhiko; Zhen, Mei; Sugita, Shuzo

    2016-01-01

    Syntaxin-1 is the central SNARE protein for neuronal exocytosis. It interacts with Munc18-1 through its cytoplasmic domains, including the N-terminal peptide (N-peptide). Here we examine the role of the N-peptide binding in two conformational states (“closed” vs. “open”) of syntaxin-1 using PC12 cells and Caenorhabditis elegans. We show that expression of “closed” syntaxin-1A carrying N-terminal single point mutations (D3R, L8A) that perturb interaction with the hydrophobic pocket of Munc18-1 rescues impaired secretion in syntaxin-1–depleted PC12 cells and the lethality and lethargy of unc-64 (C. elegans orthologue of syntaxin-1)-null mutants. Conversely, expression of the “open” syntaxin-1A harboring the same mutations fails to rescue the impairments. Biochemically, the L8A mutation alone slightly weakens the binding between “closed” syntaxin-1A and Munc18-1, whereas the same mutation in the “open” syntaxin-1A disrupts it. Our results reveal a striking interplay between the syntaxin-1 N-peptide and the conformational state of the protein. We propose that the N-peptide plays a critical role in intracellular trafficking of syntaxin-1, which is dependent on the conformational state of this protein. Surprisingly, however, the N-peptide binding mode seems dispensable for SNARE-mediated exocytosis per se, as long as the protein is trafficked to the plasma membrane. PMID:26700321

  9. Isolation and nature of intracellular alpha-aminoadipic acid-containing peptides from Paecilomyces persicinus P-10.

    PubMed Central

    Eriquez, L A; Pisano, M A

    1979-01-01

    Small intracellular peptides containing alpha-aminoadipic acid, cysteine, and a valine moiety were obtained from mycelia of Paecilomyces persicinus P-10 by ethanol or trichloroacetic acid extraction. After performic acid oxidation and ion-exchange chromatography, analysis of the peptide fractions by two-dimensional thin-layer electrophoresis and chromatography revealed the presence of three related peptides, as sulfonic acid derivatives, each containing alpha-aminoadipic acid. Each peptide was isolated in chromatographically pure form by semipreparative thin-layer electrophoresis and chromatography. The purified peptides were subjected to differential hydrolysis, dansylation, and combined dansylation-phenylisothiocyanate sequence analysis. Based on these studies, the structures of the isolated peptides were determined to be (i) glycl-delta-(alpha-aminoadipyl)-cysteinyl-beta-hydroxyvaline, (ii) glycyl-delta-(alpha-aminoadipyl)-cysteinylvaline, and (iii) delta-(alpha-aminoadipyl)-cysteinylvaline. The peptides isolated from Paecilomyces are similar to the alpha-aminoadipic acid-cysteine-valine moiety complex peptides isolated from Cephalosporium. PMID:574371

  10. Amino acid sequences of alpha-helical segments from S-carboxymethylkerateine-A. Tryptic and chymotryptic peptides from a type-II segment.

    PubMed Central

    Hogg, D M; Dowling, L M; Crewther, W G

    1978-01-01

    1. Amino acid-sequence studies were done on a peptide of mol.wt. approx. 12500 that was isolated from the highly helical fragments obtained by partial chymotryptic digestion of the low-sulphur proteins (S-carboxymethylkerateine-A) from wool. 2. The peptides obtained by tryptic and chymotryptic digestion of this large peptide were separated by ion-exchange chromatography on DEAE-cellulose at pH8.5 with an (NH4)(2)CO(3) concentration gradient and, where necessary, purified further by paper electrophoresis. 3. Determination of the sequences of many of these peptides showed that a high proportion of the cationic residues occurs in pairs. 4. Although two of the four S-carboxymethylcysteine residues are located in what appears to be a non-helical region near the N-terminus the other two S-carboxymethylcysteine residues occur in or near sequences suggesting a helical conformation. 5. Some peptides were obtained, in low yields, that appeared to be homologues of more major ones. These suggest either homologies in the helical portions of the low-sulphur proteins or the presence of closely related amino acid sequences in helical regions of completely different origins. 6. A partial sequence of the complete peptide is proposed. PMID:581263

  11. Antimicrobial peptides containing unnatural amino acid exhibit potent bactericidal activity against ESKAPE pathogens.

    PubMed

    Hicks, R P; Abercrombie, J J; Wong, R K; Leung, K P

    2013-01-01

    A series of 36 synthetic antimicrobial peptides containing unnatural amino acids were screened to determine their effectiveness to treat Enterococcus faecium, Staphylococcus aureus, Klebsiella pnemoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species (ESKAPE) pathogens, which are known to commonly infect chronic wounds. The primary amino acid sequences of these peptides incorporate either three or six dipeptide units consisting of the unnatural amino acids Tetrahydroisoquinolinecarboxylic acid (Tic) and Octahydroindolecarboxylic acid (Oic). The Tic-Oic dipeptide units are separated by SPACER amino acids with specific physicochemical properties that control how these peptides interact with bacterial cell membranes of different chemical compositions. These peptides exhibited minimum inhibitory concentrations (MIC) against these pathogens in the range from >100 to 6.25 μg/mL. The observed diversity of MIC values for these peptides against the various bacterial strains are consistent with our hypothesis that the complementarity of the physicochemical properties of the peptide and the lipid of the bacteria's cell membrane determines the resulting antibacterial activity of the peptide. Published by Elsevier Ltd.

  12. Conformational dynamics of nucleic acid molecules studied by PELDOR spectroscopy with rigid spin labels.

    PubMed

    Prisner, T F; Marko, A; Sigurdsson, S Th

    2015-03-01

    Nucleic acid molecules can adopt a variety of structures and exhibit a large degree of conformational flexibility to fulfill their various functions in cells. Here we describe the use of Pulsed Electron-Electron Double Resonance (PELDOR or DEER) to investigate nucleic acid molecules where two cytosine analogs have been incorporated as spin probes. Because these new types of spin labels are rigid and incorporated into double stranded DNA and RNA molecules, there is no additional flexibility of the spin label itself present. Therefore the magnetic dipole-dipole interaction between both spin labels encodes for the distance as well as for the mutual orientation between the spin labels. All of this information can be extracted by multi-frequency/multi-field PELDOR experiments, which gives very precise and valuable information about the structure and conformational flexibility of the nucleic acid molecules. We describe in detail our procedure to obtain the conformational ensembles and show the accuracy and limitations with test examples and application to double-stranded DNA.

  13. Conformational dynamics of nucleic acid molecules studied by PELDOR spectroscopy with rigid spin labels

    NASA Astrophysics Data System (ADS)

    Prisner, T. F.; Marko, A.; Sigurdsson, S. Th.

    2015-03-01

    Nucleic acid molecules can adopt a variety of structures and exhibit a large degree of conformational flexibility to fulfill their various functions in cells. Here we describe the use of Pulsed Electron-Electron Double Resonance (PELDOR or DEER) to investigate nucleic acid molecules where two cytosine analogs have been incorporated as spin probes. Because these new types of spin labels are rigid and incorporated into double stranded DNA and RNA molecules, there is no additional flexibility of the spin label itself present. Therefore the magnetic dipole-dipole interaction between both spin labels encodes for the distance as well as for the mutual orientation between the spin labels. All of this information can be extracted by multi-frequency/multi-field PELDOR experiments, which gives very precise and valuable information about the structure and conformational flexibility of the nucleic acid molecules. We describe in detail our procedure to obtain the conformational ensembles and show the accuracy and limitations with test examples and application to double-stranded DNA.

  14. Facile plasma-enhanced deposition of ultrathin crosslinked amino acid films for conformal biometallization.

    PubMed

    Anderson, Kyle D; Slocik, Joseph M; McConney, Michael E; Enlow, Jesse O; Jakubiak, Rachel; Bunning, Timothy J; Naik, Rajesh R; Tsukruk, Vladimir V

    2009-03-01

    A novel method for the facile fabrication of conformal, ultrathin, and uniform synthetic amino acid coatings on a variety of practical surfaces by plasma-enhanced chemical vapor deposition is introduced. Tyrosine, which is utilized as an agent to reduce gold nanoparticles from solution, is sublimed into the plasma field and directly deposited on a variety of substrates to form a homogeneous, conformal, and robust polyamino acid coating in a one-step, solvent-free process. This approach is applicable to many practical surfaces and allows surface-induced biometallization while avoiding multiple wet-chemistry treatments that can damage many soft materials. Moreover, by placing a mask over the substrate during deposition, the tyrosine coating can be micropatterned. Upon its exposure to a solution of gold chloride, a network of gold nanoparticles forms on the surface, replicating the initial micropattern. This method of templated biometallization is adaptable to a variety of practical inorganic and organic substrates, such as silicon, glass, nitrocellulose, polystyrene, polydimethylsiloxane, polytetrafluoroethylene, polyethylene, and woven silk fibers. No special pretreatment is necessary, and the technique results in a rapid, conformal amino acid coating that can be utilized for further biometallization.

  15. Thermodynamic principles for the engineering of pH-driven conformational switches and acid insensitive proteins

    PubMed Central

    Bell-Upp, Peregrine; Robinson, Aaron C.; Whitten, Steven; Wheeler, Erika L.; Lin, Janine; Stites, Wesley E.; García-Moreno E, Bertrand

    2012-01-01

    The general thermodynamic principles behind pH driven conformational transitions of biological macromolecules are well understood. What is less obvious is how they can be used to engineer pH switches in proteins. The acid unfolding of staphylococcal nuclease (SNase) was used to illustrate different factors that can affect pH-driven conformational transitions. Acid unfolding is a structural transition driven by preferential H+ binding to the acid unfolded state (U) over the native (N) state of a protein. It is the result of carboxylic groups that titrate with more normal pKa values in the U state than in the N state. Acid unfolding profiles of proteins reflect a balance between electrostatic and non-electrostatic contributions to stability. Several strategies were used in attempts to turn SNase into an acid insensitive protein: (1) enhancing global stability of the protein with mutagenesis or with osmolytes, (2) use of high salt concentrations to screen Coulomb interactions, (3) stabilizing the N state through specific anion effects, (4) removing Asp or Glu residues that titrate with depressed pKa values in the N state, and (5) removing basic residues that might have strong repulsive interactions in the N state at low pH. The only effective way to engineer acid resistance in SNase is not through modulation of pKa values of Asp/Glu but by enhancing the global stability of the protein. Modulation of pH-driven conformational transitions by selective manipulation of the electrostatic component of the switch is an extremely difficult undertaking. PMID:21802194

  16. Thermodynamic principles for the engineering of pH-driven conformational switches and acid insensitive proteins.

    PubMed

    Bell-Upp, Peregrine; Robinson, Aaron C; Whitten, Steven T; Wheeler, Erika L; Lin, Janine; Stites, Wesley E; E, Bertrand García-Moreno

    2011-11-01

    The general thermodynamic principles behind pH driven conformational transitions of biological macromolecules are well understood. What is less obvious is how they can be used to engineer pH switches in proteins. The acid unfolding of staphylococcal nuclease (SNase) was used to illustrate different factors that can affect pH-driven conformational transitions. Acid unfolding is a structural transition driven by preferential H(+) binding to the acid unfolded state (U) over the native (N) state of a protein. It is the result of carboxylic groups that titrate with more normal pK(a) values in the U state than in the N state. Acid unfolding profiles of proteins reflect a balance between electrostatic and non-electrostatic contributions to stability. Several strategies were used in attempts to turn SNase into an acid insensitive protein: (1) enhancing global stability of the protein with mutagenesis or with osmolytes, (2) use of high salt concentrations to screen Coulomb interactions, (3) stabilizing the N state through specific anion effects, (4) removing Asp or Glu residues that titrate with depressed pK(a) values in the N state, and (5) removing basic residues that might have strong repulsive interactions in the N state at low pH. The only effective way to engineer acid resistance in SNase is not through modulation of pK(a) values of Asp/Glu but by enhancing the global stability of the protein. Modulation of pH-driven conformational transitions by selective manipulation of the electrostatic component of the switch is an extremely difficult undertaking. Copyright © 2011 Elsevier B.V. All rights reserved.

  17. Helix 69 of E. coli 23S ribosomal RNA as a peptide nucleic acid target.

    PubMed

    Kulik, Marta; Markowska-Zagrajek, Agnieszka; Wojciechowska, Monika; Grzela, Renata; Wituła, Tomasz; Trylska, Joanna

    2017-04-07

    A fragment of 23S ribosomal RNA (nucleotides 1906-1924 in E. coli), termed Helix 69, forms a hairpin that is essential for ribosome function. Helix 69 forms a conformationally flexible inter-subunit connection with helix 44 of 16S ribosomal RNA, and the nucleotide A1913 of Helix 69 influences decoding accuracy. Nucleotides U1911 and U1917 are post-transcriptionally modified with pseudouridines () and U1915 with 3-methyl-. We investigated Helix 69 as a target for a complementary synthetic oligonucleotide - peptide nucleic acid (PNA). We determined thermodynamic properties of Helix 69 and its complexes with PNA. We also verified the performance of PNA targeted at Helix 69 in inhibiting translation in cell-free extracts and growth of E. coli cells. First, we examined the interactions of a PNA oligomer complementary to the G1907-A1919 fragment of Helix 69 with the sequences corresponding to human and bacterial species (with or without pseudouridine modifications). PNA invades the Helix 69 hairpin creating stable complexes and PNA binding to the pseudouridylated bacterial sequence is stronger than to Helix 69 without any modifications. Second, we confirmed the binding of PNA to 23S rRNA and 70S ribosomes. Third, we verified the efficiency of translation inhibition of these PNA oligomers in the cell-free translation/transcription E. coli system, which turned out to be in a similar range as tetracycline. Next, we confirmed that PNA conjugated to the (KFF)3K transporter peptide inhibited E. coli growth in micromolar concentrations. Overall, targeting Helix 69 with PNA or other sequence-specific oligomers could be a promising way to inhibit bacterial translation.

  18. Evidence of PPII-like helical conformation and glass transition in a self-assembled solid-state polypeptide-surfactant complex: poly(L-histidine)/docylbenzenesulfonic acid.

    PubMed

    Ramani, Ramasubbu; Hanski, Sirkku; Laiho, Ari; Tuma, Roman; Kilpeläinen, Simo; Tuomisto, Filip; Ruokolainen, Janne; Ikkala, Olli

    2008-05-01

    We present lamellar self-assembly of cationic poly(L-histidine) (PLH) stoichiometrically complexed with an anionic surfactant, dodecyl benzenesulfonic acid (DBSA), which allows a stabilized conformation reminiscent of polyproline type II (PPII) left-handed helices. Such a conformation has no intrapeptide hydrogen bonds, and it has previously been found to be one source of flexibility, e.g., in collagen and elastin, as well as an intermediate in silk processing. PLH(DBSA)1.0 complexes were characterized by Fourier transform infrared spectroscopy (FTIR), circular dichroism (CD), small-angle X-ray scattering (SAXS), transmission electron microscopy (TEM), and differential scanning calorimetry (DSC). The PPII-like conformation in PLH(DBSA)1.0 is revealed by characteristic CD and FTIR spectra, where the latter indicates absence of intrachain peptide hydrogen bonds. In addition, a glass transition was directly verified by DSC at ca. 135 degrees C for PLH(DBSA)1.0 and indirectly by SAXS and TEM in comparison to pure PLH at 165 degrees C, thus indicating plasticization. Glass transitions have not been observed before in polypeptide-surfactant complexes. The present results show that surfactant binding can be a simple scheme to provide steric crowding to stabilize PPII conformation to tune the polypeptide properties, plasticization and flexibility.

  19. Supramolecular control of self-assembling terthiophene-peptide conjugates through the amino acid side chain

    SciTech Connect

    Lehrman, Jessica A.; Cui, Honggang; Tsai, Wei-Wen; Moyer, Tyson J.; Stupp, Samuel I.

    2013-07-30

    The self-assembly of oligothiophene–peptide conjugates can be directed through the systematic variation of the peptide sequence into different nanostructures, including flat spicules, nanotubes, spiral sheets, and giant, flat sheets. Furthermore, the assembly of these molecules is not controlled by steric interactions between the amino acid side chains.

  20. A peptide & peptide nucleic acid synthesis technology for transporter molecules and theranostics--the SPPS.

    PubMed

    Pipkorn, Ruediger; Braun, Klaus; Wiessler, Manfred; Waldeck, Waldemar; Schrenk, Hans-Hermann; Koch, Mario; Semmler, Wolfhard; Komljenovic, Dorde

    2014-01-01

    Advances in imaging diagnostics using magnetic resonance tomography (MRT), positron emission tomography (PET) and fluorescence imaging including near infrared (NIR) imaging methods are facilitated by constant improvement of the concepts of peptide synthesis. Feasible patient-specific theranostic platforms in the personalized medicine are particularly dependent on efficient and clinically applicable peptide constructs. The role of peptides in the interrelations between the structure and function of proteins is widely investigated, especially by using computer-assisted methods. Nowadays the solid phase synthesis (SPPS) chemistry emerges as a key technology and is considered as a promising methodology to design peptides for the investigation of molecular pharmacological processes at the transcriptional level. SPPS syntheses could be carried out in core facilities producing peptides for large-scale scientific implementations as presented here.

  1. A Peptide & Peptide Nucleic Acid Synthesis Technology for Transporter Molecules and Theranostics - The SPPS

    PubMed Central

    Pipkorn, Ruediger; Braun, Klaus; Wiessler, Manfred; Waldeck, Waldemar; Schrenk, Hans-Hermann; Koch, Mario; Semmler, Wolfhard; Komljenovic, Dorde

    2014-01-01

    Advances in imaging diagnostics using magnetic resonance tomography (MRT), positron emission tomography (PET) and fluorescence imaging including near infrared (NIR) imaging methods are facilitated by constant improvement of the concepts of peptide synthesis. Feasible patient-specific theranostic platforms in the personalized medicine are particularly dependent on efficient and clinically applicable peptide constructs. The role of peptides in the interrelations between the structure and function of proteins is widely investigated, especially by using computer-assisted methods. Nowadays the solid phase synthesis (SPPS) chemistry emerges as a key technology and is considered as a promising methodology to design peptides for the investigation of molecular pharmacological processes at the transcriptional level. SPPS syntheses could be carried out in core facilities producing peptides for large-scale scientific implementations as presented here. PMID:24843319

  2. Amide hydrogen exchange rates of peptides in H2O solution by 1H nuclear magnetic resonance transfer of solvent saturation method. Conformations of oxytocin and lysine vasopressin in aqueous solution.

    PubMed Central

    Krishna, N R; Huang, D H; Glickson, J D; Rowan, R; Walter, R

    1979-01-01

    The NH exchange rates in aqueous media of oxytocin and 8-lysine vasopressin (LVP) have been measured by using transfer of solvent saturation method. The data are consistent with a "highly motile" dynamic equilibrium between folded and highly solvated conformations. The highly-motility limit applies to the exchange of NH hydrogens of oxytocin and LVP. Folded structures are more prevalent in oxytocin than in LVP. Partial shielding is indicated for peptide hydrogens of Asn5 and perhaps also Cys6 of oxytocin and for Cys6 of LVP. It is tentatively proposed that the folded conformation of oxytocin in aqueous media may contain a parallel beta-structure in the tocinamide ring consisting of two hydrogen bonds: one between the Tyr2 C = O and Asn5 peptide NH as originally proposed for the preferred conformation of oxytocin in dimethyl sulfoxide (D. W. Urry and R. Walter), and the second between he Cys1 C = O and the Cys6 NH. In LVP the hydrogen bond between the Tyr2 C = O and Asn5 peptide NH appears to be absent. The acylic tripeptide sequences (-Pro-X-Gly-NH2) of both hormones appear to be predominantly solvated. The second-order rate constants for acid catalyzed exchange of the primary amide hydrogens of Gln4, Asn5, and Gly9 of oxytocin are consistently greater for the trans NH than for the corresponding cis NH. This observation can be rationalized in terms of mechanisms involving protonation of either the amide oxygen, or the amide nitrogen, but with limited rotation about the C - N bond. PMID:262422

  3. Structure and conformational variability of the mycobacterium tuberculosis fatty acid synthase multienzyme complex.

    PubMed

    Ciccarelli, Luciano; Connell, Sean R; Enderle, Mathias; Mills, Deryck J; Vonck, Janet; Grininger, Martin

    2013-07-02

    Antibiotic therapy in response to Mycobacterium tuberculosis infections targets de novo fatty acid biosynthesis, which is orchestrated by a 1.9 MDa type I fatty acid synthase (FAS). Here, we characterize M. tuberculosis FAS by single-particle cryo-electron microscopy and interpret the data by docking the molecular models of yeast and Mycobacterium smegmatis FAS. Our analysis reveals a porous barrel-like structure of considerable conformational variability that is illustrated by the identification of several conformational states with altered topology in the multienzymatic assembly. This demonstrates that the barrel-like structure of M. tuberculosis FAS is not just a static scaffold for the catalytic domains, but may play an active role in coordinating fatty acid synthesis. The conception of M. tuberculosis FAS as a highly dynamic assembly of domains revises the view on bacterial type I fatty acid synthesis and might inspire new strategies for inhibition of de novo fatty acid synthesis in M. tuberculosis.

  4. Carbonyl-carbonyl interactions stabilize the partially allowed Ramachandran conformations of asparagine and aspartic acid.

    PubMed

    Deane, C M; Allen, F H; Taylor, R; Blundell, T L

    1999-12-01

    Asparagine and aspartate are known to adopt conformations in the left-handed alpha-helical region and other partially allowed regions of the Ramachandran plot more readily than any other non-glycyl amino acids. The reason for this preference has not been established. An examination of the local environments of asparagine and aspartic acid in protein structures with a resolution better than 1.5 A revealed that their side-chain carbonyls are frequently within 4 A of their own backbone carbonyl or the backbone carbonyl of the previous residue. Calculations using protein structures with a resolution better than 1.8 A reveal that this close contact occurs in more than 80% of cases. This carbonyl-carbonyl interaction offers an energetic sabilization for the partially allowed conformations of asparagine and aspartic acid with respect to all other non-glycyl amino acids. The non-covalent attractive interactions between the dipoles of two carbonyls has recently been calculated to have an energy comparable to that of a hydrogen bond. The preponderance of asparagine in the left-handed alpha-helical region, and in general of aspartic acid and asparagine in the partially allowed regions of the Ramachandran plot, may be a consequence of this carbonyl-carbonyl stacking interaction.

  5. Polymorphisms at Amino Acid Residues 141 and 154 Influence Conformational Variation in Ovine PrP

    PubMed Central

    Yang, Sujeong; Thackray, Alana M.; Hopkins, Lee; Monie, Tom P.; Burke, David F.; Bujdoso, Raymond

    2014-01-01

    Polymorphisms in ovine PrP at amino acid residues 141 and 154 are associated with susceptibility to ovine prion disease: Leu141Arg154 with classical scrapie and Phe141Arg154 and Leu141His154 with atypical scrapie. Classical scrapie is naturally transmissible between sheep, whereas this may not be the case with atypical scrapie. Critical amino acid residues will determine the range or stability of structural changes within the ovine prion protein or its functional interaction with potential cofactors, during conversion of PrPC to PrPSc in these different forms of scrapie disease. Here we computationally identified that regions of ovine PrP, including those near amino acid residues 141 and 154, displayed more conservation than expected based on local structural environment. Molecular dynamics simulations showed these conserved regions of ovine PrP displayed genotypic differences in conformational repertoire and amino acid side-chain interactions. Significantly, Leu141Arg154 PrP adopted an extended beta sheet arrangement in the N-terminal palindromic region more frequently than the Phe141Arg154 and Leu141His154 variants. We supported these computational observations experimentally using circular dichroism spectroscopy and immunobiochemical studies on ovine recombinant PrP. Collectively, our observations show amino acid residues 141 and 154 influence secondary structure and conformational change in ovine PrP that may correlate with different forms of scrapie. PMID:25126555

  6. Single Conformation Spectroscopy of Suberoylanilide Hydroxamic Acid: a Molecule Bites its Tail

    NASA Astrophysics Data System (ADS)

    Zhang, Di; Dean, Jacob; Zwier, Timothy S.

    2012-06-01

    Suberoylanilide hydroxamic acid (C_6H_5NHCO(CH_2)_6CONHOH, SAHA) is a histone deacetylase inhibitor approved by the FDA for the treatment of cutaneous T-cell lymphoma. With one hydrogen bonding group adjacent to ring and the other at the end of a long C_6 hydrocarbon tail, SAHA possesses an interesting potential energy landscape to be probed by single-conformation methods. A large number of extended structures favored by entropy are offset by a few structures