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Tetsuo Asakura - One of the best experts on this subject based on the ideXlab platform.

  • conformational change of 13c labeled 47 mer model peptides of nephila clavipes dragline silk in poly vinyl alcohol film by stretching studied by 13c solid state nmr and molecular dynamics simulation
    International Journal of Biological Macromolecules, 2019
    Co-Authors: Tetsuo Asakura, Hironori Matsuda, Akihiro Aoki, Naomi Kataoka, Akiko Imai
    Abstract:

    Abstract For determination of the conformation of irregular sequences in glycine-rich region of the Nephila clavipes spider dragline silk, the combination of 13C selectively labeled model peptides for the typical primary structure and their 13C solid-state NMR observations is very useful (T. Asakura et al. Macromolecules. 51 (2018) 3608–3619). However, spiders produce the fiber through the stretching process in nature and therefore, it is difficult to study conformational change by stretching as mimic using the model peptides because these are generally in the powder form. In this paper, 13C selectively labeled three model peptides, (Glu)4(Ala)6Gly Gly 12 Ala 13 Gly 14 GlnGlyGlyTyrGlyGlyLeuGlySerGln Gly 25 Ala 26 Gly 27 ArgGly-GlyLeuGlyGlyGln Gly 35 Ala 36 Gly 37 (Ala)6(Glu)4 with three underlined 13C labeled blocks and their poly(vinyl alcohol) blend films were prepared and the conformational changes of these peptides were monitored by stretching of the films using 13C solid-state NMR. In addition, the molecular dynamics simulation was done to evaluate change in the conformation of the sequence by stretching theoretically. The fractions of β-sheet of Ala36 and Gly37 residues in glycine-rich region adjacent to the C-terminal (Ala)6 sequence increased significantly by stretching compared with those of other 13C labeled Ala and Gly residues.

  • solid state nmr analysis of a peptide gly pro gly gly ala 6 gly derived from a flagelliform silk sequence of nephila clavipes
    Biomacromolecules, 2006
    Co-Authors: Kosuke Ohgo, Taiji Kawase, Jun Ashida, Tetsuo Asakura
    Abstract:

    Solid-state NMR is especially useful when the structures of peptides and proteins should be analyzed by taking into account the structural distribution, that is, the distribution of the torsion angle of the individual residue. In this study, two-dimensional spin-diffusion solid-state NMR spectra of 13C-double-labeled model peptides (GPGGA)6G of flagelliform silk were observed for studying the local structure in the solid state. The spin-diffusion NMR spectra calculated by assuming the torsion angles of the β-spiral structure exclusively could not reproduce the observed spectra. In contrast, the spectra calculated by taking into account the statistical distribution of the torsion angles of the individual central residues in the sequences Ala-Gly-Pro, Gly-Pro-Gly, Pro-Gly-Gly, Gly-Gly-Ala, and Gly-Ala-Gly from PDB data could reproduce the observed spectra well. This indicates that the statistical distribution of the torsion angles should be considered for the structural model of (GPGGA)6G similar to the cas...

  • the role of irregular unit gaas on the secondary structure of bombyx mori silk fibroin studied with 13c cp mas nmr and wide angle x ray scattering
    Protein Science, 2002
    Co-Authors: Tetsuo Asakura, Rena Sugino, Tatsushi Okumura, Yasumoto Nakazawa
    Abstract:

    Bombyx mori silk fibroin is a fibrous protein whose fiber is extremely strong and tough, although it is produced by the silkworm at room temperature and from an aqueous solution. The primary structure is mainly Ala-Gly alternative copolypeptide, but Gly-Ala-Ala-Ser units appear frequently and periodically. Thus, this study aims at elucidating the role of such Gly-Ala-Ala-Ser units on the secondary structure. The sequential model peptides containing Gly-Ala-Ala-Ser units selected from the primary structure of B. mori silk fibroin were synthesized, and their secondary structure was studied with 13C CP/MAS NMR and wide-angle X-ray scattering. The 13C isotope labeling of the peptides and the 13C conformation-dependent chemical shifts were used for the purpose. The Ala-Ala units take antiparallel β-sheet structure locally, and the introduction of one Ala-Ala unit in (Ala-Gly)15 chain promotes dramatical structural changes from silk I (repeated β-turn type II structure) to silk II (antiparallel β-sheet structure). Thus, the presence of Ala-Ala units in B. mori silk fibroin chain will be one of the inducing factors of the structural transition for silk fiber formation. The role of Tyr residue in the peptide chain was also studied and clarified to induce "locally nonordered structure."

  • Chain-folded lamellar structure and dynamics of the crystalline fraction of Bombyx mori silk fibroin and of (Ala-Gly-Ser-Gly-Ala-Gly)n model peptides.
    International Journal of Biological Macromolecules, 2026
    Co-Authors: Tetsuo Asakura, Tatsuya Ogawa, Akira Naito, Michael P Williamson
    Abstract:

    Abstract Solid-state NMR is a powerful analytical technique to determine the composite structure of Bombyx mori silk fibroin (SF). In our previous paper, we proposed a lamellar structure for Ala-Gly copolypeptides as a model of the crystalline fraction in Silk II. In this paper, the structure and dynamics of the crystalline fraction and of a better mimic of the crystalline fraction, (Ala-Gly-Ser-Gly-Ala-Gly)n (n = 2–5, 8), and 13C selectively labeled [3-13C]Ala-(AGSGAG)5 in Silk II forms, were studied using structural and dynamical analyses of the Ala Cβ peaks in 13C cross polarization/ magic angle spinning NMR and 13C solid-state spin-lattice relaxation time (T1) measurements, respectively. Like Ala-Gly copolypeptides, these materials have lamellar structures with two kinds of Ala residues in β-sheet, A and B, plus one distorted β-turn, t, formed by repetitive folding using β-turns every eighth amino acid in an antipolar arrangement. However, because of the presence of Ser residues at every sixth residue in (AGSGAG)n, the T1 values and mobilities of B decreased significantly. We conclude that the Ser hydroxyls hydrogen bond to adjacent lamellar layers and fix them together in a similar way to Velcro®.

Arnold De Loof - One of the best experts on this subject based on the ideXlab platform.

  • substitution of conserved glycine residue by alanine in natural and synthetic neuropeptide ligands causes partial agonism at the stomoxytachykinin receptor
    Journal of Neurochemistry, 2004
    Co-Authors: Jeroen Poels, Tom Van Loy, Vanessa Franssens, Michel Detheux, Ronald J Nachman, Hendrica B Oonk, Karl E O Akerman, Gilbert Vassart, Marc Parmentier, Arnold De Loof
    Abstract:

    A few naturally occurring insect tachykinin-related peptides, such as stomoxytachykinin (Stc-TK), contain an Ala-residue instead of the highly conserved Gly-residue that is present in most other members of this peptide family. Stc-TK is a potent, partial agonist of the stable fly (Stomoxys calcitrans) tachykinin receptor, STKR. By means of synthetic analogues, the Gly/Ala exchange, representing the addition of a single methyl group in the active core region of these peptides, was shown to be fully responsible for the generation of this partial agonism, which was also accompanied by an increase in agonistic potency. Surprisingly, this Ala-dependent reduction in maximal response levels was only observed for the agonist-induced cellular calcium rise. Stomoxytachykinin, Stc-TK, did not display partial agonism for the STKR-mediated cyclic AMP response. A possible explanation for this differential partial agonism is that the Gly-containing and Ala-replaced peptides recognize and stabilize active receptor conformations that differ in their functional coupling efficacies towards these response pathways. Drosotachykinins, Drm-TK, tachykinin-like peptides encoded in the fruit fly genome, were shown to be STKR-agonists. Interestingly, one of these peptides, which contains an Ala-residue instead of the conserved Gly-residue, also proved to be a potent, partial agonist for STKR.

  • identification of one tachykinin and two kinin related peptides in the brain of the white shrimp penaeus vannamei
    Biochemical and Biophysical Research Communications, 1998
    Co-Authors: Julie Nieto, Dirk Veelaert, Bart Devreese, Jozef Van Beeumen, Rita Derua, Etienne Waelkens, Anja Cerstiaens, Geoffrey M Coast, Jorgen Calderon, Arnold De Loof
    Abstract:

    This paper reports the purification of three myotropic neuropeptides from the white shrimpPenaeus vannamei.The central nervous systems of 3500 shrimps were extracted in an acidified solvent, after which four to five HPLC column systems were used to obtain pure peptides. A cockroach hindgut muscle contraction bioassay was used to monitor all collected fractions. The pure peptides were submitted to Edman degradation based automated microsequencing. Mass spectrometry and chemical synthesis confirmed the sequences. Ala-Pro-Ser-Gly-Phe-Leu-Gly-Met-Arg-NH2(Pev-tachykinin, 934.1 Da) belongs to the tachykinin family with identified members in all vertebrate classes and some invertebrate classes: arthropods, annelids and molluscs. A very specific Pev-tachykinin antiserum was developed, which labels 4 neurosecretory cells in the brain. Ala-Ser-Phe-Ser-Pro-Trp-Gly-NH2(Pev-kinin 1, 749.8 Da) and Asp-Phe-Ser-Ala-Trp-Ala-NH2(Pev-kinin 2, 694.7 Da) are the first crustacean kinins. Pev-kinin 2 is the first kinin with a Trp-Ala-NH2instead of a kinin-typical Trp-Gly-NH2carboxyterminus.

  • isolation and characterization of eight myoinhibiting peptides from the desert locust schistocerca gregaria new members of the cockroach allatostatin family
    Molecular and Cellular Endocrinology, 1996
    Co-Authors: Dirk Veelaert, Bart Devreese, Liliane Schoofs, Jozef Van Beeumen, Jozef Vanden Broeck, Stephen S Tobe, Arnold De Loof
    Abstract:

    Eight myoinhibiting peptides were purified by high performance liquid chromatography from a methanolic extract of 7000 brains of the desert locust, Schistocerca gregaria. Complete sequences were obtained via a novel, combined approach employing: (1) chemical microsequencing and (2) post-source decay analysis on a reflectron time-of-flight mass spectrometer using matrix-assisted laser desorption/ionisation. Each of the peptides shows C-terminal amino acid sequence similarity to cockroach and cricket allatostatins and to blowfly callatostatins. Therefore, these novel peptides were designated Schistocerca gregaria allatostatins (Scg-ASTs) or schistostatins and their primary structures were determined to be: Ala-Tyr-Thr-Tyr-Val-Ser-Glu-Tyr-Lys-Arg-Leu-Pro-Val-Tyr-Asn-Phe-Gly-Leu- NH2 (Scg-AST-2), Ala-Thr-Gly-Ala-Ala-Ser-Leu-Tyr-Ser-Phe-Gly-Leu-NH2 (Scg-AST-3), Gly-Pro-Arg-Thr-Tyr-Ser-Phe-Gly-Leu-NH2 (Scg-AST-4), Gly-Arg-Leu-Tyr-Ser-Phe-Gly-Leu-NH2 (Scg-AST-5), Ala-Arg-Pro-Tyr-Ser-Phe-Gly-Leu-NH2 (Scg-AST-6), Ala-Gly-Pro-Ala-Pro-Ser-Arg-Leu-Tyr-Ser-Phe-Gly-Leu-NH2 (Scg-AST-7), Glu-Gly-Arg-Met-Tyr-Ser-Phe-Gly-Leu-NH2 (Scg-AST-8), and Ala-Pro-Ala-Glu-His-Arg-Phe-Ser-Phe-Gly-Leu-NH2 (Scg-AST-10). Synthetic Scg-AST peptides inhibit the peristaltic movements of the oviduct of S. gregaria. Although all eight peptides show potent inhibitory effects on juvenile hormone (JH) biosynthesis by corpora allata (CA) of the cockroach Diploptera punctata, no allatostatic effects were observed on CA of the desert locust (S. gregaria).

Michael J Conlon - One of the best experts on this subject based on the ideXlab platform.

  • the alyteserins two families of antimicrobial peptides from the skin secretions of the midwife toad alytes obstetricans alytidae
    Peptides, 2009
    Co-Authors: Michael J Conlon, Anni Demandt, Per F. Nielsen, Hubert Vaudry, Jérôme Leprince, Douglas C. Woodhams
    Abstract:

    Abstract Two families of structurally related C-terminally α-amidated antimicrobial peptides have been identified in norepinephrine-stimulated skin secretions of the midwife toad Alytes obstetricans (Alytidae). The alyteserin-1 peptides (Gly-Leu-Lys-(Asp/Glu)-Ile-Phe-Lys-Ala-Gly-Leu-Gly-Ser-Leu-Val-Lys-(Gly/Asn)-Ile-Ala-Ala-His-Val-Ala-(Asn/Ser).NH 2 ) show limited structural similarity to the ascaphins from the skins of frogs of the family Leiopelmatidae. Alyteserin-2a (Ile-Leu-Gly-Lys-Leu-Leu-Ser-Thr-Ala-Ala-Gly-Leu-Leu-Ser-Asn-Leu.NH 2 ) and alyteserin-2b and -2c (Ile-Leu-Gly-Ala-Ile-Leu-Pro-Leu-Val-Ser-Gly-Leu-Leu-Ser-(Asn/Ser)-Lys-Leu.NH 2 ) show limited sequence identity with bombinin H6, present in the skins of frogs of the family Bombinatoridae. The alyteserin-1 peptides show selective growth inhibitory activity against the Gram-negative bacteria Escherichia coli (MIC = 25 μM) whereas alyteserin-2a is more potent against the Gram-positive bacteria Staphylococcus aureus (MIC = 50 μM). The hemolytic activity against human erythrocytes of all peptides tested is relatively weak (LC 50  > 100 μM). The data demonstrate that the frogs belonging to the family Alytidae are among those producing dermal antimicrobial peptides that may represent a component of the animal's system of innate immunity.

  • purification and characterization of galanin and scyliorhinin i from the hybrid sturgeon scaphirhynchus platorynchus scaphirhynchus albus acipenseriformes
    General and Comparative Endocrinology, 1999
    Co-Authors: Yuqi Wang, Per F. Nielsen, Bruce A Barton, Michael J Conlon
    Abstract:

    Abstract The sturgeons (order Acipenseriformes) are extant representatives of a group of ancient Actinopterygian (ray-finned) fish. Galanin and scyliorhinin I (a tachykinin with limited structural similarity to mammalian substance P) have been isolated from an extract of the gastrointestinal tract of a sturgeon (an F1 hybrid between the shovelnose sturgeon, Scaphirhynchus platorynchus, and the pallid sturgeon, Scaphirhynchus albus ). The primary structure of sturgeon galanin (Gly-Trp-Thr-Leu-Asn-Ser-Ala-Gly-Tyr-Leu 10 -Leu-Gly-Pro-His-Ala-Val-Asp-Gly-His-Arg 20 -Ser-Leu-Ser-Asp-Lys-His-Gly-Leu-Pro·NH 2 ) contains only two amino acid substitutions (Ser 23 → Asn and Pro 29 → Ala) compared with galanin from the bowfin, Amia calva (Amiiformes), but five amino acid substitutions compared with galanin from the trout (Teleostei). Similarly, the sturgeon tachykinin (Ser-Lys-Tyr-His-Gln-Phe-Tyr-Gly-Leu-Met·NH 2 ) contains only one amino acid substitution (Tyr 3 → Ser) compared with scyliorhinin I previously isolated from bowfin stomach but five amino acid substitutions compared with trout substance P. The data support the hypothesis that the Acipenseriformes and the basal Neopterygians (gars and bowfin) share a close phylogenetic relationship.

  • primary structure of insulin from the african lungfish protopterus annectens
    General and Comparative Endocrinology, 1997
    Co-Authors: Michael J Conlon, James E Platz, Per F. Nielsen, Hubert Vaudry, Mauro Vallarino
    Abstract:

    Among the extant Sarcopterygii, the interrelationship between the Dipnoi (lungfishes), Actinistia (coelacanths), and Tetrapoda (tetrapods) is controversial. Insulin has been purified from an extract of the pancreas of the African lungfishProtopterus annectensand its primary structure established as A-chain, Gly–Ile–Val–Glu–Gln–Cys–Cys–His–Lys–Pro10–Cys–Ser–Leu– Tyr –Glu–Leu–Glu–Asn–Tyr–Cys20–Asn–Val–Pro; and B-chain, Ala–Val–Leu–Asn–Gln–His–Leu–Cys–Gly–Ser10–His–Leu–Val– Glu– Ala–Leu–Tyr–Leu–Val–Cys20–Ala–Asp–Asn–Gly–Phe– Phe–Tyr–Lys–Pro–Ser30–Gly. Lungfish insulin contains unusual structural features, such as the dipeptide extension to the C-terminus of the A-chain and the substitution Arg → Asn at position B-23 in the putative receptor binding region of insulin, which may be expected to influence appreciably its biological potency relative to mammalian insulins. Lungfish insulin also contains amino acid substitutions such as Gly → Ala at position B-21, Glu → Asp at position B-22, and a Lys → Ser residue at position B-30, previously found in insulins from amphibia. This observation is consistent with paleontological data suggesting that lungfish and amphibia share a close phylogenetic relationship.

Nenad M Kostic - One of the best experts on this subject based on the ideXlab platform.

  • palladium ii complexes as synthetic peptidases regioselectively cleave the second peptide bond upstream from methionine and histidine side chains
    Journal of the American Chemical Society, 2002
    Co-Authors: Nebojsa M Milovic, Nenad M Kostic
    Abstract:

    Palladium(II) complexes promote hydrolysis of natural and synthetic oligopeptides with unprecedented regioselectivity; the only cleavage site is the second peptide bond upstream from a methionine or a histidine side chain, that is, the bond involving the amino group of the residue that precedes this side chain. We investigate this regioselectivity with four N-acetylated peptides as substrates: neurotransmitter methionine enkephalin (Ac-Tyr-Gly-Gly-Phe-Met) and synthetic peptides termed Met-peptide (Ac-Ala-Lys-Tyr-Gly-Gly-Met-Ala-Ala-Arg-Ala), His-peptide (Ac-Val-Lys-Gly-Gly-His-Ala-Lys-Tyr-Gly-Gly-Met(OX)-Ala-Ala-Arg-Ala), in which a Met is oxidized to sulfone, and HisMet-peptide (Ac-Val-Lys-Gly-Gly-His-Ala-Lys-Tyr-Gly-Gly-Met-Ala-Ala-Arg-Ala). While maintaining protein-like properties, these substrates are suitable for quantitative study since their coordination to Pd(II) ion can be determined (by NMR spectroscopy), and the cleavage fragments can be separated (by HPLC methods) and identified (by MALDI mass spectrometry). The only peptide bonds cleaved were the Gly3-Phe4 bond in methionine enkephalin, Gly4-Gly5 bond in Met-peptide, Gly3-Gly4 in His-peptide, and Gly3-Gly4 and Gly9-Gly10 bonds in HisMet-peptide. We explain this consistent regioselectivity of cleavage by studying the modes of Met-peptide coordination to the Pd(II) ion in [Pd(H(2)O)(4)](2+) complex. In acidic solution, the rapid attachment of the Pd(II) complex to the methionine side chain is followed by the interaction of the Pd(II) ion with the peptide backbone upstream from the anchor. In the hydrolytically active complex, Met-peptide is coordinated to Pd(II) ion as a bidentate ligand - via sulfur atom in the methionine side chain and the first peptide nitrogen upstream from this anchor - so that the Pd(II) complex approaches the scissile peptide bond. Because the increased acidity favors this hydrolytically active complex, the rate of cleavage guided by either histidine or methionine anchor increased as pH was lowered from 4.5 to 0.5. The unwanted additional cleavage of the first peptide bond upstream from the anchor is suppressed if pH is kept above 1.2. Four Pd(II) complexes cleave Met-peptide with the same regioselectivity but at somewhat different rates. Complexes in which Pd(II) ion carries labile ligands, such as [Pd(H(2)O)(4)](2+) and [Pd(NH(3))(4)](2+), are more reactive than those containing anionic ligands, such as [PdCl(4)](2)(-), or a bidentate ligand, such as cis-[Pd(en)(H(2)O)(2)](2+). When both methionine and histidine residues are present in the same substrate, as in HisMet-peptide, 1 molar equivalent of the Pd(II) complex distributes itself evenly at both anchors and provides partial cleavage, whereas 2 molar equivalents of the promoter completely cleave the second peptide bond upstream from each of the anchors. The results of this study bode well for growing use of palladium(II) reagents in biochemical and bioanalytical practice.

  • palladium ii complexes as synthetic peptidases regioselectively cleave the second peptide bond upstream from methionine and histidine side chains
    Journal of the American Chemical Society, 2002
    Co-Authors: Nebojsa M Milovic, Nenad M Kostic
    Abstract:

    Palladium(II) complexes promote hydrolysis of natural and synthetic oligopeptides with unprecedented regioselectivity; the only cleavage site is the second peptide bond upstream from a methionine or a histidine side chain, that is, the bond involving the amino group of the residue that precedes this side chain. We investigate this regioselectivity with four N-acetylated peptides as substrates:  neurotransmitter methionine enkephalin (Ac-Tyr-Gly-Gly-Phe-Met) and synthetic peptides termed Met-peptide (Ac-Ala-Lys-Tyr-Gly-Gly-Met-Ala-Ala-Arg-Ala), His-peptide (Ac-Val-Lys-Gly-Gly-His-Ala-Lys-Tyr-Gly-Gly-MetOX-Ala-Ala-Arg-Ala), in which a Met is oxidized to sulfone, and HisMet-peptide (Ac-Val-Lys-Gly-Gly-His-Ala-Lys-Tyr-Gly-Gly-Met-Ala-Ala-Arg-Ala). While maintaining protein-like properties, these substrates are suitable for quantitative study since their coordination to Pd(II) ion can be determined (by NMR spectroscopy), and the cleavage fragments can be separated (by HPLC methods) and identified (by MALDI mas...

Douglas C. Woodhams - One of the best experts on this subject based on the ideXlab platform.

  • the alyteserins two families of antimicrobial peptides from the skin secretions of the midwife toad alytes obstetricans alytidae
    Peptides, 2009
    Co-Authors: Michael J Conlon, Anni Demandt, Per F. Nielsen, Hubert Vaudry, Jérôme Leprince, Douglas C. Woodhams
    Abstract:

    Abstract Two families of structurally related C-terminally α-amidated antimicrobial peptides have been identified in norepinephrine-stimulated skin secretions of the midwife toad Alytes obstetricans (Alytidae). The alyteserin-1 peptides (Gly-Leu-Lys-(Asp/Glu)-Ile-Phe-Lys-Ala-Gly-Leu-Gly-Ser-Leu-Val-Lys-(Gly/Asn)-Ile-Ala-Ala-His-Val-Ala-(Asn/Ser).NH 2 ) show limited structural similarity to the ascaphins from the skins of frogs of the family Leiopelmatidae. Alyteserin-2a (Ile-Leu-Gly-Lys-Leu-Leu-Ser-Thr-Ala-Ala-Gly-Leu-Leu-Ser-Asn-Leu.NH 2 ) and alyteserin-2b and -2c (Ile-Leu-Gly-Ala-Ile-Leu-Pro-Leu-Val-Ser-Gly-Leu-Leu-Ser-(Asn/Ser)-Lys-Leu.NH 2 ) show limited sequence identity with bombinin H6, present in the skins of frogs of the family Bombinatoridae. The alyteserin-1 peptides show selective growth inhibitory activity against the Gram-negative bacteria Escherichia coli (MIC = 25 μM) whereas alyteserin-2a is more potent against the Gram-positive bacteria Staphylococcus aureus (MIC = 50 μM). The hemolytic activity against human erythrocytes of all peptides tested is relatively weak (LC 50  > 100 μM). The data demonstrate that the frogs belonging to the family Alytidae are among those producing dermal antimicrobial peptides that may represent a component of the animal's system of innate immunity.