bluuebunny/crossref_metadata_2025 huggingface dataset:
Regiospecific arylazo substitution into α‐amanitin with retention of inhibitory properties against eukaryotic Class II RNA polymerase
Authors: FALCK‐PEDERSEN ERIK , MORRIS PAUL W. , VENTON DUANE L.
Published: August 2010

Treatment of α‐amanitin with 1,2 equiv. benzenediazonium acetate in methanolic NaOH affords the crude 7'‐phenylazo‐α‐amanitin in 92% yield. The azoamatoxin is isolable in yields of 50% following reverse phase partition chromatography and shows the extended conjugation of an azo dye (Λmax= 295 and 384 nm in CH3OH) when compared to the parent α‐amanitin (Λmax= 305 nm). Spectrophotometric titration of the amatoxin phenolic hydroxyl shows a decrease in pKAto 7.75 as expected following azo substitution. Proton magnetic resonance at 60 MHz shows 7 aromatic protons; two AB coupled protons (δ4H= 7.83 p.p.m.; δ5H= 6.80 p.p.m., J4,5= 9) are assigned to the remaining two protons of the amatoxin indole. The 5 phenyl protons constitute a second order spin system with δ3,4,5H= 7.44–7.53 p.p.m. and δ2,6H= 7.97–8.11 p.p.m. The 7'‐phenylazo‐α‐amanitin inhibits sea urchin RNA polymerase with apparent Kis at 1.4 times 10‐8M (class II) and 1.3 times 10‐5M (class III). The Kifor the urchin class III RNA polymerase is decreased 10‐fold from the value observed with the parent α‐amanitin (Ki= 1.4 times 10‐4M), while there is no significant change for the class II enzyme. No inhibition of the class I orE. coliRNA polymerase is observed at 10‐4M. Extension of this diazo coupling reaction using other arylamines affords the preparation of a family of 7'‐arylazo‐α‐amanitins in good yield analogous to the benzenediazonium reaction. The title reaction thus provides a second facile, efficient route to modified amatoxins of defined structure with retention or enhancement of inhibitory activity against the eukaryotic RNA polymerase.
Solution conformation of poly(L‐lysyl‐L‐glutamic acid) and poly(L‐lysyl‐L‐glutamine)
Authors: RAO M.V.R. , ATREYI M. , CHAUHAN V.S. , KUMAR SATISH
Published: August 2010

Solution conformation of poly(L‐lysyl‐L‐glutamic acid) (PLGU) and poly(L‐lysyl‐L‐glutamine) (PLGN) was studied in water as a function of pH, added salt, detergents, methanol and trifluoroethanol (TFE). Both the polypeptides exhibit no ordered conformation in the pH range 1.5–12.5; salts and detergents did not have any marked effect. Replacement of side chain carboxyl by an amide group did not help in inducing PLGN to adopt a helical conformation even at pH as high as 12.0, unlike poly(L‐lysine). The helicogenic solvents, methanol and TFE, induce formation of weak helices in PLGU as well as in PLGN. It is not unlikely that H‐bonding between the side chains leads to stabilizing an unordered conformations.
Synthesis of tritiated tuftsin and macrophage inhibitory tripeptide via acetylenic intermediates
Authors: AUGER GENEVIÈVE , BLANOT DIDIER
Published: August 2010

Acetylenic analogues of tuftsin (Thr‐Dah‐Pro‐Arg) and of a macrophage inhibitory tripeptide (Thr‐Dah‐Pro) have been synthesized by conventional procedures in solution (Dah = 2,6‐diamino‐4‐hexynoic acid). These acetylenic derivatives are intermediates for the preparation of structurally unmodified, tritiated peptides. Catalytic tritiation of Thr‐Dah‐Pro‐Arg and of Thr‐Dah‐Pro has afforded the radioactive tetra‐ and tripeptides with specific activities of 11.4 Ci/mmol and 37 Ci/mmol, respectively.
Cold acclimation increases physiological responsiveness to intraventricular injection of β‐endorphin in pentobarbital anaesthetized rats
Authors: WONG T.M. , TSE S.Y.H.
Published: August 2010

β‐endorphin administered intraventricularly into pentobarbital anaesthetized rats led to a reduction in rectal temperature and a dose dependent biphasic response in metabolic rate with increase at low doses (7–10 nmol/kg) and decrease at high doses (30–300 nmol/kg) in rats acclimated to 22°. These metabolic responses to β‐endorphin were enhanced both in magnitude and in duration in rats acclimated to 5° for 3 weeks, indicating an increased responsiveness to β‐endorphin. Naloxone antagonized the metabolic effects of β‐endorphin in rats acclimated to both 22° and 5°, indicating the metabolic effects of β‐endorphin were via the naloxone sensitive opiate receptors.
Structure and conformation of linear peptides
Authors: LALITHA V. , SUBRAMANIAN E. , BORDNER J.
Published: August 2010

The tripeptide, glycyl‐glycyl‐L‐isoleucine, crystallizes as a monohydrate in the monoclinic space group P21, with a = 12.746(2), b = 6.172(1), c = 8.643(1) Å, β = 99.77(2)°, and Z = 2. The structure was solved by direct methods and refined to an R‐value of 0.039 for 917 (I > 1°) reflections. The molecule exists as a zwitterion in the crystal. The peptide units are trans and show significant deviations from planarity. The plane of peptide units and of the carboxyl group are nearly mutually perpendicular to each other. The peptide backbone torsion angles are: ø1 = ‐ 171.2°, ω1 = ‐ 176.8°, ø2 = ‐ 106.1°, ø2 = ‐ 150.7°, ø2 = ‐ 172.1°, ø3 = ‐ 70.9°, ø3 = 136.5°. For the side‐chain of isoleucine, ø1 = ‐ 58.1°, ø2 = 169.7° and the system of bonds C′‐Cα‐Cβ‐C γ1‐Cδ is trans zig‐zag. The packing arrangement involves spatial segregation of polar and nonpolar moieties.
A potential function for conformational analysis of proteins
Authors: CRIPPEN G.M. , VISWANADHAN V.N.
Published: August 2010

We have devised a residue‐residue potential function for low resolution protein conformational calculations. The interactions between residues near in sequence maintain correct secondary structure, while the long‐range terms in the potential govern the larger packing features and overall globularity. The short‐range terms were calculated by comparing the observed distributions of distances between Cα atoms in 35 protein crystal structures to the expected distributions and assigning the discrepancies to a Boltzmann distribution due to an effective potential. Long‐range terms were adjusted to ensure that the crystal structure of bovine pancreatic trypsin inhibitor has a lower total energy than perturbed conformations of the same molecule. Thus the empirical potential function implicity contains solvation and conformational entropy effects along with the usual Van der Waals and electrostatic energies. Extensive testing of the potential on trypsin inhibitor and other proteins establishes that it is generally applicable to small proteins, it does not attempt to compress or expand the conformations found by X‐ray crystallography, standard secondary structural features are maintained under the potential, and there are so many local minima that local minimization can be trusted to return a perturbed structure to the native conformation only if they differ initially by less then 1 Ad̀.
Primary structure of osteocalcin from ovine bone
Authors: MENDE LIANE M. , HUQ N. LAILA , MATTHEWS HARRY R. , CHAPMAN GEORGE E.
Published: August 2010

Osteocalcin was isolated from ovine (sheep) bone and purified by chromatography. The primary structure was determined using a manual micro‐DABITC/PITC double coupling method. Ovine osteocalcin contains two modified amino acids, 4‐hydroxyproline and γ‐carboxyglutamate. These residues were readily identified in the sequencing process and the data presented here extend the micro manual sequencing method to include identification of 4‐hydroxyproline and γ‐carboxyglutamate residues. Ovine osteocalcin consists of 49 amino acid residues and the sequence confirms the high degree of conservation previously observed between bovine and human osteocalcins.
Regeneration of native structure and biological activity by air oxidation of the reduced bovine trypsin inhibitor (Kunitz)*
Authors: BIONDI LAURA , FILIPPI BRUNO , FILIRA FERNANDO , TOLAZZI MARILENA , ROCCHI RANIERO
Published: August 2010

Derivatives of the tetraguanidinated bovine trypsin‐kallikrein inhibitor lacking up to four amino‐acid residues have been prepared by Edman degradation of the amino‐terminal Arg‐Pro‐Asp‐Phe sequence. Comparative studies on the native inhibitor, tetraguanidinated inhibitor, and truncated sequences demonstrated that none of the four N‐terminal amino‐acid residues are essential for the regeneration of native structure and biological activity by air oxidation of the reduced molecule.

However residues 2, 3, and 4, although to a different extent, are important for the protein stability and significantly affect the conformation of the tetraguanidinated inhibitor and the rate of the reactivation process.

Removal of the terminal positive charge from Arg 1 by diazotization gave a fully active deaminated, tetraguanidinated inhibitor but the lack of the salt bridge interaction between the amino and the carboxyl terminus of the protein prevented the correct folding of the reduced molecule by air oxidation as shown by activity measurements and conformational studies.

The stability constants and the standard free energies of binding of the complexes between trypsin and the different inhibitor derivatives have been determined.
Fast atom bombardment mass spectrometry of ceruletide and [Tyr 4] ceruletide
Authors: ARLANDINI EMANUELE , GIOIA BRUNO , PERSEO GIUSEPPE , VIGEVANI ARISTIDE
Published: August 2010

Fast atom bombardment (FAB) mass spectrometry has revealed a very useful technique in obtaining the mass spectrum of ceruletide (formerly caerulein), a decapeptide with a tyrosyl‐O‐sulphate residue, and of its desulphated analog. Molecular weight value for ceruletide is obtained only in the negative ion mode, whereas for [Tyr4] ceruletide this information is obtained both in negative and in positive ion detection. Positive ion mass spectra show more extended fragmentation which allows the complete amino acids sequence to be checked. The lability of the sulphate group has been confirmed, examining other sulphated peptides by field desorption mass spectrometry.
Guanidinium‐carboxylate interaction: methylguanidinium formate
Authors: BRAY D.D. , SLATTERY N. , RUSSELL C.S.
Published: August 2010

The crystal structure of methylguanidinium formate is presented as a model for arginine ‐ carboxylate interactions. The crystals are orthorhombic, P212121, with a =6.963(2), b = 7.110(2), c = 11.873(3)Å. The structure was solved by direct methods and refined by block‐diagonal least squares to R = 0.078 and Rw = 0.080. Ion pairs are formed by hydrogen bonds between the amino and CH3 NH‐ imino groups and the formate ion. Additional hydrogen bonding results in the formation of a three‐dimensional network. Comparisons are made with similar compounds.
Acylated arachins.
Authors: Shyamasundar R. , Rao .D. Rajagopal
Published: August 2010

Acetylated and succinylated arachins were examined for various physico‐chemical parameters such as viscosity, absorption and fluorescence spectra, gel electrophoretic patterns, gel filtration profiles and susceptibility to proteolysis. Acetylation and succinylation lead to dissociation of arachin into lower molecular weight fractions. The lower molecular weight fractions had a higher degree of succinylation compared to other fractions. The absorption and the fluorescence spectra were perturbed both by acetylation and by succinylation. Both fluorescence and viscosity data indicate denaturation of acylated arachins. The modified arachin was more susceptible to proteolysis by trypsin and chymotrypsin and varied in degree of hydrolysis.
Semisynthetic studies on bovine pancreatic ribonuclease
Authors: Di Bello Carlo , Lucchiari Adriana , Buso Orfeo , Tonellato Mauro
Published: August 2010

The S‐peptide of the enzyme bovine pancreatic ribonuclease has been used as a model for covalent semisynthesis. Methods for side‐chain protection, enzymatic cleavage of the peptide chain at the level of the single arginine‐10 and for selective deprotection of the α‐carboxyl function of this residue, have been examined. The partially protected [1–10] sequence has been coupled to a solid‐phase generated [11–15] sequence attached to the polymer. After deblocking from the solid‐support, the [1–15] semisynthetic peptide was complexed with native S‐protein to give a complex with high biological activity.
Conformational modifications of cyclic hexapeptide somatostatin analogs
Authors: Freidinger Roger M. , Perlow Debra Schwenk , Randall William C. , Saperstein Richard , Arison Byron H. , Veber Daniel F.
Published: August 2010

A model for the bioactive conformation of the highly active cyclic hexapeptide somatostatin analog cyclo‐(Pro‐Phe‐d‐Trp‐Lys‐Thr‐Phe) has been proposed. As a test of this model, several compounds containing lactam and N‐Me amino acid conformational modifications in the Thr‐Phe‐Pro‐Phe beta turn were synthesized. The N‐Me alanine and sarcosine substitutions for proline gave highly active analogs, while lactam dipeptides in place of Phe‐Pro decreased potency. 1H n.m.r. and CD spectra of these analogs illustrate the conformational effects in solution of these modifications. The results provide additional support for the proposed conformational model.
Antiparallel β‐sheets in the crystal structure of the heptapeptide Met‐Glu‐His‐Phe‐Arg‐Trp‐Gly (ACTH 4–10)
Authors: Admiraal G. , Vos Aafje
Published: August 2010

The conformation of the molecules in ACTH 4–10 has been determined as part of a study of the conformations of the biologically active N‐terminal fragments of the adrenocorticotropic hormone (ACTH). ACTH 4–10 crystallizes in two different superstructures. The substructure considered in the present work, is monoclinic, space group C2, a = 25.75(1) Å, b = 27.78(1) Å, c = 20.35(1) Å, β= 114.0(1)°, Z = 8 molecules ACTH 4–10 plus 22 weight per cent solvent. The crystals contain antiparallel β‐sheets, the orientations of the side groups are not found, because of disorder. The present crystal structure and those of other linear oligopeptides emphasize that antiparallel β‐sheets are energetically favourable. It is very unlikely, however, that the ACTH 4–10 crystals contain the molecules in their biologically active form.
Studies on peptides.
Authors: Kiyama Shinya , Fujii Nobutaka , Yajima Haruaki , Moriga Motoyuki , Takagi Atsushi
Published: August 2010

The heptacosapeptide amide corresponding to the entire amino acid sequence of chicken secretin was synthesized by assembling four peptide fragments followed by deprotection with thioanisole‐mediated trifluoromethanesulfonic acid in TFA. The deprotected peptide was purified by gel‐filtration on Sephadex G‐25, followed by ion‐exchange chromatographies on CM‐cellulose and DEAE‐Sepha‐rose. Synthetic peptide stimulated the flow of pancreatic alkaline juice and decreased systemic blood pressure in rats.
Synthesis and immunological characterization of two peptides which are models for two of the four major antigenic sites of a scorpion toxin
Authors: Granier C. , Bahraoui E. , Van Rietschoten J. , Rochat H. , Ayeb M. El
Published: August 2010

We have synthesized, by the solid phase procedure, then purified and chemically characterized two peptides. They mimic two regions of toxin II of the scorpion Androctonus australis Hector, one around disulfide bridge 12–63 and another at sequence 50–59. Each of these two regions was supposed to include an antigenic site. We have shown that the synthetic replicas of these regions are individually recognized by a part of IgGs raised against native toxin II. This is a strong argument for the involvement of these areas in the antigenicity of the toxin. Furthermore, region‐specific IgGs purified by affinity chromatography on the two Sepharose linked peptides were able to bind 125I‐labelled toxin II.
Methylthiomethyl, a new carboxyl protecting group in peptide synthesis
Authors: Dossena Arnaldo , Palla Gerardo , Marchelli Rosangela , Lodi Tiziana
Published: August 2010

Nα‐protected amino acid methylthiomethyl esters (MTM) were obtained in good yields under mild conditions using the «ButBr/Me2SO» reagent. Selective removal of the N‐protecting group was achieved in HCl/anhydrous ethyl ether and the MTM ester hydrochlorides were successfully used in the synthesis of dipeptides.
N‐acyl‐diketopiperazines.
Authors: Cerrini Silvio , Fedeli Walter , Lucente Gino , Mazza Fernando , Pinnen Francesco , Zanotti GIancarlo
Published: August 2010

N‐(N‐phenylacetyl‐L‐alanyl)‐cyclo‐(L‐phenylalanyl‐D‐prolyl) (I) was synthesized in one step starting from the linear precursor phenylacetyl‐L‐alanyl‐L‐phenyl‐alanyl‐L‐proline. X‐ray crystallographic analysis of (I) shows that the diketopiperazine ring adopts a boat conformation appreciably more puckered than that found in the unacylated cyclo(L‐Pro‐D‐Phe). The side chain of Phe residue is in quasi‐axial flagpole orientation with the aromatic ring folded over the diketopiperazine ring. 1H‐n.m.r. data indicate that the same conformation is preferred in chloroform solution. The proline ring assumes a β‐envelope conformation. No intramolecular interactions between the diketopiperazine system and the aromatic ring of the N‐phenylacetyl‐alanyl side chain have been evidenced. Crystals: space group P21 with a= 9.956(3), b= 8.809(2), c= 13.615(2) Å, β= 111.0(1)° and Z = 2. The final R and Rw are 0.037 and 0.052, respectively.
Conformational properties of aminosuccinyl peptides.
Authors: Capasso Sante , Mattia Carlo Andrea , Mazzarella Prof.Lelio , Zagari Adriana
Published: August 2010

The synthesis and crystal structure of Boc‐L‐aminosuccinyl‐glycine methyl ester is reported. The crystal structure was solved by direct methods and refined to a final R value of 0.064 for the 761 observed reflections (I ≥ 2.5 σ(I)). The molecule is highly extended and the succinimide ring adopts a puckered conformation intermediate between that of a “pure envelope” and a “pure twist” conformation. The molecule was also studied by empirical potential energy calculations. One of the minima of the potential energy map closely corresponds to a type II‘β reverse turn. The results are discussed in connection with the stereochemical constraints imposed by the presence of a cyclic imide structure to the conformational freedom of a polypeptide chain.
Cold acclimation increases physiological responsiveness to intraventricular injection of β‐endorphin in pentobarbital anaesthetized rats. 1. Cardiovascular functions
Authors: Tse S.Y.H. , Wong T.M.
Published: August 2010

Intraventricular injection of β‐endorphin (3, 7, 10 and 30 nmol/kg) into the third ventricular of pentobarbital‐anaesthetized male Sprague‐Dawley rats resulted in a dose‐dependent increase in mean arterial pressure (MAP) while injection of the same volume of 0.9% NaC1 solution did not cause significant changes in MAP. Naloxone, which did not produce any significant change in MAP, antagonized the vasopressor effect of β‐endorphin, indicating that the response is mediated via the naloxone sensitive opiate receptors. Rats acclimated to cold (5°) for 3 weeks showed a potentiated and prolonged increase in MAP following β‐endorphin injection, indicating an increased responsiveness to the peptide. This increased responsiveness in the cardiovascular system is probably of adaptive value in cold acclimation. Naloxone itself did not alter MAP either, but abolished the cardiovascular response to β‐endorphin completely in cold acclimated rats, indicating an increased effectiveness in its antagonistic effect following cold acclimation as well.
4‐Sulfobenzyl ester – an anionic protecting group for peptide synthesis
Authors: Bindewald Roland , Hubbuch Arnulf , Danho Waleed , Büllesbach Erika E. , Föhles Josef , Zahn Helmut
Published: August 2010

The preparation of the 4‐sulfobenzyl esters of 18 amino acid derivatives is described. This carboxyl protecting group was introduced according to Hubbuchet al.(1980). The caesium or dicyclohexylammonium salts ofN‐terminal protected amino acids were reacted with 4‐(bromomethyl)benzenesulfonate (1). AfterN‐terminal deblocking, the amino acid‐4‐sulfobenzyl esters were isolated as zwitterions. The protecting group was removable by catalytic hydrogenation and by saponification. The 4‐sulfobenzyl esters could be easily converted to amides and hydrazides. They were stable to 2 M hydrogen bromide in acetic acid as well as to a 10‐fold excess of trifluoromethane sulfonic acid in trifluoro‐acetic acid. The behaviours of+H2‐Gly‐Phe‐Leu‐OBzl‐SO‐3and the corresponding methyl, benzyl and 4‐nitrobenzyl esters were compared under various conditions.
Nature of the B10 amino acid residue.
Authors: Burke G. Thompson , Schwartz Gerald , Katsoyannis Panayotis G.
Published: August 2010

Human [10‐asparagine‐B] insulin ([Asn10‐B] insulin), an analogue which differs from the parent molecule in that the histidine residue at position 10 of the B chain (B10) is replaced by asparagine, has been synthesized and isolated in purified form. In vitro biological assays indicated a potency of ca. 35% compared to insulin. We have previously shown that the replacement of histidine at position B10 by lysine resulted in an analogue displaying ca. 15% of the biological activity of natural hormone, while the substitution of leucine in this position produced a molecule exhibiting ca. 45% potency in in vivo assays. The data indicate that molecular size of the amino acid residue at position B10 may be important in the maintenance of a structure commensurate with high biological activity. Polarity at this position appears to be rather unimportant while a strongly basic group appears to be deleterious.
Modified calmodulin calcium binding domain III.
Authors: Pavone Vincenzo , Di Nola Angela , Andini Salvatore , Ferrara Luciano , Di Blasio Benedetto , Benedetti Ettore , Pucci Piero
Published: August 2010

The dodecapeptide Ac‐Asp‐Lys‐Asp‐Gly‐Asn‐Gly‐Tyr‐Ile‐Ser‐Ala‐Ala‐Gaba‐OH is a modified calmodulin calcium binding domain III. The synthesis of the peptide by the solid phase method with a total protection scheme using PAM‐resin is reported. The purified compound has been characterized by 1H n.m.r. spectroscopy, both in the presence and in the absence of calcium ions, at various pHs. No strong specific interaction seems to occur between the peptide and Ca++ ions in water solutions.
1H n.m.r. investigation of conformational features of cyclic enkephalinamide analogs
Authors: Mosberg Henry I. , Schiller Peter W.
Published: August 2010

The conformational basis for the differing opioid receptor selectivities of the cyclic cystine‐containing analogs, [D‐Cys2, D(or L)‐Cys5] enkephalinamide and the related penicillamine‐containing analogs, [D‐Pen2, D(or L)‐Cys5] enkephalinamide (penicillamine =β,β dimethylcysteine) was investigated by 1H n.m.r. in aqueous solution. Comparison of chemical shift, temperature dependence of amide proton chemical shift, and coupling constant data suggests similar overall conformations for corresponding penicillamine‐ and cystine‐containing analogs. Differences in conformation and flexibility do appear in the carboxamide terminal region of the corresponding analogs, which may account for their selectivities for different classes of opioid receptors.
Isolation and characterization of monodeamidated derivatives of bovine pancreatic Ribonuclease A
Authors: Venkatesh Yeldur P. , Vithayathil Paul J.
Published: August 2010

The isolation and characterization of the initial intermediates formed during the irreversible acid denaturation of enzyme Ribonuclease A are described. The products obtained when RNase A is maintained in 0.5 M HCl at 30° for periods up to 20 h have been analyzed by ion‐exchange chromatography on Amberlite XE‐64. Four distinct components were found to elute earlier to RNase A; these have been designated RNase Aa2, Aa1c, Aa1b, and Aa1a in order of their elution. With the exception of RNase Aa2, the other components are nearly as active as RNase A. Polyacrylamide gel electrophoresis at near‐neutral pH indicated that RNase Aa1a, Aa1b, and Aa1c are monodeamidated derivatives of RNase A; RNase Aa2 contains, in addition, a small amount of a dideamidated component. RNase Aa2, which has 75% enzymic activity as compared to RNase A, consists of dideamidated and higher deamidated derivatives of RNase A. Except for differences in the proteolytic susceptibilities at an elevated temperature or acidic pH, the monodeamidated derivatives were found to have very nearly the same enzymic activity and the compact folded structure as the native enzyme. Fingerprint analyses of the tryptic peptides of monodeamidated derivatives have shown that the deamidations are restricted to an amide cluster in the region 67–74 of the polypeptide chain. The initial acid‐catalyzed deamidation occurs in and around the 65–72 disulfide loop giving rise to at least three distinct monodeamidated derivatives of RNase A without an appreciable change in the catalytic activity and conformation of the ribonuclease molecule. Significance of this specific deamidation occurring in highly acidic conditions, and the biological implications of the physiological deamidation reactions of proteins are discussed.
Analogs of viroidin.
Authors: Kahl Jens Uwe , Vlasov Genadi P. , Seeliger Annemarie , Wieland Theodor
Published: August 2010

The vitroxins, toxic cyclic heptapeptides from Amanita virosa fungi, contain a 3,4‐dihydroxy‐L‐proline, an imino acid scarcely accessible in greater amounts. Therefore in the syntheses of the analogs described this imino acid was replaced by 4‐cis‐hydroxy‐L‐proline, a component of the analogously toxic phallotoxins. The following syntheses are described: 1a= cyclo (L‐alanyl‐D‐threonyl‐D‐seryl‐L‐4‐cis‐hydroxy‐L‐prolyl‐L‐alanyl‐2‐methylthio‐L‐tryptophyl‐L‐leucyl), 1b=1a but with 2‐methylsulfonyl‐L‐tryptophan in position 6, 2a= cyclic (L‐valyl‐D‐threonyl‐D‐seryl‐4‐cis‐hydroxy‐L‐prolyl‐L‐alanyl‐2‐methylthio‐L‐tryptophyl‐hydroxyl‐L‐leucyl), and 2b=2a, but with 2‐methylsulfonyl‐L‐tryptophan in position 6. In a test for binding strength to rabbit muscle F‐actin 2b and 2a exhibited about 40% of that of demethylphalloin. Analogs 1a and 2b have not been tested.
pH‐dependent changes in properties of concanavalin A in the acid pH range
Authors: AUER HENRY E. , SCHILZ TIMOTHY
Published: August 2010

Properties characteristic of the structure and function of dimeric concanavalin A have been studied as a function of pH in the acid pH range using preparations comprising intact subunits or enriched in fragmented chains. For intact subunits, the glycogen binding ability falls to zero with a midpoint at pH 4.7, the release of Mn+2, Ca+2 and the fluorescent ligand 4‐methylumbelliferyl‐α‐D‐mannopyranoside from the lectin coincides over a pH range centered at pH 3.9, and the CD spectra of the aromatic amino acid residues increase sharply in amplitude between pH 4.0 and 1.5. Nevertheless, the sedimentation coefficient and peptide CD spectrum change insignificantly in the pH range 5 to 2, indicating that dimeric concanavalin A retains its secondary structure and overall hydrodynamic shape essentially unchanged upon acidification. The behavior of concanavalin comprising primarily fragmented chains is not significantly different from that of intact subunits, although it precipitates glycogen less efficiently. It is concluded that dimeric concanavalin A does not undergo a concerted change in structure upon acidification, but rather that it passes through a series of states differing from one another in their local conformations. The distinction in binding between the monosaccharide and the polysaccharide is attributed to participation of a secondary binding site in the latter case. A change in optical activity at 283 nm in the pH range 5–6 is ascribed to disruption of intersubunit interactions of Tyr 67 as the protein undergoes the dimer‐tetramer equilibrium.
Synthesis of human pancreatic growth hormone‐releasing factor and two omission analogs by segment‐coupling method in aqueous solution
Authors: BLAKE JAMES , WESTPHAL MANFRED , LI CHOH HAO
Published: August 2010

The human growth hormone‐releasing factor (GRF) peptides [GlyS15]‐GRF‐(1–15) (IV), trifluoroacetyl‐GRF‐(20–44) (VI), trifluoroacetyl‐GRF‐(18–44) (VIII), and trifluoroacetyl‐GRF‐(16–44) (X) were synthesized by the solid‐phase method. Each of the peptides was reacted with citraconic anhydride and the trifluoroacetyl group was removed by reaction with 10% hydrazine in water. The citraconylated GRF‐(1–15) peptide was coupled to the (20–44), (18–44) or (16–44) peptides by reaction with silver nitrate/N‐hydroxysuccinimide to give GRF‐(1–15)‐(20–44) (XII), GRF‐(1–15)‐(18–44) (XIII), or GRF‐(1–44), respectively. GRF‐(1–44) was shown to stimulate the release of rat growth hormone from rat pituitary cells with an ED50= 8.8 times 10‐11 M. Peptides XII and XIII were inactive, either as agonists or as antagonists of the action of GRF‐(1–44).
Human β‐endorphin
Authors: YAMASHIRO DONALD , LI CHOH HAO , WESTPHAL MANFRED
Published: August 2010

Three analogs of human β‐endorphin (βh‐EP) were synthesized by the solidphase method: [Gln8, Trp27]‐βh‐EP (I), [Gln8,Arg9,Trp27]‐βh‐EP (II), and [Gln9,Arg11,Trp27]‐β‐EP (III). Radioreceptor binding assay with use of tritiated βh‐EP as primary ligand gave relative potencies as follows: βh‐EP, 100; I, 778; II, 467; III, 449. Relative potencies in an analgesic assay were: β‐EP, 100; I, 114; II, 165; III, 83. The 8–11 segment of βh‐EP can tolerate a net increase in charge of + 2 without diminishing analgesic potency. The substitution of Glu8may be one of the more dependable means of designing β‐endorphin antagonists.
Effects of β‐endorphin on the contraction and electrical activity of the isolated perfused rat heart
Authors: LEE A.Y.S. , ZHAN C.Y. , WONG T.M.
Published: August 2010

β‐Endorphin at pharmacological doses reduced both the left ventricular systolic and diastolic pressures of the isolated rat heart. The reduction in the diastolic ventricular pressure was dose dependent. β‐Endorphin also altered the electrical activities of the isolated heart, disturbing the normal electrocardiogram pattern. The degree of disturbance was also dose dependent. Naloxone itself did not produce any effect. However it antagonized the depressant effects and completely abolished the effect of β‐endorphin on the electrical activity of the heart, indicating that β‐endorphin acts via the naloxone sensitive receptors. The results suggest a possible regulatory role of intracardiac endogenous opioid peptides in the cardiac functions.
Solid phase synthesis of retro‐inverso peptide analogues
Authors: BONELLI FABIO , PESSI ANTONELLO , VERDINI ANTONIO S.
Published: August 2010

The solid phase synthesis of a partially modified retro‐inverso analogue of the bradykinin potentiating peptide BPP9a, [gLys6, (R,S)‐mPhe7, Ala8] BPP9a is described. The analogue, which is active in vitro and in vivo, displays prolonged resistance towards cleavage by angiotensin converting enzyme (ACE).
Enzymatically active subunits of Bacillus stearothermophilus enolase bound to Sepharose
Authors: VERONESE FRANCESCO M. , BOCCÙ ODDONE SCHIAVON ENVIRO , BENASSI CARLO A. , FONTANA ANGELO
Published: August 2010

The octameric enolase from Bacillus stearothermophilus was immobilized onto Sepharose 4B activated by the cyanogen bromide reaction under conditions for achieving essentially a single‐point attachment. The immobilized enzyme was dissociated with guanidine hydrochloride to yield bound monomeric enolase. The Sepharose‐bound subunit regained activity upon removal of the denaturant. It was also possible to rehybridize immobilized monomers to native octamers. Of note, the thermal stability of the immobilized enolase subunit does not appreciably differ from that of the parent soluble octameric enzyme. Thus, these results indicate that single subunits of thermophilic enolase are active and that oligomerization is not a prerequisite for the enzymic activity as well as for thermal stability.
Coupling in the absence of tertiary amines: II
Authors: BODANSZKY MIKLOS , BODANSZKY AGNES
Published: August 2010

Formate salts of amino‐components obtained by acidolysis of tert. butyloxy‐carbonyl‐derivatives with formic acid were converted to the tetrazolate salts which in turn were acylated with active esters. Addition of a tertiary base was not necessary for ready coupling. Further improvement was achieved by using a solution of tetrazole in formic acid for the removal of the tert.butyloxy‐carbonyl group.
Denaturation of concanavalin A by urea at acid pH
Authors: AUER HENRY E. , SCHILZ TIMOTHY
Published: August 2010

The denaturation of dimeric concanavalin A induced by urea at pH 3 has been studied using optical activity and sedimentation velocity. Under the conditions employed Mn+2 and Ca+2 are dissociated from the protein, but the basic structural elements are little changed from those prevailing in the functional lectin at pH 5.5 [H.E. Auer and T. Schilz, preceding paper in this issue]. The protein passes through three stages as the urea concentration is varied from 0 to 10 M. Below 4M urea the only effect observed is the loss of optical activity of the aromatic amino acid residues. At 4 M, a conformational change occurs producing extensive aggregation, which persists to 7 M. At 8–10 M urea a disordered monomeric protein molecule prevails. The protein could be reactivated provided that dilution to native conditions was very rapid and the protein concentration remained very low.

Kinetics of denaturation were monitored by optical activity at 218, 225 and 283 nm, Transients with one, two or three components were observed, which were resolved by nonlinear regression according to sequential first‐order decay laws. First order character was confirmed by independence of the kinetic parameters from protein concentration over a two‐ to four‐fold range. Enthalpies and entropies of activation for the various steps were also determined. The transients at the three wavelengths monitor changes in β structure, β turns and aromatic groups, respectively. The urea dependence of the rate constants is unique in most cases. It is concluded that different structural elements of the concanavalin A molecule unfold independently from one another.
Type II‘β‐bend conformation of tert.‐butyloxycarbonyl‐L‐amino‐succinyl‐L‐alanyl‐glycine methyl ester in the solid state
Authors: CAPASSO S. , MAZZARELLA L. , SICA F. , ZAGARI A.
Published: August 2010

Boc‐L‐Asu‐L‐Ala‐Gly‐OMe crystallizes in the monoclinic space group P21 with cell dimensions a = 14.315(3) Å, b = 9.280(2) Å, c = 14.358(3) Å, β= 103.63 (1) d̀, V= 1853.4(9) Å3, with two molecules in the asymmetric unit. The conformation of the two molecules is characterized by a type II' β‐bend, similar to that predicted earlier by potential energy calculations, stabilized by an intramolecular hydrogen bond. I.r. and 1H‐n.m.r. data show that the folded conformation is also stable in chloroform solution.
β‐Endorphin
Authors: HAMMONDS R. GLENN , FERRARA PASCUAL , LI CHOH HAO
Published: August 2010

Inhibition of binding of βh‐endorphin or Leu‐enkephalin by βh‐endorphin analogs of various chain lengths in membrane preparations of the neuroblastoma x glioma NG108–15 cells has been investigated. The removal of even a single residue from theC‐terminus results in the inability of the resulting peptide to completely displace βh‐endorphin. In addition, the proportion of nondisplaceable binding increases with decreasing chain length.
Sugars decrease the thermal denaturation and aggregation of arachin
Authors: KELLA NAVIN KUMAR D. , POOLA INDRAVATHAMMA
Published: August 2010

The effect of three sugars D‐xylose, D‐glucose and D‐sucrose (with increasing hydroxyl content) on the thermal denaturation and protein‐protein interactions of arachin at pH 3.6 were studied by various physicochemical techniques. These included gel melting temperature, turbidity, electrophoresis, viscosity, u.v. difference, fluorescence and circular dichroic measurements. Sugars decreased the heat‐induced aggregation of arachin as shown by the gel melting temperature, turbidity and electrophoretic measurements. Arachin became more compact in the presence of sugars as shown by the reduced viscosity, u.v. difference and fluorescence spectral measurements. Far u.v.‐CD spectral measurements suggested that sugars increase the ordered structure. Viscosity and spectral measurements also showed that heating and cooling in the presence of sugars at pH 3.6 decrease the thermal unfolding of arachin. The effectiveness of the sugar to bring about all the described changes increased with its hydroxyl content. The preventive effect of sugars on the thermal denaturation and aggregation of arachin probably arises from their capacity to strengthen the intra‐protein hydrophobic and hydrogen bonding.
Isolation, purification and 13C‐and 1H‐n.m.r. assignments of peptide [1–24] of human serum albumin
Authors: LAUSSAC JEAN‐PIERRE , SARKAR BIBUDHENDRA
Published: August 2010

Isolation, purification and 360 MHz 1H‐ and 13C‐n.m.r. spectra of the residue corresponding to the NH2‐terminal peptide fragment [1–24] of human serum albumin are reported. The various resonances have been assigned to individual amino acid residues and their spatial microenvironment has been determined in a straightforward manner on the basis of (i) pH dependent chemical shifts; (ii) combined use of multiple and selective proton‐decoupled 1H‐ and 13C‐n.m.r. spectra; (iii) the characteristic pK values exhibited by protons adjacent to sites of ionization in the molecule; and (iv) comparison of the spectra with the NH2‐terminal tripeptide segment of human albumin. The pK values of different ionizable groups all fall in the normal range expected for each titrating sites and support a model of peptide fragment [1–24] in which there is no special structure‐forming strong associations. These results are in agreement with those obtained by CD spectroscopy.
Cyclic analogues of bradykinin
Authors: CHIPENS G.I. , MUTULIS F.K. , MYSHLYAKOVA N.V. , MISINA I.P. , VITOLINA R.O. , KLUSHA V.J. , KATAYEV B.S.
Published: August 2010

A study of the biological activity of six bradykinin and kallidin cycloanalogues has revealed that all of them exhibit prolonged hypotensive activity except the inactive cyclo‐(ω‐aminododecanoyl‐ω‐aminododecanoyl‐bradykinin) (CADADB) when assayed in anaesthetized rats in vivo. The threshold dose of cyclo‐bradykinin (CB) in these experiments was found to be 250 μg/kg of body weight. A single dose (500 μg/kg) of this cyclopeptide produces a decrease in the arterial pressure of rats up to 40 mm Hg for more than 2 h. The hypotensive action of other cyclopeptides under the same conditions is characterized by the following values: cyclo‐[∍‐(L‐Lys1, Gly6)‐bradykinin] (CLGB), α‐L‐Arg‐cyclo‐[∍‐(L‐Lys1, Gly6)‐bradykinin] (ACLGB) and cyclo‐∍‐kallidin (CK) – 5 μg/kg, 50 μg/kg, 40 mm Hg, over 2 h; cyclo‐(ω‐aminododecanoyl‐bradykinin) (CADB) – 50 μg/kg, 250 μg/kg, 40 mm Hg, 35 min. Some cyclopeptides appeared to possess myotropic activity in vitro on isolated rat uterus preparations: CB (α = 0.52, pD2 = 7.86), CK (α = 1.0, pD2 = 7.56), CADB (α = 0.81, pD2 = 6.41). CLGB, ACLGB and CADADB failed to show myotropic activity. In contrast to CLGB and CADB, CK was hypotensive in dogs. CB acts as a weak antagonist of bradykinin (pA2 = 5.09 ± 0.06) on rat uterus in vitro.

It is believed that the hypotensive action within this series of compounds is primarily determined by the cycle size, whilst their myotropic activity is due to the presence of a “common” fragment Arg‐Pro‐Pro.

CD spectra of the bradykinin cycloanalogues were obtained in aqueous solution. On CLGB and CK spectra effects of CaCl2, solvent and pH changes were also examined. The CD spectra of the analogues resemble the spectrum of bradykinin, indicating a certain similarity of secondary structures of these compounds.
Synthetic homo‐oligomethionine chemoattractants
Authors: BISMARA CLAUDIO , BONORA GIAN MARIA , TONIOLO CLAUDIO , BECKER ELMER L. , FREER RICHARD J.
Published: August 2010

Using solution methods, we have synthesized several series of L‐methionine homooligopeptides from the dipeptide to the hexapeptide with either a free α‐carboxyl function or different C‐blocking groups (methoxy, benzyloxy, benzy‐lamino) and with N‐blocking groups of either the amide type (formyl, pivaloyl) or the urethane type (tert.‐butyloxycarbonyl). Compounds were compared to determine the combined effect of main‐chain length and presence and nature of N‐ and C‐blocking groups on conformation‐activity relationship. Each peptide was tested for its ability to induce rabbit peritoneal polymorphonuclear leukocytes in the presence of cytochalasin B to secrete granule enzymes. In parallel, a conformational analysis was carried out in the solid state and in solution, using infrared absorption and circular dichroism. The biological and conformational data are discussed in relation to a recently proposed model of the chemotactic peptide receptor of rabbit neutrophils.
Studies of repeating synthetic peptides designed to adopt a cross‐β conformation
Authors: CARBONE F.R. , LEACH S.J.
Published: August 2010

The conformational preferences of peptides forming parts of the sequence Val‐Val‐Asn‐Pro‐Gly‐Gln‐Val‐Val have been studied by circular dichroism (CD) and nuclear magnetic resonance (n.m.r.) spectroscopy. The sequence is intended as a model for cross‐β structures and keratins. CD spectra characteristic of type II β‐bends are produced by Pro‐Gly sequences: these have a maximum at 196–203 nm and a single minimum at 222–225 nm. Peptides with the sequence Asn‐Pro‐Gly have a CD spectrum attributable to a type I(III) β‐bend with two minima at 200–205 nm and 220 nm respectively. N.m.r. data support the suggestion that the type I(III) bend is stabilised by H‐bonding between the oxygen of the side‐chain amide of Asn and the hydrogen of the main chain NH of the Gly (or other third) residue. Thus, sequences where such H‐bonding is possible show shielding of the Asn amide protons, non‐equivalence of CβH2 protons and low temperature dependencies of NH protons of the third residue for each peptide examined. Calculations of rotamer populations for the Asn amide side chain support these conclusions.
Digestion of elastin by porcine pancreatic elastase I and elastase II
Authors: VERED M. , BURSTEIN Y. , GERTLER A.
Published: August 2010

Elastin was fully solubilized by digestion with elastase I or elastase II. Each digest was separated into high‐molecular weight and low‐molecular weight fractions that were characterized by the correspondence to their amino acid content, N‐terminal sequence and C‐terminal amino acids. It was found that although the relative amount of amino acids in the low‐molecular weight fraction obtained by digestion with elastase I was lower than in digestion with elastase II, no major difference in the type of bonds cleaved in the low‐ or high‐molecular weight fractions of each digest could be seen. There is, however, a remarkable difference in the type of bond cleaved by the two enzymes. While elastase I cleaves mostly Ala‐Ala and also Ala‐Gly bonds, elastase II hydrolyzes Leu‐Ala, Leu‐Gly, Phe‐Ala, Phe‐Gly and Tyr‐Ala, Tyr‐Gly bonds. Theoretical calculations led us to suggest both digests are composed of cross‐linked peptides that vary not only in the molecular size but also in the number of cross‐links found in peptides of the same size.
Synthesis of 3,4‐iminocyclohexyl‐glycine and its N‐benzyloxy‐carbonyl derivative
Authors: DZIEDUSZYCKA MARIA , MARTELLI SANTE , BOROWSKI EDWARD
Published: August 2010

3,4‐Iminocyclohexyl‐glycine and its N‐benzyloxycarbonyl derivative were synthesized by the addition of iodine isocyanate to the double bond of properly protected 3‐cyclohexenyl‐glycine, followed by the conversion of addition product to a corresponding iodocarbamate and further cyclization to aziridine. 3,4‐Iminocyclohexyl‐glycine is the first representative of a novel group of amino acids analogs, aziridinoamino acids. The compound is glutamine analog and exhibits weak inhibitory activity in regard to glucosamine synthetase.
Synthesis and biological activity of N‐acyl derivatives of a Saccharomyces cerevisiae mating pheromone
Authors: SHENBAGAMURTHI PONNIAH , KUNDU BIJOY , BECKER JEFFREY M. , NAIDER FRED
Published: August 2010

We report a general method for acylation of the N‐amino group of the lysyl residue in peptides. The procedure involves acylation using p‐nitrophenyl esters and 1‐hydroxybenzotriazole in organic solvents to yield a series of fatty acyl mating pheromones of Saccharomyces cerevisiae. The fatty acyl group does not influence coupling of peptide fragments. Biological activities of the synthesized α‐factor mating pheromones derivatized with acetyl, butyryl, caprylyl and lauryl groups are nearly equivalent to the activity of unacylated α‐factor. The N‐stearyl‐α‐factor is biologically inactive. The procedures reported in this communication can be used to increase hydrophobicity of lysine‐containing peptides when the lysyl group is not essential for activity.
Prediction of strong antigenic determinant of seminalplasmin and ribonuclease from the amino acid sequence
Authors: PANDIT MADHUSUDAN W.
Published: August 2010

It has been reported earlier [Hopp, T.P. & Woods, K.R. (1981) Proc. Natl. Acad. Sci. US 78, 3824–3828] that the antigenic determinants of a protein can be delineated by examining the average local hydrophilicity values along the peptide chain. I have used this method to predict the strong antigenic determinants of two proteins, seminalplasmin and ribonuclease, of known sequence. In the former case, the N‐terminal segment 1–14, and in the latter case the segments 27–38 and 80–86, are predicted to possess the antigenic determinants of the two proteins. Experimental verification already exists for the former case.
Conformational energy analysis and mechanical properties of polyglycine II
Authors: NAKAMURA HIDEAKI , GO NOBUHIRO
Published: August 2010

For the purpose of calculating the crystal structure of polyglycine II, a system of three parallel chains are considered. The structure obtained by the energy minimization of this system agrees well with experimentally determined crystal structure. Hydrogen bonds of the type of Cα–H. O, which are discussed in literature as existing in the crystal structure, are neglected in the calculation. The fact that the crystal structure is well reproduced by this calculation indicates that hydrogen bonds of this type, if any, are weak in this crystal structure. The relative contributions of various energy terms to the stabilization of the structure are investigated. Interchain hydrogen bonds do not play a significant role in the determination of the crystal structure, but the interchain nonbonded interactions do. Mechanical properties are studied by calculating a response to a static external force applied to the system. Young's modulus E = 4.2 times 1011‐dyncm‐2 and Poisson's ratio s̀=0.25 are obtained. Relative contributions of various energy terms to elastic properties are discussed.
Effect of sodium dodecyl sulfate on the heat denaturation and aggregation of arachin at pH 3.6
Authors: KELLA NAVIN KUMAR D. , RAO M.S. NARASINGA
Published: August 2010

The effect of an anionic detergent, sodium dodecyl sulfate, on the heat‐induced denaturation and protein‐protein interactions of arachin at pH 3.6 was studied by melting temperature, turbidity, electrophoresis and fluorescence spectral measurements. Sodium dodecyl sulfate induced aggregation in arachin at room temperature, in the concentration range 1 times 10‐5to 1 times 10‐2M, as shown by turbidity, electrophoresis and fluorescence spectral measurements. However, higher detergent concentrations (> 1 times 10‐2M SDS) decreased the aggregation. Electrophoretic experiments at pH 3.6 showed that sodium dodecyl sulfate at a concentration of 5 times 10‐2M changes the protein charge from positive to negative. Fluroescence spectral measurements suggested that the detergent at 5 times 10‐2M level unfolds arachin. Heating and cooling arachin at pH 3.6 in the presence of sodium dodecyl sulfate increased the protein‐protein interactions as compared to those at room temperature, in the detergent concentration range 1 times 10‐5to 1 times 10‐2M; at higher detergent concentrations there was a sharp decrease. The observed effects of sodium dodecyl sulfate on the structure and conformation and heat‐induced protein‐protein interactions of arachin at the experimental pH may arise out of the detergent binding to the protein involving both ionic and hydrophobic interactions.