bluuebunny/crossref_metadata_2025 huggingface dataset:
Synthesis of potent agonists of Substance P by replacement of Met11with Glu(OBzl) andN‐terminal glutamine with Glp of theC‐terminal hexapeptide and heptapeptide of Substance P
Authors: STAVROPOULOS GEORGE , KARAGIANNIS KOSTAS , ANAGNOSTIDES STAVROS , MINISTROUSKI ITZHAK , SELINGER ZVI , CHOREV MICHAEL
Published: August 2010

The analogues [Glp6,Glu(OBzl)11]SP(6‐11)and [Glp5,Glu(OBzl)11]SP(5‐11)) of theC‐terminal hexapeptide and heptapeptide of Substance P have been synthesized by conventional solution methods. In each analogue theN‐terminal glutamine has been replaced by pyroglutamic acid, while the COOCH2C6H5ester group has replaced the SCH3group of the Met11side chain. Thein vitroactivity of both analogues has been determined on three biological preparations: guinea pig ileum (GPI), rat vas deferens (RVD) and rat portal vein (RPV). The results showed that both analogues are highly potent and selective agonists on GPI through the NK‐1 receptor. They are more potent than SP itself, with 1.54 and 1.25 respective values of relative potency on GPI. Their selectivity has been studied by utilizing atropine‐treated guinea pig ileum (GPI + At). The analogues showed low activity on RVD and RPV tissues, which represent NK‐2 and NK‐3 monoreceptor assay, respectively. © Munksgaard 1995.
Self‐activation of N‐phosphoamino acids and N‐phosphodipeptides in oligopeptide formation
Authors: ZHAO YUFEN , JU YONG , LI YANMEI , WANG QIAN , YIN YINWU , TAN BO
Published: August 2010

N‐Phosphorylamino acids and N‐phosphoryldipeptides can self‐activate to give the oligopeptide. An intramolecular mixed anhydride intermediate might exist during the self‐activating process. A peptide bond was formed from the N‐terminus to the C‐terminus by the‘1 +n’ mechanism, which was similar to the rRNA‐peptide biosynthesis pathway. © Munksgaard 1995.
Synthesis of a new dipeptide template, its X‐ray structure, and modeling studies on its conformational features
Authors: ESSER FRANZ , CARPY ALAIN , BRIEM HANS , KÖPPEN HERBERT , POOK KARL‐HEINZ
Published: August 2010

The diastereoselective synthesis is reported of a dipeptide template which is closely related to H‐Gly‐Trp‐OH. Intramolecular bond formation between α‐C of Gly and ring position 2 of the Trp unit has been achieved by a Pictet‐Spengler‐type electrophilic aromatic substitution. The absolute configuration of the N‐Moc protected dipeptide template 9·2H2O was determined by single‐crystal X‐ray crystallography and found to be (2S,5S). The cis orientation of the amino and carboxy termini prompted us to investigate the potential of 9 as a β‐turn mimic. MD calculations on the model pseudopeptide Ac‐Ala‐Gly‐Trp‐Ala‐NHMe 11 suggest that an unusually tight turn should be favoured rather than a β‐turn. The proper protective situation as a prerequisite for the incorporation of the template into a peptide has been established, and comments about its chemical properties are given. © Munksgaard 1995.
Effect of adjacent histidine and cysteine residues on the spontaneous degradation of asparaginyl‐ and aspartyl‐containing peptides
Authors: BRENNAN TODD V. , CLARKE STEVEN
Published: August 2010

Aspartate and asparagine residues in polypeptides are subject to nonenzymatic reactions that lead to deamidation, isomerization, peptide bond cleavage and racemization. Much of this reactivity is due to the propensity for the initial formation of a cyclic succinimide intermediate. We have been interested in determining the effect of the side chains of neighboring histidine and cysteine residues in facilitating these reactions, particularly in the possibility that they can act as general acids and bases. In this study, we found little or no effect of histidine residues preceding an asparagine residue in hexapeptides derived from the sequence of adrenocorticotropic hormone, while a histidine residue preceding an aspartic acid residue was found to increase the rate of succinimide formation 8‐ to 11‐fold. The presence of a histidine residue following either an asparagine or aspartic acid residue did not effect the rate of succinimide formation by peptide‐bond nitrogen attack, but did increase the rate of the competing side‐chain nitrogen attack leading to cleavage in the asparaginyl‐containing peptide. We found that the effect of a cysteine residue following an asparagine or aspartic acid residue was in general similar to that of a serine residue, although the cleavage reaction appeared to be enhanced. These results suggest that His‐Asp sequences may be particularly labile to spontaneous degradation in proteins and peptides, possibly owing to the ability of the histidine residue to facilitate succinimide formation by protonating the OH− leaving group on the side chain carboxylic acid of the aspartic acid residue. Finally, we have also utilized these results, along with previously accumulated data on succinimide formation in related peptides, to correlate the rate of succinimide formation with the predicted acidity of the peptide bond nitrogen atom that is involved in the initial nucleophilic attack. © Munksgaard 1995.
Structural and charge requirements for antimicrobial and hemolytic activity in the peptide PKLLETFLSKWIG, corresponding to the hydrophobic region of the antimicrobial protein bovine seminalplasmin
Authors: SITARAM NARASIMHAIAH , SUBBALAKSHMI CHILUKCURI , NAGARAJ RAMAKRISHNAN
Published: August 2010

Several analogs of the 13‐residue antimicrobial and hemolytic peptide PKLLETFLSKWIG (SPF), which is the most hydrophobic region of the 47‐residue antimicrobial protein seminalplasmin [Sitaram, N. & Nagaraj, R. (1990) J. Biol. Chem. 265, 10438‐104423 have been synthesized. The antimicrobial and hemolytic properties of the peptides were investigated with a view to gain an insight into the structural and charge requirements for these activities of SPF. Peptides in which E was replaced by K exhibited considerably improved antimicrobial activity with no concomitant increase in hemolytic activity. A peptide in which the aromatic amino acids were replaced by leucine exhibited antimicrobial activity like those of the peptides which had aromatic amino acids. Interchange in the positions of E and K and total replacement of K by E resulted in complete loss of activity. The peptides having antimicrobial activities showed appreciable helical content in a hydrophobic environment, whereas inactive peptides did not. Thus, by suitable‘engineering’ the biological activity of a short 13‐residue peptide can be altered to yield peptides specifically having only antimicrobial activity with increased potency. © Munksgaard 1995.
Specificity and formation of unusual amino acids of an amide ligation strategy for unprotected peptides
Authors: TAM JAMES P. , RAO CHANG , LIU CHUAN‐FA , SHAO JUN
Published: August 2010

An important step in the recently developed ligation strategy known as domain ligation strategy to link unprotected peptide segments without activation is the ring formation between the C‐terminal ester aldehyde and the N‐terminal amino acid bearing a 8‐thiol or 8‐hydroxide. A new method was developed to define the specificity of this reaction using a dye‐labeled alanyl ester aldehyde to react with libraries of 400 dipeptides which contained all dipeptide combinations of the 20 genetically coded amino acids. Three different ester aldehydes of the dye‐labeled alanine: α‐formylmethyl (FM), β‐formylethyl (FE), and β,β,β‐dimethyl and formylethyl esters (DFE), were examined. The DFE ester was overly hindered and reacted with N‐terminal Cys dipeptides (Cys‐X). Interestingly, it also reacted slowly with the sequences of X‐Gly where Gly was the second amino acid and the X‐Gly amide bond participated in the ring formation. Although the FE ester reacted similarly as the FM ester in the ring formation, the subsequent O,N‐acyl transfer was at least 30‐fold slower than those of the FM‐ester. The FM α‐formyl methyl ester was the most suitable ester and was reactive with dipeptides of six N‐terminal amino acids: Cys, Thr, Trp, Ser, His and Asn. The order and extent of their reactivity were highly dependent on pH, solvent and neighboring participation by the adjacent amino acid. In general, they could be divided into three categories. (1) N‐Terminal Cys and Thr were the most reactive. Cys reacted very rapidly and completely within 0.5 h to form thiazolidine in both aqueous and high content of water‐miscible organic solvents. Thr reacted to form oxazolidine slowly in aqueous buffer (t1/2 > 300 h) but rapidly and completely within 20 h in organic‐water solvents. (2) N‐Terminal Trp, His and Ser were comparatively much less reactive than Cys or Thr. Trp reacted slowly and completely in aqueous buffer but significantly more slowly and incompletely in water‐organic solvents. Both His and Ser reacted very slowly and incompletely in both solvent systems. (3) Finally, Asn reacted nearly insignificantly in both solvent systems. The significant rate enhancement by the water‐miscible organic solvent on Thr was particularly important to allow the synthesis of disulfide‐rich protein domains. Furthermore, the ring formation with N‐terminal Trp, His and Asn provided a convenient route to prepare their bicyclic and unusual heterocyclic derivatives for structure‐activity study.
Synthesis and separation of protected tripeptide epimers by RP‐HPLC
Authors: GRIEHL C. , MERKEL S.
Published: August 2010

Twenty three epimeric pairs of protected tripeptides of the type Z‐Ala‐Xaa‐Val‐OMe have been synthesized by conventional coupling procedures in solution. Optimal conditions for baseline separation of LLL/LDL epimers were found by high‐performance liquid chromatography on a RP‐18 column with aqueous methanol as mobile phase. The hydrophobic parameters (log kw) were determined for all tripeptides.
Synthesis and characterization of the human elastin W4 sequence
Authors: GOWDA D.C. , LUAN C.‐H. , FURNER R.L. , PENG S.Q. , JING N. , HARRIS C.M. , PARKER T.M. , URRY D.W.
Published: August 2010

Following the nomenclature of Sandberg, the W4 sequence of human elastin, LVPGGPGFGPGVVGVPGAGVPGVGVPGAGIPVVPGAGIPGAGVPGVVSPEG, has been synthesized by solid‐phase methods and characterized by carbon‐13 nuclear magnetic resonance, amino‐acid analysis, mass spectra and elemental analysis. This sequence was then polymerized to greater than 50 kDa as determined by retention in 50 kDa molecular weight cut‐off dialysis tubing. It has been successfully cross‐linked by γ‐irradiation (20 Mrad) to form an elastomeric matrix. designated as X20‐poly(W4). Physical characterizations such as stress/strain, thermoelasticity, acid‐base titration and inverse temperature transition studies have been carried out on this elastomer, which is homologous to the striking, poly(VPGVG), W4 sequence of bovine and porcine elastins. These results are compared with previous results on the polypentapeptide of elastin, (VPGVG)n, and it has been demonstrated that X20‐poly(W4) also is a dominantly entropic elastomer. Finally, the working model for the structure of this human elastin sequence was derived computationally using molecular mechanics and dynamics calculations.

Thus the human W4 sequence appears to be structurally and functionally equivalent to the bovine and porcine W4 sequences in spite of the less regular repeating pentamer sequence. © Munksgaard 1995.
Crystal and molecular structure of Boc‐Phe‐Val‐OMe; comparison of the peptide conformation with its dehydro analogue
Authors: RAJASHANKAR K.R. , CHAUHAN V.S , RAMAKUMAR S.
Published: August 2010

The crystal structure of the peptide Boc‐Phe‐Val‐OMe determined by X‐ray diffraction methods is reported in this paper. The crystals grown from aqueous methanol are orthorhombic, space group P 2121 21, a= 11.843(2), b= 21.493(4), c= 26.676(4) A and V= 6790 Å3. Data were collected on a CAD4 diffractometer using MoKα radiation (λ= 0.7107 Å) up to Bragg angle θ=26°. The structure was solved by direct methods and refined by a least‐squares procedure to an R value of 6.8% for 3288 observed reflections. There are three crystal‐lographically independent peptide molecules in the asymmetric unit. All the three molecules exhibit extended conformation. The sidechain of the Val2 residue shows two different conformations. The conformation of the peptide Boc‐Phe‐Val‐OMe is compared with the conformation of Ac‐ΔPhe‐Val‐OH. It is observed that while Boc‐Phe‐Val‐OMe exhibits an extended conformation, Ac‐ΔPhe‐Val‐OH shows a folded conformation. The results of this comparison highlight the conformation constraining property of the ΔPhe residue. Interestingly, even though Boc‐Phe‐Val‐OMe and Ac‐ΔPhe‐Val‐OH are conformation ally different, they exhibit similar packing patterns in the solid state. © Munksgaard 1995.
Isolation and characterization of two distinct lectins with antiproliferative activity from the cultured mycelium of the edible mushroomTricholoma mongolicum
Authors: WANG H.X. , NG T.B. , LIU W.K. , OOU V.E.C. , CHANG S.T.
Published: August 2010

Two lectins, TML‐1 and TML‐2, were isolated fromTricholoma mongolicumby ion‐exchange chromatography and gel filtration. They are dimers with molecular weight near 37 000. The hemagglutinating activities of TML‐1 and TML‐2 are sensitive to lactose inhibition and are stable between 10 and 80 °C. They exhibit antiproliferative activities against mouse monocyte‐macrophage PU5–1.8 cells and mouse mastocytoma P815 cellsin vitro.The two lectins differ in the content of proline and tyrosine residues. Both are non‐glycoproteins and have hydroxyproline residues. © Munksgaard 1995.
Design and synthesis of a β‐strand inducer Application to ICAM‐1/LFA‐1 mediated cellular adhesion
Authors: MICHNE WILLIAM F. , SCHROEDER JOSEPH D.
Published: August 2010

The binding of lymphocyte function associated antigen (LFA‐1) to intercellular adhesion molecule (ICAM‐1) is responsible for several types of cellular adhesion. Amino‐acid substitution mutants of ICAM‐1 have established the importance of several sequences in this protein. We selected the binding region of Glu34 for further study. One published model of domain 1 placed Glu34 near the end of a β‐strand. We designed and synthesized three tripeptide derivatives centered on the Glu34 sequence and attached a platform which, through hydrogen bonds, induces a rigid β‐strand conformation. Variable temperature NMR methods coupled with NOESY 2D NMR data enabled determination of the solution conformation of these compounds. ©Munksgaard 1996.
Proteinase‐catalyzed peptide synthesis in concentrated solutions of urea and other denaturing agents
Authors: ANISIMOVA VERONIKA V. , FILIPPOVA IRINA YU. , LYSOGORSKAYA ELENA N. , OKSENOIT ELENA S. , KOLOBANOVA SVETLANA V. , STEPANOV VALENTIN M.
Published: August 2010

Pepsin successfully catalyzed the synthesis of several hydrophobic octa‐ and decapeptides in dimethylformamide‐water solutions containing concentrated urea at pH 4.65. The factors that influence peptide synthesis in the presence of urea aere studied using condensation of the tripeptides Z‐Ala‐Ala‐Phe‐OH and H‐Leu‐Ala‐Ala‐OCH3 as a model. The dependence of Z‐Ala‐Ala‐Phe‐Leu‐Ala‐Ala‐OCH3 yield on pepsin concentration and pH, as well as the behavior of pepsin during peptide synthesis were studied. It was shown that pepsin catalyzed the synthesis of Z‐Ala‐Ala‐Phe‐Leu‐Ala‐Ala‐OCH3 in guanidine hydro‐chloride and sodium dodecyl sulfate solutions. Other proteinases, subtilisin and thermolysin, were applied for the synthesis of p‐nitroanilides of tri‐ and tetrapeptides in urea solutions.

Proteinase‐catalyzed peptide synthesis in the presence of denaturing agents might help to overcome the limitations caused by poor solubility of the starting peptide derivatives, although this effect is sometimes counterbalanced by the product solubility. ©Munksgaard 1996.
Use of the Hmb backbone‐protecting group in the synthesis of difficult sequences
Authors: SIMMONDS ROBIN G.
Published: August 2010

Aggregation due to hydrogen‐bonded interchain association is thought to be the cause of difficult sequences in solid‐phase peptide synthesis. Hmb (2‐hydroxy‐4‐methoxybenzyl) was introduced recently as a backbone‐protecting group for Fmoc/tBu strategies which inhibits this association. Hmb derivatives of four amino acids are now available commercially. This paper describes the syntheses of two difficult sequences which were achieved by judicious use of the Hmb protecting group. (Hmb)Gly. which is attractive because of the ease of coupling the following residue, did not always produce the expected long‐range effect. Moving the Hmb protection to a suitable preceding position in the sequence overcame this shortcoming. ©Munksgaard 1996.
A convenient method for determining cyclic peptide conformation from 1D 1H‐NMR information
Authors: SEFLER ANDREA M. , LAURI GEORGES , BARTLETT PAUL A.
Published: August 2010

A rapid and convenient method for determining the backbone conformation of cyclic peptides results from the combination of ID 1H NMR information and molecular modeling. Φ, Angle torsional constraints calculated from 3JHN,Hx coupling constants are used to determine the position of multiple‐welled potential energy penalty functions that are imposed on the force field used in the structure refinement (Amber* with GB/SA solvation model). Monte Carlo searches and minimizations lead to a collection of structures that are clustered by backbone similarity and then filtered according to hydrogen‐bonding constraints determined by the chemical shift temperature dependencies of the amide protons. This approach was applied to five cyclic peptides whose structures had been determined previously using more extensive 2D NMR techniques, and the importance of the torsional. H‐bonding, and solvation restraints were assessed. For the four peptides that adopt a predominant conformation, this method reproduced the reported structures closely; lack of convergence for the fifth structure reflected the multiple backbone conformations that this macrocycle adopts. © Munksgaard 1996.
Dissociation, unfolding and inactivation of creatine kinase in urea solutions
Authors: WANG XI‐CHENG , YANG JIAN , ZHOU HAI‐MENG
Published: August 2010

It was previously reported that, during unfolding of creatine kinase in guanidinium chloride or urea solutions, inactivation occured before noticeable conformational change could be detected, suggesting that the conformation at the active site is more easily perturbed and, hence, more flexible than the molecule as a whole [Tsou (1986) Trends Biochem. Sci.11, 417–429]. In the present paper, the urea‐gradient electrophoresis and the isoenzyme hybrid of creatine kinase has been studied. The results show that at low urea concentrations, creatine kinase is still in the dimeric state or only slightly dissociated. The dissociation and inactivation of creatine kinase during denaturation by urea are also compared. It was found that the enzyme was nearly inactivated in low urea concentrations before noticeable dissociation was detected. It therefore appears that in low urea concentrations, inactivation of creatine kinase is not due to the dissociation of the active dimer. The present result supports the hypothesis of the conformational flexibility of the active site in this enzyme. © Munksgaard 1996.
Observation of a mixed antiparallel and parallel β‐sheet motif in the crystal structure of Boc‐Ala‐Ile‐Aib‐OMe
Authors: DATTA S. , SHAMALA N. , GURUNATH R. , BALARAM P.
Published: August 2010

A diastereomeric mixture of the tripeptide Boc‐Ala‐Ile‐Aib‐OMe crystallized in the space group Pl from CH3OH/H2O. The unit cell parameters are a= 10.593(2) Å, b= 14.377(3) Å, c= 17.872(4) Å, α= 104.41(2)°, β= 90.55(2)°, γ= 106.91(2)°, V= 2512.4 Å3, Z=4. X‐Ray crystallographic studies shows the presence of four molecules in the asymmetric unit consisting of two pairs of diastereomeric peptides, Boc‐l‐Ala‐l‐Ile‐Aib‐OMe and Boc‐l‐Ala‐d‐Ile‐Aib‐OMe. The four molecules in the asymmetric unit form a rarely found mixed antiparallel and parallel β‐sheet hydrogen bond motif. The Ala and (l,d)‐Ile residues in all the four molecules adopt the extended conformations, while the φ, ψ values of the Aib residues are in the right‐handed helical region. In one of the molecules the Ile sidechain adopts the unusual gauche conformation about the Cβ‐Cγ bond. © Munksgaard 1996.
Inhibition of prophage induction by synthetic pentapeptide
Authors: ABDEL‐RAHMAN S. , EL‐DIDAMONY G.
Published: August 2010

Six peptide chains different in length and number of sequences were synthesized by the continuous‐flow solid‐phase method. Their effects on prophage induction were tested. It was found that pentapeptide 1H‐Val‐Val‐Asn‐Asp‐Leu‐OH, at a concentration of 25 μ g/mL inhibited the spontaneous induction of prophage Lambda in E. coli strain W 3110(γ) and φ RLZLS in R. Zeguminosarum (lysogenic local isolate) by 99.5% and 65.7%, respectively. Moreover, this concentration was also able to inhibit the replication of the developed phages in the indicator strains. It was suggested that this peptide may block the attachment site of the induced phages or inhibit Rec A Protease. © Munksgaard 1996.
Linear epitopes of sperm whale myoglobin identified by polyclonal antibody screening of random peptide library
Authors: YAO ZHI‐JIAN , CHAN MOEY‐CHU , KAO MANDY C.C. , CHUNG MAXEY C.M.
Published: August 2010

Distinct enhancement of antibody‐specific clones was apparent during the screening against random peptide libraries with antigen‐specific polyclonal antibodies. Several sequence motifs obtained from these screenings were homologous with the primary sequence of myoglobin. Two of these motifs have been confirmed as antigenic determinants by competitive inhibition tests using eight‐branched synthetic peptides. One of the peptides has a sequence that corresponds to amino acid residues 42‐50, KFDRFKHLK, of the myoglobin sequence. This is a new epitope of myoglobin that is reported for the first time. The epitope is located precisely in the ‘turn’ or ‘loop’ region between helices C and D of the crystal structure of myoglobin. The second antibody binding site has a sequence of DIAAKYKELGYQG, and this is located between residues 141‐153, which is the C‐terminus of myoglobin. This epitope encompassed two linear epitopes of myoglobin, amino acid residues 145‐151 and 147‐153, that have been reported earlier based on immunochemical characterisation of cleavage fragments of the protein. These results clearly indicate that epitope mapping using polyclonal antibodies against random peptide libraries can identify new epitopes precisely, as well as confirm epitopes of myoglobin obtained earlier using established methodologies. © Munksgaard 1996.
Binding of human serum amyloid A (hSAA) and its high‐density Iipoprotein3 complex (hSAA‐HDL3) to human neutrophils. Possible implication to the function of a protein of an unknown physiological role
Authors: PRECIADO‐PATT LIANA , PRAS MORDECHAI , FRIDKIN MATI
Published: August 2010

Serum amyloid A (SAA) is an acute‐phase serum protein which exists in the body in a complex with high‐density lipoprotein (HDL3). It is involved in chronic inflammation and neoplastic diseases in an as yet unknown manner. Toward an understanding of the possible physiological role of SAA we initiated a study of its association with blood proinflammatory cells with which it may interact functionally in vivo. In the following we describe the binding characteristics of recombinant human SAA to human neutrophils (polymorphonuclear leukocytes; PMNLs) and their plasma membranes. Scatchard analysis of rSAA binding and displacement curves revealed Kd in the nanomolar range. The C‐terminal domain of the protein, i.e. amino acid residues 77‐104, which might reside in serum following SAA degradation and amyloid A formation, was found to inhibit efficiently the binding of the whole protein to neutrophils. The interaction of SAA, and of its related peptides while complexed in HDL3, with human PMNs was also studied. The results suggest that SAA may be involved, in an as yet unknown manner, in the neutrophil‐associated inflammatory mechanism.
Characterization of PO1, a new peptide ligand of the apamin‐sensitive Ca2+ activated K+ channel
Authors: ZERROUK H. , LARABA‐DJEBARI F. , FREMONT V. , MEKI A. , DARBON H. , MANSUELLE P. , OUGHIDENI R. , RIETSCHOTEN J. VAN , ROCHAT H. , MARTIN‐EAUCLAIRE M.F.
Published: August 2010

A new peptide ligand of the small conductance Ca2+ activated K+ channels has been purified from the venom (obtained by manual rather than electrical stimulation of the scorpion Androctonus mauretanicus mauretanicus), by following the inhibition of the 125I‐apamin binding to its receptor on rat brain synaptosomes. Only one step on a C18 reversed‐phase high‐performance liquid chromatography column was necessary to obtain PO1. Its K0.5 for the apamin binding site was 100 nM. The amino acid sequence of PO1 is different from those of leiurotoxin and PO5. For the first time the same peptide was also purified from the venoms of two other species of North African scorpions, Androctonus austmlis and Buthus occitanus tunetanus. PO1 was chemically synthesized by the solid‐phase technique and fully characterized. A model of PO1 was constructed by amino acid replacement using PO5 nuclear magnetic resonance studies as the starting model. Structure‐activity relationships between these toxins and their receptor are discussed.
Alanine scan and N‐methyl amide derivatives of Ac‐bombesin[7‐14]. Development of a proposed binding conformation at the neuromedin B (NMB) and gastrin releasing peptide (GRP) receptors
Authors: HORWELL DAVID C. , HOWSON WILLIAM , NAYLOR DORICA , OSBORNE SIMON , PINNOCK ROBERT D. , RATCLIFFE GILES S. , SUMAN‐CHAUHAN NIRMALA
Published: August 2010

Alanine and N‐methylation scans together with molecular modelling were implemented in order to propose a binding conformation of the minimum active fragment of bombesin (BB), Ac‐BB[7‐14], to the gastrin releasing peptide (GRP) and neuromedin B (NMB) receptors. These data are also used to critically evaluate the previously proposed binding conformations such as x‐helix and antiparallel β‐sheets. This shows that the previously reported conformations do not satisfy the experimental data. A new binding conformation of Ac‐BB[7‐14] is proposed consisting of three consecutive y‐turns followed by a bend and finishing with two y‐turns. This low energy conformation (analogous to a fragment of thymidylate synthase, 2TSC) of bombesin stabilized by five internal hydrogen bonds, and with the side chains of residues Trp8 and Leu13 held on the same side of the peptide, is in agreement with the experimentally observed data. This and the results of molecular modelling may aid in the synthesis of conformationally restricted high affinity bombesin analogues and/or high affinity template‐based GRP or NMB receptor agonists and antagonists.
Synthesis of novel (N‐farnesyl) amino acids and their incorporation into peptides
Authors: BYK GERARDO , SCHERMAN DANIEL
Published: August 2010

N‐Alkylation, and particularly N‐farnesylation, of bioactive peptides might be of wide interest: first, to increase peptide bioavailability by decreasing their elimination and by favouring their transport through biological membranes; second, to target peptides to cellular membranes; and third, to generate therapeutic double‐substrate inhibitors of enzymes such as ras‐farnesyltransferase. We report the synthesis of novel N‐farnesyl amino acids [(N‐Frn) amino acids]. We have synthesized (N‐Frn)MetOCH3, (N‐Frn)ValOBz and (N‐Frn)PheOCH3, by alkylation of the corresponding natural amino acid esters. In order to demonstrate the feasibility of the introduction of (N‐Frn) amino acids into peptides, we have synthesized representative dipeptide analogs: Cys‐(N‐Frn)ValOBz, Phe‐(N‐Frn)ValOBz, Lys‐(N‐Frn)ValOBz, Phe‐(N‐Frn)MetOCH3, Glu‐(N‐Frn)MetOCH3, Ser‐(N‐Frn)MetOCH3, Trp‐(N‐Frn)PheOCH3, and Pro‐(N‐Frn)PheOCH3, We also describe the synthesis of the model peptide Cys‐Val‐Phe‐(N‐Frn)MetOCH3, which is derived from the tetrapeptide CysValPheMet inhibitor of human p21ras‐farnesyl transferase. © Munksgaard 1996.
Evidence for a C‐terminal turn in PBAN An NMR and distance geometry study
Authors: CLARK BARBARA A. , PRESTWICH GLENN D.
Published: August 2010

The solution conformation of the pheromone biosynthesis activating neuropeptide (PBAN) of the moth Helicoverpa zea has been determined using homonuclear two‐dimensional nuclear magnetic resonance techniques and distance geometry‐restrained energy minimization. The insect peptide hormone showed a random distribution of conformers in aqueous solution, whereas in a less polar medium of trifluoroethanol and water, a reordering process was observed. In particular, the C‐terminal region (Phe‐Ser‐Pro‐Arg‐Leu‐NH2) adopts a type I′β‐turn conformation, residues 20‐27 are in a helix conformation, and residues 1‐19 exhibit a high degree of flexibility. Direct observation of the C‐terminal β‐turn configuration of PBAN is consistent with a previous report that showed a rigid, cyclic analog of the C‐terminal pentapeptide of PBAN retained pheromonotropic activity [Nachman, R.J., Kuniyoshi, H., Roberts, V.A., Holman, G.M. & Suzuki, A. (1993). Biochem. Biophys. Res. Commun. 661‐666], Furthermore, our results show no interaction between the C‐terminal turn and the rest of the polypeptide chain, thus providing further evidence that the C‐terminal turn is indeed the important conformation recognized by the PBAN receptor. © Munksgaard 1996.
Crystal structures of a nonapeptide helix containing α, α‐di‐n‐butylglycine (Dbg), Boc‐G1y‐Dbg‐Ala‐Val‐Ala‐Leu‐Aib‐Val‐Leu‐OMe
Authors: KARLE ISABELLA L. , GURUNATH R. , PRASAD SUDHANAND , RAO R. BALAJI , BALARAM P.
Published: August 2010

Two crystal structures of a nonapeptide (anhydrous and hydrated) containing the amino acid residue α,α‐di‐n‐butylglycyl, reveal a mixed 310‐α‐helical conformation. Residues 1‐7 adopt φ, ψ values in the helical region, with Val(8) being appreciably distorted. The Dbg residue has φ, ψ values of ‐40, ‐37° and ‐46, ‐407° in the two crystals with the two butyl side chains mostly extended in each. Peptide molecules in the crystals pack into helical columns. The crystal parameters are: C50H91N9O12, space groupP21, witha= 9.789(1)Å;,b= 20.240(2) Å.c= 15.998(3) Å.β= 103.97(1):Z= 2,R=10.3% for 1945 data observed < 3σ(F) and C50H91N9O12· 3H2O, space groupP21witha= 9.747(3)Å,b= 21.002(8) Å,c= 15.885(6) Å, β= 102.22(3).Z= 2.R=13.6% for 2535 data observed < 3σ(F) The observation of a helical conformation at Dbg suggests that the higher homologs in the α,α‐dialkylated glycine series also have a tendency to stabilize peptide helices. © Munksgaard 1996.
Photochemical conjugation of peptides to carrier proteins using 1,2,3‐thiadiazole‐4‐carboxylic acid Immunoreactivity of free C‐terminal epitope with specific antibodies
Authors: HANSEN PAUL R. , FLYGE HANNE , HOLM ARNE , LAURITZEN EDGAR , LARSEN BJARNE DUE
Published: August 2010

A new heterobifunctional cross‐linking reagent, 1,2,3‐thiadiazole‐4‐carboxylic acid, for the photochemical conjugation of peptides to proteins is described. The title compound can be coupled directly to a protected peptide resin during solid‐phase peptide synthesis (SPPS) using standard coupling procedures. The probe is stable to TFA deprotection/cleavage mixtures containing ethanedithiol commonly used in Fmoc‐SPPS. Furthermore, tritium may easily be introduced into the thiadiazole ring by base‐catalyzed hydrogen‐exchange. Upon irradiation at 245‐300 nm, parent 1,2,3‐thiadiazole rapidly eliminates N2, generating very reactive thioketene which reacts with amines to give a thioamide in high yield, even when the photolysis is carried out in hydroxylic solvents.

In order to investigate the potential of the title compound as a heterobifunctional cross‐linking reagent a model study with angiotensin II (AII) was conducted. The photoreactive peptide Nα‐4‐carbonyl‐1,2,3‐thiadiazole‐AII (TDA‐AII) was synthesized by Fmoc‐SPPS and conjugated to bovine serum albumin (BSA) by photolysis at 245 and 300 nm. By use of a capture competition ELISA, the C‐terminal Pro‐Phe epitope of photoconjugated AII with the sequence DRVYIHPF was shown to bind specifically to antiAII antibodies (anti‐AII abs), although antibodies against both the C‐ and N‐terminal epitopes were present in the assay. A dipeptide His‐Leu carboxy‐extension form of AII, angiotensin I (AI), only bound to anti‐AII abs at 100‐200 times higher concentrations, showing that the C‐terminal epitope was blocked by the dipeptide. © Munksgaard 1996.
Identification of the residues involved in homodimer formation of recombinant human erythropoietin
Authors: DERBY PATRICIA L. , STRICKLAND THOMAS W. , ROHDE MICHAEL F. , STONEY KENDALL , RUSH ROBERT S.
Published: August 2010

Recombinant human erythropoietin (rHuEPO) is biologically functional when in a monomeric state; upon extensive heating, rHuEPO forms a dimer. The nature of this dimeric linkage was investigated after isolation of the dimer by gel filtration. The dimer fraction was subjected to tryptic digestion, and the peptides were separated by reversed‐phase HPLC. SDS‐PAGE, N‐terminal sequencing, capillary electrophoresis and mass spectrometry (both liquid‐chromatographic electrospray and matrix‐assisted laser desorption ionization) were employed to compare the tryptic peptides from heat‐treated rHuEPO and untreated rHuEPO. Results demonstrated that elevated heat broke the intramolecular disulfide bond between Cys‐7 and Cys‐161 and an intermolecular disulfide bond then formed from these residues, producing a covalently linked rHuEPO homodimer. Dimer formation was also mathematically modeled and shown to fit a simple equilibrium. © Munksgaard 1996.
Design and evaluation of small peptides mapping the exposed surface of IL‐8
Authors: THOMPSON SCOTT K. , VEBER DANIEL F. , JUREWICZ ANTHONY J. , PEISHOFF CATHERINE E. , LEE JUDITHANN M. , WHITE JOHN R.
Published: August 2010

In an effort to determine which regions of IL‐8 are involved in interactions with its receptors, eight peptides were designed to correspond to distinct exposed regions of the IL‐8 monomer, using the proton NMR‐derived structure of the dimer as a basis. The peptides were evaluated singularly, and as equimolar mixtures of two to six peptides, in an IL‐8 receptor binding assay and found to have no binding interaction with either α or β IL‐8 receptor as single peptides or mixtures of two peptides. In contrast, one of these peptides having the sequence AVLPRSAKEL, which corresponds to the N‐terminal 10 amino acid residues of the 77 amino acid form of IL‐8, exhibited potent chemotactic activity in human neutrophils. These results indicate that there is no contiguous ligand that can be designed based on the NMR and X‐ray determined structure of IL‐8 and that there may be multiple receptors responsible for neutrophil activation and chemotaxis.
Structural redesign and stabilization of the overlapping tandem β‐turns of RNA polymerase II
Authors: DOBBINS JOHN R. , MURALI NAGARAJAN , LONG ERIC C.
Published: August 2010

Peptides representing single repeat units of the carboxy‐terminal domain (CTD) of RNA polymerase II (Tyr‐Ser‐Pro‐Thr‐Ser‐Pro‐Ser‐Tyr‐NH2, 1) contain overlapping Ser‐Pro‐Xaa‐Xaa β‐turn forming sites which permit their overall structure to closely resemble members of the quinoxaline class of antitumor DNA bisintercalators. We have modified this native sequence at the i+2 positions of each β‐turn unit by substituting Gly or D‐Ala in an attempt to preorganize this structure in aqueous solution. CD and NMR spectroscopic investigations confirmed the presence of type II β‐turns within each of the substituted peptides in contrast to the native sequence which contains a relatively low population of turn structure. In addition, an examination of singly substituted peptides suggests that an increase in the population of β‐turn structure within the amino‐terminal Ser‐Pro‐Xaa‐Xaa site also increased the formation of β‐turn structure in the carboxy‐terminal (unmodified) Ser‐Pro‐Xaa‐Xaa site; in comparison, substitution in the carboxy‐terminal site did not influence structure in the remaining portion of the peptide. Overall, these results suggest that the structures formed could provide unique. preorganized linkers for the construction of novel DNA‐interactive bisintercalators. © Munksgaard 1996.
Solution structure of the phosphorylated sites of ribosomal protein S6 by 1H NMR spectroscopy
Authors: KATAHIRA RITSUKO , FLOTOW HORST , THOMAS GEORGE , NOSAKA ATSUKO Y.
Published: August 2010

An increase in the rate of protein synthesis is found to be accompanied by phosphorylation of the 40S ribosomal protein S6. Treatment of S6 by cyanogen bromide produced three fragments, and one of the fragments of S6, which is a C‐terminal portion of S6 (Mr≅ 4000), contains all phosphorylation sites of S6. The C‐terminal fragment of S6 contains seven serines. S6 kinase phosphorylates S6 specifically, i.e. five serines in the C‐terminal of S6 are phosphorylated. The three‐dimensional structure of S6 peptide was studied in 50% trifluoroethanol/50% H2O solution by 1H NMR with combined use of distance geometry and restrained molecular dynamics calculations. NMR results indicated that it takes an α‐helix between Glu5 and Arg21 and a distorted helical structure for the following three residues, but no rigid structure was present from Ser25 through the C‐terminus and for the N‐terminal region (Lys1‐Lys4). The specificity of the phosphorylation of the peptide is discussed from a structural aspect. © Munksgaard 1996.
Extrinsic factors derived from mouse embryonal carcinoma cell lines maintain pluripotency of mouse embryonic stem cells through a novel signal pathway
Authors: Kawazoe Shinjirou , Ikeda Nobuhito , Miki Kengo , Shibuya Masayuki , Morikawa Kumi , Nakano Seiji , Oshimura Mitsuo , Hisatome Ichiro , Shirayoshi Yasuaki
Published: August 2010

Embryonic carcinoma (EC) cells, which are malignant stem cells of teratocarcinoma, have numerous morphological and biochemical properties in common with pluripotent stem cells such as embryonic stem (ES) cells. However, three EC cell lines (F9, P19 and PCC3) show different developmental potential and self‐renewal capacity from those of ES cells. All three EC cell lines maintain self‐renewal capacity in serum containing medium without Leukemia Inhibitory factor (LIF) or feeder layer, and show limited differentiation capacity into restricted lineage and cell types. To reveal the underlying mechanism of these characteristics, we took the approach of characterizing extrinsic factors derived from EC cells on the self‐renewal capacity and pluripotency of mouse ES cells. Here we demonstrate that EC cell lines F9 and P19 produce factor(s) maintaining the undifferentiated state of mouse ES cells via an unidentified signal pathway, while P19 and PCC3 cells produce self‐renewal factors of ES cells other than LIF that were able to activate the STAT3 signal; however, inhibition of STAT3 activation with Janus kinase inhibitor shows only partial impairment on the maintenance of the undifferentiated state of ES cells. Thus, these factors present in EC cells‐derived conditioned medium may be responsible for the self‐renewal capacity of EC and ES cells independently of LIF signaling.
Novel interleukin‐4 and interleukin‐1 receptor antagonist gene variations associated with non‐cardia gastric cancer in Japan: Comprehensive analysis of 207 polymorphisms of 11 cytokine genes
Authors: Seno Hiroshi , Satoh Kiichi , Tsuji Shingo , Shiratsuchi Takayuki , Harada Yosuke , Hamajima Nobuyuki , Sugano Kentaro , Kawano Sunao , Chiba Tsutomu
Published: August 2010

Background and Aim:  Helicobacter pylori (H. pylori)‐induced chronic atrophic gastritis is a high‐risk factor for gastric cancer. Immune responses to H. pylori are involved in gastric mucosal inflammation, and might affect clinical outcome, including the development of gastric cancer. The present study examines the significance of gene polymorphisms of various cytokines in the development of gastric cancer following H. pylori infection.

Methods:  One hundred Japanese non‐cardia gastric cancer patients and 93 dyspeptic patients as controls were enrolled in the study (age range 50–75 years). All patients were positive for H. pylori. Genomic DNA was extracted from peripheral whole blood leukocytes, and we comprehensively analyzed 207 single nucleotide polymorphisms (SNP) in 11 cytokine genes; interleukin (IL)‐1α, IL‐1β, IL‐1 receptor antagonist (RN), IL‐4, IL‐4R, IL‐8, IL‐10, IL‐12, TNF‐α, TNF‐β, and IFN‐γ, using either invader assay (163 SNP), direct sequencing (22 SNP), or PCR‐restriction fragment length polymorphism (22 SNP).

Results:  Among the 207 SNP examined, the IL‐4 gene diplotypes (984 and 2983 AA/GA) had a significant negative association with gastric cancer development (odds ratio =0.3, 95% confidence interval =0.1–0.9). When we adopted the dyspeptic patients over 66 years of age as the controls, the IL‐1RN gene diplotypes (−1102 and 6110 CG/GA) also had a significant negative association (odds ratio =0.2, 95% confidence interval =0.1–0.7).

Conclusion:  A comprehensive analysis of 207 SNP of 11 cytokine genes revealed that variations in IL‐4 and IL‐1RN genes are negatively associated with the risk of developing gastric cancer following H. pylori infection. Distinct host cytokine responses in the gastric mucosa might have a role in H. pylori‐induced carcinogenesis.
Effects of recombinant human erythropoietin on echocardiographic findings in elderly dialysis patients
Authors: LIM Paik‐Seong , YEH Chi‐Hsiang , HUNG Tung‐Shiao
Published: August 2010

Summary: Twenty anaemic, normotensive and elderly patients on both haemodialysis and CAPD with left ventricular hypertrophy (LVH) were undergoing systematic echocardiographic analysis during treatment with recombinant human erythropoietin (rHuEPO) for 12 months. Before and after the rHuEPO treatment, M‐mode and pulsed Doppler echocardiographic were performed and haemodynamic parameters were closely followed. Partial correction of anaemia resulted in a decrease in the LV end‐diastolic diameter (LVEDD) from 56.0±2.0–49.2 ± 2.0mm (P<0.05) in HD patients and 50.5 ± 4.0‐43.6 ± 3.0mm in CAPD patients (P= NS). Concomitantly the calculated mass index (LVMi) was reduced significantly from 232.3 ± 21.8‐185.9 ± 11.4 g/m2 in elderly haemodialysis patients. Heart rate and arterial pressure did not change during the study period. However, in CAPD patients there was no significant change in LVMi during the study period. We conclude that long‐term amelioration of anaemia in dialysis patients could induce a regression of left ventricular hypertrophy. However, in CAPD patients, both myocardial and haemodynamic effects were less evident. CAPD could possibly modulate the haemodynamic changes associated with the rHuEPO treatment or CAPD itself impose less left ventricular wall stress in these elderly patients.
Cell kinetics and tissue contraction following renal parenchymal cell death
Authors: KELYNACK Kristen J , HEWITSON Tim D , NICHOLLS Kathy M , BECKER Gavin J , JONES Colin L
Published: August 2010

SUMMARY: The kinetics of interstitial extracellular matrix remodelling in the kidney following parenchymal cell death were studied after injury was produced by a single application of liquid nitrogen to the surface of the kidney. the size of the injury decreased to 30% of the original size at 4 days and to 5% at 16 days after injury. the primary response to injury consisted of an acute polymorphonuclear infiltration followed by monocyte/macrophage and myofibroblast accumulation from day 2 to 8. Collagen III accumulation increased progressively until day 16 and was inversely related to lesion size. These results demonstrate the time course of cellular events and collagen synthesis in renal tissue remodelling following renal parenchymal cell death.
11β hydroxysteroid dehydrogenase type 2 enzyme is expressed in normotensive and hypertensive patients with renal disease
Authors: STEIN‐OAKLEY Alicia N , MAGUIRE Julie A , DOWLING John , PERRY Greg J , KROZOWSKI Zygmunt , THOMSON Napier M
Published: August 2010

SUMMARY: The enzyme 11β hydroxysteroid dehydrogenase type 2 (11βHSD2) converts glucocorticoids to their 11‐keto metabolites, permitting binding of aldosterone to the non‐selective mineralocorticoid receptor. Recent studies have suggested that 11βHSD deficiency may be implicated in essential hypertension and renal disease. Using an antibody against a peptide deduced from the cDNA sequence, the renal expression of 11βHSD2 was examined in patients with advanced stages of renal disease and associated hypertension, patients with primary hypertension resulting in renal damage, and patients with preserved or mildly damaged renal architecture without concomitant hypertension. Despite variable degrees of tubulointerstitial damage, 11βHSD2 was expressed in distal convoluted tubules and collecting ducts in all patient groups. Collapsed tubules retained strong immunoreactivity, but decreased staining was apparent in dilated tubules. There was weak staining of the thick ascending limb of Henle. Variable glomerular expression of 11βHSD2 was observed. Our results therefore suggest that absence of renal 11βHSD2 is not a prominent feature of hypertensive nephrosclerosis, or primary renal disease associated with hypertension.
Lipid risk factors for atherosclerotic carotid artery disease in patients on continuous ambulatory peritoneal dialysis
Authors: YORIOKA Noriaki , TANIGUCHI Yoshihiko , ITO Takafumi , SHAO Jin Chang , MASAKI Takao , YAMASHITA Kazuomi , YAMAMOTO Maki , YAMAKIDO Michio
Published: August 2010

SUMMARY: We investigated the relationship between abnormalities of lipid metabolism and carotid atherosclerosis in 20 patients on continuous ambulatory peritoneal dialysis (11 men and nine women). Carotid ultrasonography was used to determine the combined thickness of the intima and media (I‐M thickness). the apo A‐I/apo B ratio showed a significant negative correlation with I‐M thickness (P<0.05). In the patients with carotid plaque, the triglycerides (TG) level and the remnant‐like particle cholesterol level (RLP‐C) were significantly higher than in the patients without plaque (P<0.05), and high density lipoprotein (HDL) cholesterol and the apo AI/apo B ratio were significantly lower than in the patients without plaque (P<0.05). Moreover, there was a strong relationship between the severity of plaque and TG, the apo AI/apo B ratio, and RLP‐C. Thus, abnormal lipid metabolism may contribute to progressive atherosclerosis, while TG, the apo AI/apo B ratio, and RLP‐C. levels may be useful indicators of atherosclerotic risk in peritoneal dialysis patients.
Non-Voluntary Return? The Politics of Return to Afghanistan
Authors: Blitz Brad K. , Sales Rosemary , Marzano Lisa
Published: August 2010

The forced removal of 35 Afghan nationals from the UK in April 2003 calls into question the viability of the government's voluntary repatriation schemes and undermines the voluntary nature of return programmes. This article draws on the results of research conducted in 2002 to explore the views of the Afghan community about return. We evaluate three motivations for promoting return programmes: justice-based arguments, where return is the ‘end of the refugee cycle’; human capital explanations, which focus on individual decisions to reverse the effects of brain-drain; and burden-relieving explanations, where return is an alternative to repatriation. Our findings suggest that domestic interest based arguments, rather than those founded on the protection of human rights, are driving the policy-making agenda. Returns are portrayed as a means of relieving the burden on welfare services, and placating an increasingly anti-immigrant public opinion. As well as individuals forcibly removed from Britain, other Afghans are being urged to return by means of financial inducements, and sometimes under the threat of repatriation. In this context, we can discern a new category of ‘non-voluntary’ returns where individual choice has little real meaning.