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Peptide Backbone (peptide + backbone)
Selected AbstractsEvaluation of carrier ampholyte-based capillary electrophoresis for separation of peptides and peptide mimetics,ELECTROPHORESIS, Issue 18 2008an Koval Abstract Carrier ampholyte-based capillary electrophoresis (CABCE) has recently been introduced as an alternative to CE (CZE) in the classical buffers. In this study, isoelectric BGEs were obtained by fractionation of Servalyt pH 4,9 carrier ampholytes to cuts of typical width of 0.2 pH unit. CABCE feasibility was examined on a series of insect oostatic peptides, i.e. proline-rich di- to decapeptides, and phosphinic pseudopeptides , tetrapeptide mimetics synthesized as a mixture of four diastereomers having the ,P(O)(OH),CH2, moiety embedded into the peptide backbone. With identical selectivity, the separation efficiency of CABCE proved to be as good as classical CE for the insect oostatic peptides and better for diastereomers of the phosphinic pseudopeptides. In addition, despite the numerous species present in the narrow pH cuts of carrier ampholytes, CABCE seems to be free of system zones that could hamper the analysis. Peak symmetry was good for moderately to low mobile peptides, whereas some peak distortion due to electromigration dispersion, was observed for short peptides of rather high mobility. [source] The ester-bonded palmitoyl side chains of Pam3CysSerLys4 lipopeptide account for its powerful adjuvanticity to HLA class,I-restricted CD8+ T,lymphocytesEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 7 2003Anca Reschner Abstract Molecularly defined adjuvants are urgently required to implement immunization protocols by which CD8+ T,cells induction is envisaged. We show here that the synthetic lipopeptide Pam3CysSerLys4 (P3CSK4) strongly enhances the expansion of antigen-specific IFN-,+CD8+ cells in vitro. These effects critically depend on the presence of two ester-bonded palmitoylated side chains. In fact, T,cell expansion is impaired in the presence of derivatives bearing two non-palmitoylated fatty acid chains, while derivatives with only one amide-bonded palmitoylated residue are completely inactive and behave like the non-lipidated peptide backbone. P3CSK4 is not mitogenic for T,lymphocytes and can modulte DC immune biological properties. Indeed, doses as low as 100,ng/ml increase CD86, CD83 and CD40 surface expression on DC, fail to induce CCR7, and trigger a defined pattern of soluble factors associated to immune effector functions. In particular, substantial amounts of TNF-,, IL-6, CCL2 and CXCL10, in the absence of IFN-,, IFN-,, IL-15, IL-12p70 and CX3CL1, can be measured. Accordingly, antigen-specific CD8+ T,cells expanded in vitro express CCR2 and CXCR3 chemokine receptors. Altogether our data suggest that human DC are able to respond to chemically different synthetic lipopeptide analogs and that optimal adjuvanticity to CD8+ T,cell induction is achieved by the palmitoylated structures. [source] On the molecular basis of the recognition of angiotensin II (AII)FEBS JOURNAL, Issue 5 2003NMR structure of AII in solution compared with the X-ray structure of AII bound to the mAb Fab13 The high-resolution 3D structure of the octapeptide hormone angiotensin II (AII) in aqueous solution has been obtained by simulated annealing calculations, using high-resolution NMR-derived restraints. After final refinement in explicit water, a family of 13 structures was obtained with a backbone RMSD of 0.73 ± 0.23 Å. AII adopts a fairly compact folded structure, with its C-terminus and N-terminus approaching to within ,,7.2 Å of each other. The side chains of Arg2, Tyr4, Ile5 and His6 are oriented on one side of a plane defined by the peptide backbone, and the Val3 and Pro7 are pointing in opposite directions. The stabilization of the folded conformation can be explained by the stacking of the Val3 side chain with the Pro7 ring and by a hydrophobic cluster formed by the Tyr4, Ile5 and His6 side chains. Comparison between the NMR-derived structure of AII in aqueous solution and the refined crystal structure of the complex of AII with a high-affinity mAb (Fab131) [Garcia, K.C., Ronco, P.M., Verroust, P.J., Brunger, A.T., Amzel, L.M. (1992) Science257, 502,507] provides important quantitative information on two common structural features: (a) a U-shaped structure of the Tyr4-Ile5-His6-Pro7 sequence, which is the most immunogenic epitope of the peptide, with the Asp1 side chain oriented towards the interior of the turn approaching the C-terminus; (b) an Asx-turn-like motif with the side chain aspartate carboxyl group hydrogen-bonded to the main chain NH group of Arg2. It can be concluded that small rearrangements of the epitope 4,7 in the solution structure of AII are required by a mean value of 0.76 ± 0.03 Å for structure alignment and ,,1.27 ± 0.02 Å for sequence alignment with the X-ray structure of AII bound to the mAb Fab131. These data are interpreted in terms of a biological ,nucleus' conformation of the hormone in solution, which requires a limited number of structural rearrangements for receptor,antigen recognition and binding. [source] Evidence of a functional requirement for a carbamoylated lysine residue in MurD, MurE and MurF synthetases as established by chemical rescue experimentsFEBS JOURNAL, Issue 22 2001Sébastien Dementin Enzymes MurD, MurE, MurF, folylpolyglutamate synthetase and cyanophycin synthetase, which belong to the Mur synthetase superfamily, possess an invariant lysine residue (K198 in the Escherichia coli MurD numbering). Crystallographic analysis of MurD and MurE has recently shown that this residue is present as a carbamate derivative, a modification presumably essential for Mg2+ binding and acyl phosphate formation. In the present work, the importance of the carbamoylated residue was investigated in MurD, MurE and MurF by site-directed mutagenesis and chemical rescue experiments. Mutant proteins MurD K198A/F, MurE K224A and MurF K202A, which displayed low enzymatic activity, were rescued by incubation with short-chain carboxylic acids, but not amines. The best rescuing agent was acetate for MurD K198A, formate for K198F, and propionate for MurE K224A and MurF K202A. In the last of these, wild-type levels of activity were recovered. A complementarity between the volume of the residue replacing lysine and the length of the carbon chain of the acid was noted. These observations support a functional role for the carbamate in the three Mur synthetases. Experiments aimed at recovering an active enzyme by introducing an acidic residue in place of the invariant lysine residue were also undertaken. Mutant protein MurD K198E was weakly active and was rescued by formate, indicating the necessity of correct positioning of the acidic function with respect to the peptide backbone. Attempts at covalent rescue of mutant protein MurD K198C failed because of its lack of reactivity towards haloacids. [source] Modeling an active conformation for linear peptides and design of a competitive inhibitor for HMG-CoA reductaseJOURNAL OF MOLECULAR RECOGNITION, Issue 4 2008Valeriy V. Pak Abstract This study presents an approach that can be used to search for lead peptide candidates, including unconstrained structures in a recognized sequence. This approach was performed using the design of a competitive inhibitor for 3-hydroxy-3-methylglutaryl CoA reductase (HMGR). In a previous design for constrained peptides, a head-to-tail cyclic structure of peptide was used as a model of linear analog in searches for lead peptides with a structure close to an active conformation. Analysis of the conformational space occupied by the peptides suggests that an analogical approach can be applied for finding a lead peptide with an unconstrained structure in a recognized sequence via modeling a cycle using fixed residues of the peptide backbone. Using the space obtained by an analysis of the bioactive conformations of statins, eight cyclic peptides were selected for a peptide library based on the YVAE sequence as a recognized motif. For each cycle, the four models were assessed according to the design criterion ("V" parameter) applied for constrained peptides. Three cyclic peptides (FGYVAE, FPYVAE, and FFYVAE) were selected as lead cycles from the library. The linear FGYVAE peptide (IC50,=,0.4,µM) showed a 1200-fold increase the inhibitory activity compared to the first isolated LPYP peptide (IC50,=,484,µM) from soybean. Experimental analysis of the modeled peptide structures confirms the appropriateness of the proposed approach for the modeling of active conformations of peptides. Copyright © 2008 John Wiley & Sons, Ltd. [source] Backbone-modified amylin derivatives: implications for amyloid inhibitor design and as template for self-assembling bionanomaterials,JOURNAL OF PEPTIDE SCIENCE, Issue 11 2007Ronald C. Elgersma Abstract This report reviews our approach to the design, synthesis and structural/morphological analysis of backbone-modified amylin(20,29) derivatives. Depending on the position in the peptide backbone and the type of amide bond isostere/modification, the amylin(20,29) peptides behave either as inhibitors of amyloid fibril formation, which are able to retard amyloid formation of native amylin(20,29), or as templates for the formation of self-assembled supramolecular structures. Molecular fine-tuning of the hydrogen-bond accepting/donating properties allows the control over the morphology of the supramolecular aggregation motifs such as helical ribbons and tapes, ribbons progressing to closed peptide nanotubes, (twisted) lamellar sheets or amyloid fibrils. Copyright © 2007 European Peptide Society and John Wiley & Sons, Ltd. [source] Mass spectrometric identification of the trypsin cleavage pathway in lysyl-proline containing oligotuftsin peptidesJOURNAL OF PEPTIDE SCIENCE, Issue 4 2007Marilena Manea Abstract Trypsin cleaves specifically peptide bonds at the C -terminal side of lysine and arginine residues, except for -Arg-Pro- and -Lys-Pro- bonds which are normally resistant to proteolysis. Here we report evidence for a -Lys-Pro- tryptic cleavage in modified oligotuftsin derivatives, Ac-[TKPKG]4 -NH2) (1), using high-resolution mass spectrometry and HPLC as primary methods for analysis of proteolytic reactions. The proteolytic susceptibility of -Lys-Pro- bonds was strongly dependent on flanking residues, and the flexibility of the peptide backbone might be a prerequisite for this unusual cleavage. While -Lys-Gly- bonds in 1 were rapidly cleaved, the modification of these Lys residues by the attachment of a ß-amyloid(4,10) epitope to yield -Lys(X)-Gly derivatives prevented cleavage of this bond, and provided trypsin cleavage of -Lys-Pro- bonds, the pathway of this degradation being independent on the type of Lys- N, -side chains (acetyl group, amino acid, peptide). Substitution of the Lys residues by Ala at the P,2 positions decreased the tryptic cleavage, while replacement of the bulky side chain of Thr at the P2 positions strongly increased the cleavage of -Lys-Pro- bonds. Circular dichroism (CD) data of the modified oligotuftsin derivatives are in accord with enhanced flexibility of the peptide backbone, as a prerequisite for increased susceptibility to cleavage of -Lys-Pro- bonds. These results obtained of oligotuftsin derivatives might have implications for the proteolytic degradation of target peptides that require specific conformational preconditions. Copyright © 2007 European Peptide Society and John Wiley & Sons, Ltd. [source] Solution, solid phase and computational structures of apicidin and its backbone-reduced analogsJOURNAL OF PEPTIDE SCIENCE, Issue 6 2006Michael Kranz Abstract The recently isolated broad-spectrum antiparasitic apicidin (1) is one of the few naturally occurring cyclic tetrapeptides (CTP). Depending on the solvent, the backbone of 1 exhibits two ,-turns (in CH2Cl2) or a ,-turn (in DMSO), differing solely in the rotation of the plane of one of the amide bonds. In the X-ray crystal structure, the peptidic COs and NHs are on opposite sides of the backbone plane, giving rise to infinite stacks of cyclotetrapeptides connected by three intermolecular hydrogen bonds between the backbones. Conformational searches (Amber force field) on a truncated model system of 1 confirm all three backbone conformations to be low-energy states. The previously synthesized analogs of 1 containing a reduced amide bond exhibit the same backbone conformation as 1 in DMSO, which is confirmed further by the X-ray crystal structure of a model system of the desoxy analogs of 1. This similarity helps in explaining why the desoxy analogs retain some of the antiprotozoal activities of apicidin. The backbone-reduction approach designed to facilitate the cyclization step of the acyclic precursors of the CTPs seems to retain the conformational preferences of the parent peptide backbone. Copyright © 2005 European Peptide Society and John Wiley & Sons, Ltd. [source] Study of the conformational profile of the norbornane analogues of phenylalanineJOURNAL OF PEPTIDE SCIENCE, Issue 6 2002Arnau Cordomí Abstract The conformational profile of the eight stereoisomeric 2-amino-3-phenylnorbornane-2-carboxylic acids (2-amino-3-phenylbicyclo[2.2.1]heptane-2-carboxylic acids) has been assessed by computational methods. These molecules constitute a series of four enantiomeric pairs that can be considered as rigid analogues of either L - or D -phenylalanine. The conformational space of their N -acetyl methylamide derivatives has been explored within the molecular mechanics framework, using the parm94 set of parameters of the AMBER force field. Local minimum energy conformations have been further investigated at the ab initio level by means of the Hartree-Fock and second order Moller-Plesset perturbation energy calculations using a 6,31G(d) basis set. The results of the present work suggest that the bulky norbornane structure induces two kinds of conformational constraints on the residues. On one hand, those of a steric nature directly imposed by the bicycle on the peptide backbone and, on the other hand, those that limit the orientations attainable by the phenyl ring which, in turn, reduces further the flexibility of the peptide backbone. A comparative analysis of the conformational profile of the phenylnorbornane amino acids with that of the norbornane amino acids devoid of the ,-phenyl substituent suggests that the norbornane system hampers the residue to adopt extended conformations in favour of C7-like structures. However, the bicycle itself does not impart a clear preference for any of the two possible C7 minima. It is the aromatic side chain, which is forced to adopt an almost eclipsed orientation, that breaks this symmetry introducing a marked preference for a single region of the (,, ,) conformational space in each of the phenylalanine norbornane analogues investigated. Copyright © 2002 European Peptide Society and John Wiley & Sons, Ltd. [source] Formulation considerations for proteins susceptible to asparagine deamidation and aspartate isomerizationJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 11 2006Aditya A. Wakankar Abstract The asparagine (Asn) deamidation and aspartate (Asp) isomerization reactions are nonenzymatic intra-molecular reactions occurring in peptides and proteins that are a source of major stability concern in the formulation of these biomolecules. The mechanisms for the deamidation and isomerization reactions are similar since they both proceed through an intra-molecular cyclic imide (Asu) intermediate. The formation of the Asu intermediate, which involves the attack by nitrogen of the peptide backbone on the carbonyl carbon of the Asn or the Asp side chain, is the rate-limiting step in both the deamidation and the isomerization reactions at physiological pH. In this article, the influence of factors such as formulation conditions, protein primary sequence, and protein structure on the reactivity of Asn and Asp residues in proteins are reviewed. The importance of formulation conditions such as pH and solvent dielectric in influencing deamidation and isomerization reaction rates is addressed. Formulation strategies that could improve the stability of proteins to deamidation and isomerization reactions are described. The review is intended to provide information to formulation scientists, based on protein sequence and structure, to predict potential degradative sites on a protein molecule and to enable formulation scientists to set appropriate formulation conditions to minimize reactivity of Asn and Asp residues in protein therapeutics. © 2006 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 95:2321,2336, 2006 [source] Molecular structure study of dimethoxyphenyl-substituted phosphonodipeptides by infrared, Raman, and surface enhanced Raman spectroscopiesJOURNAL OF RAMAN SPECTROSCOPY, Issue 5 2006Edyta Podstawka Abstract In this study, Fourier-transform Raman (FT-RS) and infrared (FT-IR) absorption spectroscopies were employed to characterize the molecular structures of L -Ala-(3,4-dimethoxy)- L -Phe-PO3H2 (A) and L -Ala-(3,4-dimethoxy)-(des-CH2)- L -Phe-PO3H2 (B) phosphonodipeptides, where L -Phe denotes L -phenylalanine and L -Ala, L -alanine. The vibrational band assignments have been proposed. In order to determine the structures of these dimethoxyphenyl-substituted phosphonodipeptides adsorbed on a colloidal silver surface, surface enhanced Raman spectra (SERS) were measured. The analysis of SERS band intensities showed that these dimethoxyphenyl-substituted phosphonodipeptides directly interacted with the silver surface through the aromatic ring of Phe that adopted an orientation almost perpendicular to the silver surface. We also showed that intense enhancement of the ,as(NH2), ,as(PO32,), and ,(PO) modes of L -Ala-(3,4-dimethoxy)- L -Phe-PO3H2 suggested that these groups were mainly involved in the interaction with the silver colloid. Additionally, we proved that in the SERS spectrum of L -Ala-(3,4-dimethoxy)-(des-CH2)- L -Phe-PO3H2, several vibrations of the CH3 group were enhanced, indicating a flattened orientation of the peptide backbone on the silver surface. Copyright © 2005 John Wiley & Sons, Ltd. [source] Protein glycosylation analysis by HILIC-LC-MS of Proteinase K-generated N - and O -glycopeptidesJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010Gerhild Zauner Abstract Analysis of protein glycosylation is essential in order to correlate certain disease types with oligosaccharide structures on proteins. Here, a method for the MS characterization of site-specific protein glycosylation is presented. Using asialofetuin and fetuin as model substances, a protocol for glycopeptide dissection was developed based on unspecific proteolysis by Proteinase K. The resulting glycopeptides were then resolved by nanoscale hydrophilic interaction liquid chromatography-electrospray multistage MS. The early elution range of O -glycopeptides was clearly separated from the late elution range of N -glycopeptides. Glycopeptides were analyzed by ion trap-MS/MS, which revealed fragmentations of glycosidic linkages and some peptide backbone cleavages; MS3 spectra predominantly exhibited cleavages of the peptide backbone and provided essential information on the peptide sequence. The previously reported N - and O -glycan attachment sites of fetuin could be confirmed; moreover using our method, the occupation of a new, additional O -glycosylation site serine 296 was found. In conclusion, this approach appears to be a valuable technique for in-depth analysis of the site-specific N -glycosylation and O -glycosylation of individual glycoproteins. [source] Human CD4+ T-cell epitope repertoire on the C2 domain of coagulation factor VIIIJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 8 2003M. T. Reding Summary., Approximately 25% of severe hemophilia A patients develop antibodies (Ab) that neutralize the procoagulant function of factor (F)VIII (inhibitors). Autoimmune FVIII inhibitors may develop in individuals without congenital FVIII deficiency and cause acquired hemophilia. Low titers of anti-FVIII Ab may be present in hemophilia A patients without inhibitors and in healthy blood donors. FVIII-specific CD4+ T-cells drive the synthesis of anti-FVIII Ab. We examined the epitope repertoire of CD4+ T-cells from 15 healthy subjects, 10 hemophilia A patients without inhibitors, 11 hemophilia A patients with inhibitors, and six acquired hemophilia patients. Blood CD4+ T-cells were challenged in proliferation assays with a panel 16 overlapping synthetic peptides, spanning the sequence of the FVIII C2 domain. The sequence region 2291,2330 contained the most frequently and strongly recognized peptides in each of the four subject groups. Crystallographic B factor data and the location of these peptides within the three-dimensional structure of the C2 domain confirm that this region has a high degree of solvent exposure and flexibility within the peptide backbone, which are structural features typical of immunodominant universal CD4+ epitopes. Furthermore, this sequence region overlaps inhibitor-binding sites, suggesting that CD4+ T-cells recognizing peptide sequences within this region might be involved in inhibitor synthesis. The sequence regions 2191,2210 (recognized strongly by each study group except hemophilia A patients with inhibitors) and 2241,2290 (recognized primarily by acquired hemophilia patients and healthy subjects) share the same structural features, and also overlap inhibitor-binding sites. Although similar, there appear to be important differences in the CD4+ epitope repertoires of congenital and acquired hemophilia patients. [source] A statistically derived parameterization for the collagen triple-helixPROTEIN SCIENCE, Issue 11 2002Jan K. Rainey Abstract The triple-helix is a unique secondary structural motif found primarily within the collagens. In collagen, it is a homo- or hetero-tripeptide with a repeating primary sequence of (Gly-X-Y)n, displaying characteristic peptide backbone dihedral angles. Studies of bulk collagen fibrils indicate that the triple-helix must be a highly repetitive secondary structure, with very specific constraints. Primary sequence analysis shows that most collagen molecules are primarily triple-helical; however, no high-resolution structure of any entire protein is yet available. Given the drastic morphological differences in self-assembled collagen structures with subtle changes in assembly conditions, a detailed knowledge of the relative locations of charged and sterically bulky residues in collagen is desirable. Its repetitive primary sequence and highly conserved secondary structure make collagen, and the triple-helix in general, an ideal candidate for a general parameterization for prediction of residue locations and for the use of a helical wheel in the prediction of residue orientation. Herein, a statistical analysis of the currently available high-resolution X-ray crystal structures of model triple-helical peptides is performed to produce an experimentally based parameter set for predicting peptide backbone and C, atom locations for the triple-helix. Unlike existing homology models, this allows easy prediction of an entire triple-helix structure based on all existing high-resolution triple-helix structures, rather than only on a single structure or on idealized parameters. Furthermore, regional differences based on the helical propensity of residues may be readily incorporated. The parameter set is validated in terms of the predicted bond lengths, backbone dihedral angles, and interchain hydrogen bonding. [source] Comparison of collision- versus electron-induced dissociation of Pt(II) ternary complexes of histidine- and methionine-containing peptides,RAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 19 2009Linda Feketeová Incubation of the histidine-containing peptides (GH, HG, GGH, GHG, HGG) and methionine-containing peptides (GM, MG, GGM, GMG, MGG) with the platinum complexes [Pt(terpy)Cl]+ (A) and [Pt(dien)Cl]+ (B) followed by electrospray ionisation (ESI) led to a number of singly and doubly charged ternary platinum peptide complexes, including [Pt(L)M]2+ and [Pt(L)M,H]+ (where L,=,the ligand terpy or dien; M is a peptide). Each of the [Pt(L)M]2+ complexes was subjected to electron capture dissociation (ECD), collision-induced dissociation (CID) and electron-induced dissociation (EID), while each of the [Pt(L)M,H]+ complexes was subjected to CID and EID. Results from ECD suggest that the free electron is captured by the metal ion thus weakening the bonds to its ligands. In the case of the ligand terpy, which binds more strongly than dien, this weakening leads to the loss of the peptide. The minor products in the ECD spectra of [Pt(terpy)M]2+ complexes do show fragmentation along the peptide backbone, but the ions observed are of the a-, b-, and y-type. For the complexes with methionine-containing peptides, a marker ion, [Pt(L)SCH3]+, was found which is indicative of binding of Pt to the methionine side chain. For the histidine-containing peptides, an ion containing platinum, the auxiliary ligand, and the histidine imine was observed in many instances, thus indicating the binding of the histidine side chain to the metal, but other modes of Pt coordination (N-terminus) were also found to be competitive. These findings are consistent with a recent finding (Sze et al. J. Biol. Inorg. Chem. 2009; 14: 163) that Pt occupies the methionine-rich copper(I)-binding site rather than histidine-rich copper(II)-binding site in the CopC protein. Copyright © 2009 John Wiley & Sons, Ltd. [source] Atmospheric pressure matrix-assisted laser desorption/ionization ion trap mass spectrometry of sulfonic acid derivatized tryptic peptidesRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 23 2001T. Keough Atmospheric pressure matrix-assisted laser desorption/ionization (AP-MALDI) and ion trap mass spectrometry have been used to study the fragmentation behavior of native peptides and peptide derivatives prepared for de novo sequencing applications. Sulfonic acid derivatized peptides were observed to fragment more extensively and up to 28 times more efficiently than the corresponding native peptides. Tandem mass spectra of native peptides containing aspartic or glutamic acids are dominated by cleavage on the C-terminal side of the acidic residues. This significantly limits the amount of sequence information that can be derived from those compounds. The MS/MS spectra of native tryptic peptides containing oxidized Met residues show extensive loss of CH3SOH and little sequence-specific fragmentation. On the other hand, the tandem mass spectra of derivatized peptides containing Asp, Glu and oxidized Met show much more uniform fragmentation along the peptide backbone. The AP-MALDI tandem mass spectra of some derivatized peptides were shown to be qualitatively very similar to the corresponding vacuum MALDI postsource decay mass spectra, which were obtained on a reflector time-of-flight instrument. However, the ion trap mass spectrometer offers several advantages for peptide sequencing relative to current reflector time-of-flight instruments including improved product ion mass measurement accuracy, improved precursor ion selection and MSn. These latter capabilities were demonstrated with solution digests of model proteins and with in-gel digests of 2D-gel separated proteins. Copyright © 2001 John Wiley & Sons, Ltd. [source] Nociceptin and its natural and specifically-modified fragments: Structural studiesBIOPOLYMERS, Issue 12 2010E. Podstawka-Proniewicz Abstract The vibrational structures of Nociceptin (FQ), its short bioactive fragments, and specifically-modified [Tyr1]FQ (1-6), [His1]FQ (1-6), and [His1,4]FQ (1-6) fragments were characterized. We showed that in the solid state, all of the aforementioned peptides except FQ adopt mainly turn and disordered secondary structures with a small contribution from an antiparallel ,-sheet conformation. FQ (1-11), FQ (7-17) [His1]FQ (1-6), and [His1,4]FQ (1-6) have an ,-helical backbone arrangement that could also slightly influence their secondary structure. The adsorption behavior of these peptides on a colloidal silver surface in an aqueous solution (pH = ,8.3) was investigated by means of surface-enhanced Raman scattering (SERS). All of the peptides, excluding FQ (7-17), chemisorbed on the colloidal silver surfaces through a Phe4 residue, which for FQ, FQ (1-11), FQ (1-6), [Tyr1]FQ (1-6), and [His1]FQ (1-6) lies almost flat on this surface, while for FQ (1-13) and FQ (1-13)NH2 adopts a slightly tilted orientation with respect to the surface. The Tyr1 residue in [Tyr1]FQ (1-6) does not interact with the colloidal silver surface, suggesting that the Tyr1 and Phe4 side chains are located on the opposite sides of the peptide backbone, which can be also true for His1 and Phe4 in [His1]FQ (1-6). The lone pair of electrons on the oxygen atom of the ionized carbonyl group of FQ (1-13) and FQ (7-17) appears to be coordinated to the colloidal silver nanoparticles, whereas in the case of the remaining peptides, it only assists in the adsorption process, similar to the NH2 group. We also showed that upon adsorption, the secondary structure of these peptides is altered. © 2010 Wiley Periodicals, Inc. Biopolymers 93: 1039,1054, 2010. This article was originally published online as an accepted preprint. The "Published Online" date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com [source] Model peptide-based system used for the investigation of metal ions binding to histidine-containing polypeptidesBIOPOLYMERS, Issue 6 2010Manuela Murariu Abstract The reaction of histidine-containing polypeptides with toxic and essential metals and the molecular mechanism of complexation has yet to be determined, particularly with respect to the conformational changes of the interacting macromolecules. Therefore, a system of oligopeptides containing histidine residues in various positions of Ala or Gly sequences has been designed and used in heavy metal comparatively binding experiments. The role of spacing residues (Gly and Ala repeats) in selecting the various conformations was investigated. The newly synthesized peptides and metal ion adducts have been characterized by Fourier transform infrared spectroscopy (FTIR) as well as electrospray ion trap mass spectrometry (ESI,MS) and circular dichroism (CD). The analysis of CD-spectra of the four peptides in water revealed that the secondary structure depends much on the position of each amino acid in the peptide backbone. Our peptides system reveals various binding mechanisms of metal ions to peptides depending on the position of histidine residue and the corresponding conformations of Ala or Gly sequences. Biological and medical consequences of conformational changes of metal-bound peptides are further discussed. Thus, the binding of heavy metals to four peptides may serve as a model system with respect to the conformational consequences of the metal addition on the amino acid repeats situated in prion protein. © 2010 Wiley Periodicals, Inc. Biopolymers 93:497,508, 2010. This article was originally published online as an accepted preprint. The "Published Online" date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com [source] 4-Fluorophenylglycine as a Label for 19F NMR Structure Analysis of Membrane-Associated PeptidesCHEMBIOCHEM, Issue 11 2003Sergii Afonin Abstract The non-natural amino acid 4-fluorophenylglycine (4F-Phg) was incorporated into several representative membrane-associated peptides for dual purpose. The 19F-substituted ring is directly attached to the peptide backbone, so it not only provides a well-defined label for highly sensitive 19F NMR studies but, in addition, the D and L enantiomers of the stiff side chain may serve as reporter groups on the transient peptide conformation during the biological function. Besides peptide synthesis, which is accompanied by racemisation of 4F-Phg, we also describe separation of the epimers by HPLC and removal of trifluoroacetic acid. As a first example, 18 different analogues of the fusogenic peptide "B18" were prepared and tested for induction of vesicle fusion; the results confirmed that hydrophobic sites tolerated 4F-Phg labelling. Similar fusion activities within each pair of epimers suggest that the peptide is less structured in the fusogenic transition state than in the helical ground state. In a second example, five doubly labelled analogues of the antimicrobial peptide gramicidin S were compared by using bacterial growth inhibition assays. This cyclic ,-sheet peptide could accommodate both L and D substituents on its hydrophobic face. As a third example, we tested six analogues of the antimicrobial peptide PGLa. The presence of d- 4F-Phg reduced the biological activity of the peptide by interfering with its amphiphilic ,-helical fold. Finally, to illustrate the numerous uses of l- 4F-Phg in 19F NMR spectroscopy, we characterised the interaction of labelled PGLa with uncharged and negatively charged membranes. Observing the signal of the free peptide in an aqueous suspension of unilamellar vesicles, we found a linear saturation behaviour that was dominated by electrostatic attraction of the cationic PGLa. Once the peptide is bound to the membrane, however, solid-state 19F NMR spectroscopy of macroscopically oriented samples revealed that the charge density has virtually no further influence on the structure, alignment or mobility of the peptide. [source] Synthesis and Structural Model of an ,(2,6)-Sialyl-T Glycosylated MUC1 Eicosapeptide under Physiological ConditionsCHEMISTRY - A EUROPEAN JOURNAL, Issue 19 2006Sebastian Dziadek Dr. Abstract To study the effect of O-glycosylation on the conformational propensities of a peptide backbone, a 20-residue peptide (GSTAPPAHGVTSAPDTRPAP) representing the full length tandem repeat sequence of the human mucin MUC1 and its analogue glycosylated with the (2,6)-sialyl-T antigen on Thr11, were prepared and investigated by NMR and molecular modeling. The peptides contain both the GVTSAP sequence, which is an effective substrate for GalNAc transferases, and the PDTRP fragment, a known epitope recognized by several anti-MUC1 monoclonal antibodies. It has been shown that glycosylation of threonine in the GVTSAP sequence is a prerequisite for subsequent glycosylation of the serine at GVTSAP. Furthermore, carbohydrates serve as additional epitopes for MUC1 antibodies. Investigation of the solution structure of the sialyl-T glycoeicosapeptide in a H2O/D2O mixture (9:1) under physiological conditions (25,°C and pH 6.5) revealed that the attachment of the saccharide side-chain affects the conformational equilibrium of the peptide backbone near the glycosylated Thr11 residue. For the GVTSA region, an extended, rod-like secondary structure was found by restrained molecular dynamics simulation. The APDTR region formed a turn structure which is more flexibly organized. Taken together, the joined sequence GVTSAPDTR represents the largest structural model of MUC1 derived glycopeptides analyzed so far. [source] Modulating Charge Transfer through Cyclic D,L -,-Peptide Self-AssemblyCHEMISTRY - A EUROPEAN JOURNAL, Issue 4 2005W. Seth Horne Abstract We describe a concise, solid support-based synthetic method for the preparation of cyclic d,l -,-peptides bearing 1,4,5,8-naphthalenetetracarboxylic acid diimide (NDI) side chains. Studies of the structural and photoluminescence properties of these molecules in solution show that the hydrogen bond-directed self-assembly of the cyclic d,l -,-peptide backbone promotes intermolecular NDI excimer formation. The efficiency of NDI charge transfer in the resulting supramolecular assemblies is shown to depend on the length of the linker between the NDI and the peptide backbone, the distal NDI substituent, and the number of NDIs incorporated in a given structure. The design rationale and synthetic strategies described here should provide a basic blueprint for a series of self-assembling cyclic d,l -,-peptide nanotubes with interesting optical and electronic properties. [source] Molecular Modeling Suggests Conformational Scaffolds Specifically Targeting Five Subtypes of Somatostatin ReceptorsCHEMICAL BIOLOGY & DRUG DESIGN, Issue 3 2007Gregory V. Nikiforovich Several analogs of somatostatin with conformational constraints in their peptide backbones have been modeled to determine energetically feasible conformations. Comparison of low-energy backbone structures of these peptides suggested unique conformations of the central Phe/Alai -D-Trpi+1 -Lysi+2 -Thri+3 fragment characteristic for specific interactions of somatostatin with each of the five distinct subtypes of somatostatin receptors (SSTRs). The conformations obtained were in good agreement with experimental data obtained earlier by NMR measurements and/or X-ray crystallography. The results help rationalize experimental observations on the specificity of binding of various somatostatin analogs with different subtypes of the SSTRs. They also serve as templates for the design of conformationally constrained non-peptide scaffolds that effectively and selectively interact with different subtypes of SSTRs. Such scaffolds can be convenient carriers of radiolabels and near-infrared labels in specific agents for imaging tumors expressing different SSTR subtypes. [source] | ||||||||||||||||