bluuebunny/crossref_metadata_2025 huggingface dataset:
Prediction and improvement of protected peptide solubility in organic solvents*
Authors: NARITA MITSUAKI , ISHIKAWA KAZUNORI , CHEN JUNN‐YANN , KIM YUKIO
Published: August 2010

A predictive method of solubility for protected peptide fragments of globular proteins was described. The solubility prediction was performed on the basis of both the randomness of peptide structures in a solid state and the existence of tertiary peptide bonds such as X‐Pro and X‐(Z)Y bonds, in which X and Y are arbitrary amino acid residues and Z is a suitable protecting group for the X‐Y peptide bond. In order to predict the randomness, the coil conformational parameter, Pe, was utilized. Solubility prediction by this method was success fully applied to the two classes of protected peptides composed solely of hydrophobic and of various amino acid residues. The solubility test results also indicate that the protection of peptide bonds at suitable intervals is effective in achieving a remarkable improvement in the solubility of extraordinarily insoluble peptides. Lastly, the strategy for the selection of the coupling routes in the protein syntheses was proposed.
Sequence‐dependence of secondary structure formation II*
Authors: MUTTER M. , ALTMANN K.‐H.
Published: August 2010

The conformational influence of the insertion of two guest amino acids into a homooligopeptide consisting of L‐Val residues was investigated by circular dichroism and infrared spectroscopy. L‐Ala, L‐Ile and L‐Leu were chosen as guest amino acids and their β‐structure disrupting properties studied with respect to size and symmetry of their side‐chains. These studies were complemented by investigations concerning the aggregation behaviour of the various peptides by means of gel permeation chromatography. It is shown that the tendency of the various peptides to form β‐structures and to aggregate increases with increasing similarity in the spatial size of the side‐chains between guest and host amino acids. The impact of those results on the prediction of peptide secondary structure as well as their importance for peptide synthesis is briefly discussed.
Allyl ester as temporary protecting group for the β‐carboxy function of aspartic acid
Authors: KUNZ HORST , WALDMANN HERBERT , UNVERZAGT CARLO
Published: August 2010

The use of the allyl ester as a temporary protecting function in aspartyl peptides is described. The β‐allyl ester was introduced into Boc‐Asp‐Phe‐NH2 4a, Boc‐Asp‐Gly‐NH2 4b and Boc‐Asp‐Ser‐NH2 4c by means of the caesium salt method. It was removed from these molecules 5 using noble metal catalysts under weakly basic, acidic or even neutral conditions likewise in high yield. The formation of an aminosuccinyl derivative during the deblocking reactions was not observed. Treatment of the dipeptide Boc‐Asp(OAll)‐AS‐NH2 with an excess of triethylamine at room temperature induced the ring closure to the cyclic imide.
Synthetic and binding studies on the calcium binding site I of bovine brain calmodulin
Authors: BORIN G. , PEZZOLI A. , MARCHIORI F. , PEGGION E.
Published: August 2010

The synthesis of the cyclic 20–31 sequence of bovine brain calmodulin corresponding to the loop of the hypothetical calcium binding site I of the protein has been accomplished by classical solution methods. The interaction of the synthetic cyclic fragment with calcium ions has been investigated by CD spectroscopy in water and in 98% trifluoroethanol solution. Calcium ions have no effect on the dichroic absorption of aqueous solution of the cyclic dodeca‐peptide in the wavelength range 200–250 nm. In 98% trifluoroethanol the CD spectrum of the cyclic compound in the absence of calcium ions is almost identical to that of the linear dodecapeptide in the presence of saturation concentrations of calcium. This result supports our previous hypothesis of a folding of the linear sequence upon interaction with the metal ion. The cyclic peptide also interacts with calcium ions in 98% trifluoroethanol forming a 1:1 complex.
Human retinoblastomas have binding sites for the COOH‐terminal segment of human β‐endorphin
Authors: WESTPHAL MANFRED , LI CHOH HAO
Published: August 2010

Two human retinoblastoma cell lines (Y79 and McA) were evaluated for the presence of binding sites for human beta‐endorphin (βh‐EP). Using tritiated βh‐EP (3H‐βh‐EP) and synthetic β‐EP analogues, it was possible to demonstrate binding sites for 3H‐βh‐EP with an ED50 of 3.5 nM in Y79 cells and 8 nM in McA cells respectively. The non‐opioid segment [βh‐EP‐(6–31)] retained about 20% relative potency in Y79 and 40% in McA in displacing the tritiated hormone when compared with βh‐EP. Camel β‐EP had a relative potency of less than 1% and βh‐EP‐(1–27) was inactive in both cells in doses as high as 4μm. Taken together with previous reports on similar binding sites in human neuroblastoma and glioblastoma cell lines, it appears that cell lines of neural origin have binding sites for the COOH‐terminal of human β‐EP.
Evidence for the existence of [Gln9] ‐β‐lipotropin in human pituitary glands
Authors: CHUNG DAVID , LI CHOH HAO
Published: August 2010

The isolation of two peptides similar in amino acid composition to that of human β‐lipotropin is presented. Peptide patterns after enzymatic digestions of these two peptides by Staphylococcus aureus protease and by trypsin were nearly identical. Paper electrophoresis and amino acid analyses of acidic peptides generated from the enzymatic digestions of these two peptides indicate that there is an amide difference between the two peptides. It is proposed that this amide difference is in amino acid residue number nine, and that one is the human β‐lipotropin and the other its [Gln9] analog.
Generating plausible protein folds by secondary structure similarity
Authors: SHERIDAN ROBERT P. , DIXON J. SCOTT , VENKATARAGHAVAN R.
Published: August 2010

We extend a standard method of comparing protein sequences to develop a method for quantitatively comparing the secondary structures of two proteins and finding the best alignment between them. We introduce a new approach to generating a set of plausible folds for a protein given only an assigned secondary structure (a query) and use this secondary structure comparison method to search the set of proteins with known tertiary structures for an array of secondary structure elements similar to the query. A large fraction of the similar arrays found in known proteins have folds that can be taken as candidate folds for the query. The approach is analogous to “protein extension” based on primary structure. We show, for example, how the method can produce plausible folds for the secondary structure of flavodoxin, including the correct one.
Kinetic study of peptide synthesis in the presence of imidazole
Authors: GIRIN S.K. , SHVACHKIN Y.P.
Published: August 2010

The interaction of N‐protected amino acid active esters with imidazole has been found to be a reversible reaction. The rate constant values for a direct reaction of tert.‐butyloxycarbonylglycine p‐nitrophenyl ester with imidazole and for a reverse reaction of tert.‐butyloxycarbonylglycine imidazolide with p‐nitrophenol are reported. The effect of imidazole on the kinetics of active ester and imidazolide reactions with l‐threonine methyl ester were studied. The influence of imidazole on the kinetics of O‐acylation of N‐benzyloxycarbonyl‐l‐serine amide was also investigated. The data obtained enable the optimum strategy to be chosen in the synthesis of peptides containing hydroxy‐amino acid and histidine residues.
Destabilization of collagen structure by amides and detergents in solution
Authors: NANDI P.K. , GRANT M.E. , ROBINSON D.R.
Published: August 2010

The effects of amides and detergents on collagen to gelatin transition have been studied at neutral pH. Simple amides denature the protein. The substitution of H‐atoms by the alkyl groups at the nonpolar end of amide increases the effectiveness of the compounds in destabilizing the collagen structure whereas substitution of the H‐atom at the polar amide end shows marginal effects on the collagen transition. The capabilities of these reagents to denature collagen are much less pronounced than their effects on denaturing globular proteins. Anionic detergents are found to destabilize collagen at very low concentrations (below their cmc values). In this respect, the effects of the detergents on collagen are comparable to the denaturing effects of the detergents on globular proteins. The effect of detergents increases with the increase in the length of the alkyl chain. The structure of the anion in the detergent is also important as seen from the lower potency of the sulfonate containing detergent compared to the sulfate containing detergent in denaturing collagen. Cationic and nonionic detergents do not denature collagen.
Peptide conformations. 33. Conformational analysis of cyclic enkephalin analogs of the type Tyr‐cyclo‐(‐N ω ‐Xxx‐Gly‐Phe‐Leu‐)*
Authors: KESSLER HORST , HÖLZEMANN GÜNTER , ZECHEL CHRISTIAN
Published: August 2010

Four conformationally restricted cyclic enkephalin analogs of the type Tyr‐cyclo(‐Nω ‐Xxx‐Gly‐Phe‐Leu‐) with Xxx = l‐Orn, d‐Orn, l‐Lys and d‐Lys have been synthesized by conventional methods and the conformation of the Bocprotected, the deprotected as well as the des Tyr1 analogs analysed by proton and nitrogen n.m.r. spectroscopy. The assignments, of the resonances were performed by two‐dimensional homo‐ and heteronuclear correlated spectroscopy in the normal (COSY, SECSY) as well as a modified (for the detection of small couplings) version. NOE difference spectroscopy was used to distinguish the amino acid residues with aliphatic side chains. The n.m.r. parameters suggest a rather rigid conformation of the ornithine analogs with two internal NH protons whereas the lysine peptides appear to be more flexible. The structure of the Orn2‐analogs can be described by a Gly3‐CO HN‐Leu5‐γi‐turn and a hydrogen bond Orn2‐CO HN ‐Orn2. The d‐lysine compound seems to include a β‐turn (Gly3‐CO HN‐d‐Lys2). The relation of the different conformational properties of the four cyclic peptides and their biological activities are briefly discussed.
In search of new methods in peptide synthesis. A review of the last three decades
Authors: BODANSZKY MIKLOS
Published: August 2010

An attempt was made to discern ideas and trends in the development of peptide synthesis and to recognize general principles of the discipline. Introduction of efficient methods of activation and coupling during the early years of the reviewed period was followed by only moderate further improvements. Major advances were achieved by the discovery of novel methods of protection and by techniques of facilitation. Improvements in the methods of deblocking hold considerable promise and might bring significantly closer the goal of peptide synthesis: the direct preparation of homogeneous products.
Crystal structure and conformation of the cyclic tetramer of a repeat tripeptide of elastin, cyclo(l‐valyl‐l‐prolylglycyl)4
Authors: COOK WILLIAM J. , TRAPANE TINA L. , PRASAD KARI U.
Published: August 2010

X‐ray diffraction data were used to determine the crystal structure of cyclo‐(l‐Val‐l‐Pro‐Gly)4, the cyclic tetramer of a repeat tripeptide of elastin. The crystals are monoclinic, space group C2, with a = 29.639(3), b = 7.099(1), c = 20.325 (2) Å, and β = 130.4(4)°. The structure was solved by direct methods and refined by least squares to R = 0.082 for 2603 observed reflections. The cyclic dodecapeptide contains two β(II) turns. Hydrophilic and hydrophobic channels that run parallel to the b axis are formed by the stacking of cyclic peptides on twofold axes.
Protection and deprotection of horse cytochrome c
Authors: BOON P.J. , TESSER G.I.
Published: August 2010

The last step in the semisynthesis of horse cytochrome c analogues (formation of the bond 65–66) requires the conformation of the complex between two complementary fragments, (1–65) lactone and (66–104). The fragments can be obtained from a limited degradation with cyanogen bromide. The amino component in this reaction can also be obtained from organo chemical synthesis in which the C‐terminal fragment (81–104) is required in a selectively protected form. The latter is available from a cyanogen bromide degradation of ubiquitously protected cytochrome c. The details of the protection/deprotection reaction and the properties of nonadecamethylsulfonylethyloxycarbonyl cytochrome c are described.
Mechanism of the reaction catalyzed by acyl‐CoA: lysolecithin acyltransferase from rabbit lung.
Authors: ESTRADA P. , ACEBAL C. , ARCHE R.
Published: August 2010

This paper deals with the first attempt to elucidate the chemical mechanism of acyl‐CoA:lysolecithin acyltransferase from rabbit lung, a key enzyme in the metabolism of lung surfactant. For this purpose, the pH dependence of kinetic constants as well as the chemical modification of the protein have been studied on a partially‐purified preparation.

From these experiments, the pKs on which the activity of the enzyme relies have been calculated, giving values of pK1 ⋍ 5.5 and pK2 ⋍ 10. Analysis of the effect of organic solvents on these pKs and the calculation of the enthalpies of ionization, together with the chemical modification experiments, lead to the conclusion that pK1 is due to an histidine residue, whereas pK2 arises from the amino group of the adenine ring of palmitoyl‐CoA. Moreover, chemical modification demonstrated an essential cysteine.

A tentative chemical mechanism, in accordance with these results, is proposed and it is hypothesized, in view of other results obtained in our laboratory and from the literature, that the chemical mechanism of acyl transfer to sn‐2 position may be common to other enzymes of glycerolipid metabolism.
Effect of alkylation with different sized substituents on the conformation of ovomucoid, lysozyme and ovotransferrin
Authors: FRETHEIM KRISTEN , EGELANDSDAL BJØRG , HARBITZ OLE
Published: August 2010

Conformational changes induced in ovomucoid, lysozyme and ovotransferrin on reductive addition of different sized substituents have been studied employing differential scanning calorimetry (DSC) and circular dichroic spectroscopy (CD). The thermograms obtained by DSC revealed that extensive introduction of methyl, isopropyl, cyclopentyl, cyclohexyl, benzyl or n‐butyl groups has a detrimental effect on thermal stability (enthalpy of denaturation); the effect generally increases with the size of the substituent. Circular dichroic spectra were affected only to a very limited extent by the modifications, near‐u.v. spectra remaining much the same while far‐u.v. spectra displayed minor changes. The general conclusion drawn is that the modifications had only limited effects on the conformation of the proteins while, nonetheless, perturbing (or breaking) long‐range intramolecular interactions so as to destabilize the structure. Derivatization of lysozyme and ovotransferrin with some of the larger groups has been reported to result in spontaneous precipitation of the proteins [Fretheim, K., Iwai, S. & Feeney, R.E. (1979) Int. J. Peptide Protein Res. 14, 451–456]. The present investigation indicates that precipitation was caused by (partial) denaturation (and ensuing aggregation) as a consequence of modification.
Synthesis and biological activity of linear and cyclic enkephalins modified at the Gly3‐Phe4amide bond
Authors: RICHMAN S.J. , GOODMAN M. , NGUYEN THI M.‐D. , SCHILLER P.W.
Published: August 2010

As part of our continuing effort to define structure‐activity relationships for enkephalin and design enzymatically resistant analogs, we report the synthesis and biological activities of linear and cyclic enkephalin analogs modified at the Gly3‐Phe4amide bond. The partial retro‐inverso enkephalin analog Tyr‐d‐Ala‐gGly‐(R,S)‐mPhe‐Leu‐NH2 and its cyclic counterpart, Tyr‐cyclo[d‐A2bu‐gGly‐(R,S)‐mPhe‐Leu‐], were synthesized as diastereomeric mixtures using solution methodology. The racemic benzylmalonate allowed the linear analog to be synthesized by fragment coupling at the reversed bond. Cyclization of the second analog was carried out at high concentration, eliminating formation of polymer by the use of an insoluble base. All gem‐diaminoalkyl residues were prepared by conversion of peptidyl amides with benzene iodonium bis(trifluoroacetate). Diastereomers of both compounds were separable by reverse phase HPLC but those of the linear compound racemized rapidly under conditions of testing and were therefore tested together. All analogs tested had activities ranging from 6 to 14% of the activity of Leu enkephalin, indicating that the Gly3‐Phe4amide bond is important, though not crucial, for receptor binding.
Synthesis of a docosapeptide comprising the hydrophobic membrane spanning region of glycophorin A
Authors: GALARDY RICHARD E. , KORTYLEWICZ ZBIGNIEW P.
Published: August 2010

The docosapeptide which constitutes the membrane spanning region (amino acid residues 73–94) of the human red blood cell protein glycophorin A has been synthesized. This may be the first example of the synthesis of the entire membrane embedded domain of a membrane spanning protein. Three fully protected fragments were prepared by stepwise elongation using dicyclohexyl‐carbodiimide and p‐nitrophenyl ester activation of Nα‐tert.‐butyloxycarbonyl amino acids. The three fragments represent amino acid residues 73–79, 80–86, and 87–94 in the sequence of glycophorin A and contain a large proportion of valine, leucine, and isoleucine residues but contain no amino acids with ionizable side chain functional groups. The three fragments were condensed using both the azide method and the dicyclohexylcarbodiimide method to give fully protected docosapeptide. Benzyl groups protecting the side chains of the docosapeptide were removed by prolonged hydrogenolysis to give the desired product Nα‐tert.‐butyloxycarbonyldocosapeptide ethyl ester. High resolution proton n.m.r. spectra of the protected fragments in 100% deuterochloroform showed all resonances to be broadened with the amide resonances broadened beyond recognition. In perdeuterodimethylsulfoxide all resonances were relatively sharp with all amide resonances visible and showing coupling constants of 7–8 Hz. Solvent titration of the proton spectra of two of the fragments from 100% perdeuterodimethylsulfoxide to 100% deuterochloroform demonstrated a transition to the broadened spectrum, accompanied by a decrease in the coupling constant of the amide protons (JNH‐CHα) suggesting solvent dependent onset of intramolecular secondary structure, possibly accompanied by aggregation. A proton n.m.r. spectrum of the docosapeptide in perdeuterodimethylsulfoxide shows a few resolved amide resonances with coupling constants of 7–9 Hz. Solvent titration with perdeuterochloroform again suggests a transition to a rigid intramolecular secondary structure.
Solid phase synthesis of ovine corticotropin releasing factor
Authors: MORELL JOHN L. , BROWN JUDITH H.
Published: August 2010

Ovine corticotropin releasing factor was synthesized by the stepwise solid phase method under conditions anticipated to maximize yield. Final yields of fully active hormone were 34% based on peptide which could be removed from the resin and 24% on initial resin substitution.
Photoreactive enkephalin analogue: [D‐Ala2,p‐N3‐Phe 4‐Met 5] enkephalin
Authors: YEUNG CLEMENT W.T.
Published: August 2010

A photoreactive [D‐Ala2,p‐N3‐Phe4‐Met5]enkephalin was synthesized by classical solution peptide synthetic methods. The hydroxysuccinimide ester was used in all the coupling steps in the presence of a weak base, triethylamine. The deprotected enkephalin analogue was purified on high performance liquid chromatography using a Waters, C18μBondapak reverse phase column and its purity was assessed by thin‐layer chromatography. The composition of the analogue was determined and confirmed by elemental analysis and amino acid analysis. Its photoreactivity was demonstrated by the time dependent ultraviolet spectral changes on exposure to light. Competition receptor binding showed that [D‐Ala2,p‐N3‐Phe4‐Met5]enkephalin was 4‐fold more potent than [D‐Ala2, Met5]‐enkephalin in competing for binding to the enkephalin binding site. The data presented suggest that this photoreactive enkephalin analogue may be suitable for use in thein situphotoaffinity labeling of the enkephalin receptor.
Use of proline‐specific endopeptidase in the isolation of all four “native” disulfides of hen egg white lysozyme
Authors: HAEFFNER‐GORMLEY LORRAINE , PARENTE LINDA , WETLAUFER DONALD B.
Published: August 2010

The disulfide peptides from the tryptic digestion of cyanogen bromide‐treated hen egg white lysozyme (HEWL) were isolated by reverse phase high performance liquid chromatography (HPLC) and identified by amino acid analysis. Three peptides containing the I–VIII, II–VII, and III–V + IV–VI disulfide bonds were obtained. The two‐disulfide peptide was further digested with proline‐specific endopeptidase (PCE)(EC 3.4.21.26). Amino acid analysis of digest peptides separated by HPLC showed four peptides with the IV–VI disulfide bond as well as a peptide with the III–V disulfide bond. The IV–VI peptides were produced by hydrolysis of several alanine‐X bonds as well as the prolyl‐cystine bond. Our studies show that alanyl peptide bonds to lysyl, seryl, and leucyl residues are susceptible to hydrolysis by PCE preparations, thus substantially extending its known specificity range. The two‐disulfide peptide was also digested sequentially with thermolysin and PCE; the resulting IV–VI and III–V peptides were identified by HPLC and amino acid analysis. PCE showed substantial activity at pH 5.3 as well as at pH 8.3. The lower pH is useful in studies of proteins or peptides where base‐catalyzed reactions must be limited.
Coupling in absence of tertiary amines:
Authors: BODANSZKY MIKLOS , BODANSZKY AGNES
Published: August 2010

Highly acid sensitive amine protecting groups, such as the triphenylmethyl or the biphenylylisopropyloxycarbonyl group, are cleaved by the N‐protected carboxyl component particularly if deprotection is carried out in trifluoro‐ethanol. It was also found possible to apply certain activated derivatives of the carboxyl component (symmetrical anhydrides and 2,4‐dinitrophenyl esters) for deblocking and concurrent acylation in the same solvent.
Studies on opioid receptor selectivity of β‐endorphin antagonists
Authors: HAMMONDS R. GLENN , LI CHOH HAO
Published: August 2010

Opioid receptor selectivity of several β‐endorphin (β‐EP) analogs which antagonize β‐EP‐induced analgesia has been assessed using partially selective binding assays. Although the apparent affinity dissociation constant of β‐EP in these assays varies from 0.2 to 360 nm, the potency of β‐EP antagonists relative to β‐EP remains largely unchanged. It is unlikely that differences in receptor affinities can account for the antagonist properties of these analogs in vivo.
Models of molecular evolution
Authors: KRICHELDORF HANS R. , AU MICHAEL , MANG THOMAS
Published: August 2010

Acetyl, benzyloxycarbonyl and tosyl protected DL‐alanine, DL‐leucine, DL‐methionine, DL‐phenylalanine, and DL‐valine were condensed with DL‐amino acid methyl esters. Cyanamide, diethylcyanamide, diisopropyl carbodiimide, dicyclohexyl carbodiimide, and N‐cyclohexyl‐N‘‐(2‐morpholino ethyl) carbodiimide‐N“‐methotosylate served as condensing reagents. Water, methanol, and dimethylformamide were used as reaction media. The stereochemical course of these dipeptide syntheses was elucidated by means of 13C‐n.m.r. spectroscopy. The formation of L‐L and D‐D bonds (isotactic sequences) was favoured in ca. 80% of all condensations. L‐L/L‐D (D‐D/D‐L) ratios of up to 6:1 were found.
Synthesis of peptide labelled with p‐iodophenylalanine which corresponds to the first amphipathic region of apolipoprotein C‐I
Authors: HARDING D.R.K. , HANCOCK W.S. , SPARROW J.T.
Published: August 2010

The amino terminal fragment (1–15) of apolipoprotein C‐I, H‐Thr‐Pro‐Asp‐Val‐Ser‐Ser‐Ala‐Leu‐Asp‐Lys‐Leu‐Lys‐Glu‐Phe14‐Gly was prepared by the solid phase method. However, the phenylalanine residue at position 14 was replaced with p‐iodophenylalanine in the chemical synthesis. The preparation of t‐butyloxy‐carbonyl‐p‐iodophenylalanine is described. After completion of the synthesis, the product was deprotected and cleaved from the resin with liquid HF. The peptide was purified by gel filtration on Sephadex G‐25 and preparative high performance liquid chromatography. The purified material was shown to be homogeneous by amino acid analytical data and by chromatography in three different analytical reversed phase HPLC systems. The peptide was then labelled by the chloroamine‐T procedure and good incorporation of 125I was obtained. After purification of the product by gel filtration on Biogel P‐2, the labelled pentadecapeptide was tested for ability to bind to Very Low Density Lipoproteins (VLDL) in the following manner: VLDL was isolated from normolipemic serum by ultracentrifugal flotation and 100‐μul samples were incubated with labelled material dissolved in 200 μul of 0.5 m NaCl, 0.02 m sodium phosphate buffer, pH 7.4 at 37° for 30min. The VLDL fraction was again isolated by ultracentrifugal flotation and the incorporation of radioactivity into the lipoprotein was measured. Under these conditions, a sample of [3H]‐native apolipoprotein C‐I was incorporated to an extent of 83% of the total sample, while the [125I]‐pentadecapeptide exhibited an incorporation of 8.7%.
Synthesis of aldehydic peptides inhibiting renin
Authors: FEHRENTZ JEAN‐ALAIN , HEITZ ANNIE , CASTRO BERTRAND
Published: August 2010

Reduction of peptidyl N,O‐dimethyl hydroxamates with lithium aluminium hydride in diethyl ether at 0° allowed the preparation of peptidyl aldehydes in excellent yield and optical purity. These aldehydic peptides are able to inhibit renin activity. They are the shortest renin inhibitors known to date.
Conformational preferences of cyclopropyl peptides
Authors: VARUGHESE K.I. , SRINIVASAN A.R. , STAMMER C.H.
Published: August 2010

Crystal structure analysis of (E)‐DL‐1‐benzamido‐1‐methoxycarbonyl‐2‐chlorocyclopropane (C12H12NO3Cl) is reported. The (about N1‐C1 bond) and ø' (about C1‐C11 bond) torsional angles for this compound are ‐ 62.5° and ‐ 33.0°, respectively, and are close to the ø values of the 310 helix and the α‐helix. Semi‐empirical potential energy calculations are performed on a cyclopropyl dipeptide which is a special case of α,α‐disubstituted dipeptide where the α‐carbon and the two substituent carbon atoms form a 3‐membered ring. Our calculations show that different types of helics: α‐, γ‐, ←‐, ω‐, 310‐ and δ‐helices, are energetically favorable. Another interesting possibility is the formation of a cyclic pentapeptide with five‐fold symmetry. The effect of substitutions on Cβ atom are also studied with the help of potential energy maps. Selective substitutions on Cβ atom may be used effectively to restrict either ± or ø values into a very narrow range.
Equilibrium and kinetic study of sodium‐and potassium‐induced conformational changes of apo‐α‐lactalbumin
Authors: HIRAOKA Y. , SUGAI S.
Published: August 2010

Equilibrium and kinetics of Na+‐and K+‐induced conformational changes of apo‐α‐lactalbumin were studied by means of circular dichroism. While apo‐α‐lactalbumin was considerably unfolded in the absence of Na+ or K+ in 20 mM Tris at pH 8.0 and 25°, both the monovalent cations restored the tertiary structure of the protein. Apparent binding constants of Na+ and K+ to the apoprotein were estimated from the equilibria of the Na+‐ and K+‐induced conformational changes. Based on kinetic data of the conformational changes induced by the monovalent cations, binding mechanism of the ions to the apo‐protein was examined. Bound alkali‐metal ions stabilize the native‐like state and an activated state in the unfolding‐refolding reaction of the apoprotein.
Semisynthesis of linear gramicidins using diphenyl phosphorazidate (DPPA)
Authors: WEISS LAWRENCE B. , KOEPPE ROGER E.
Published: August 2010

Sequential HPLC analysis has been used to optimize a synthetic scheme for the preparation of semisynthetic position 1 analogues of the channel‐forming pentadecapeptide, gramicidin A. Diphenyl phosphorazidate (DPPA) was more efficient than dicyclohexylcarbodiimide (DCC) for the coupling of N‐formyl amino acids to des(formyl‐valyl) gramicidin A. The DPPA coupling proceeds rapidly, and with high yield, at 0°. The absence of significant (<0.1%) racemization is demonstrated by the lack of electrophysiologically active formyl‐L‐valine gramicidin A in preparations of formyl‐D‐valine gramicidin A. This scheme has proved useful for the preparation of microgram to gram amounts of position‐1‐substituted gramicidin analogues suitable for crystallography and electrophysiology. Details of the techniques for the preparation of these highly purified analogues are discussed.
Stability of vicilin, a legume seed storage protein, with step‐wise electrostatic modification
Authors: ISMOND M.A.H. , MURRAY E.D. , ARNTFIELD S.D.
Published: August 2010

Step‐wise chemical modification of lysine residues with maleic anhydride was used to examine the influence of electrostatic perturbation on the structural integrity of vicilin from fababeans. There were no significant changes in denaturation temperature (Td), enthalpy of denaturation (ΔH) and one‐half band width (1/2 bw) values up to 63.0% modification. Exposure of hydrophobic surface residues decreased initially to 43.9% maleylation, indicative of minor conformational fluctuations with increased surface negativity. Maleylation of vicilin to 93.8% resulted in a significant decrease in ΔH and Td values and an increase in 1/2 bw and surface hydrophobicity (So) parameters. Although progressive labelling appeared to promote unfolding and subunit dissociation, maximum modification did not result in complete vicilin denaturation. The possible existence of a critical maleylation point was attributed to the cumulative effects of electrostatic repulsion and/or modification of specific key residues.
Synthesis of hydrophobic enkephalin amides
Authors: SOLE NURIA , REIG FRANCISCA , ANTON JOSE Ma GARCIA
Published: August 2010

The important role of the C‐terminal residue of enkephalins in bringing about selectivity for the specific opioid receptors and the possibility that certain hydrophobic modifications could enhance the ability of the molecules to cross biological barriers, prompted us to synthesize a series of amides: Tyr‐D‐Met‐Gly‐Phe‐Pro‐NH‐(CH2)n‐CH3 where n is 7, 9, 11 and 13 respectively, derived from the very potent (D‐Met2, Pro‐NH25) enkephalin analogue. The synthesis has been carried out by the solution phase procedures with isolation and characterization of intermediates and final products. Mixed anhydrides and DCC/HOBt have been used as condensation agents. Homogeneity of final products was checked by t.l.c. and reversed phase HPLC under programmed gradient elution. A considerable decrease in the chemical yields in the last coupling step for the C12 and C14 derivatives has been observed, probably due to the steric hindrance of the aliphatic chain at the C‐terminal position.
Conformation, and dissociation‐recombination of chemically deglycosylated ovine lutropin
Authors: BEWLEY THOMAS A. , SAIRAM M.R.
Published: August 2010

The conformations of intact and acid dissociated, as well as the relative rates of conformational repair of reassociating, ovine lutropin, and a chemically deglycosylated form of lutropin, have been studied by circular dichroism, zero‐order, and difference absorption spectroscopy. Deglycosylation produces only small conformational changes in the intact or acid dissociated form of the molecule. While conformational repair of reassociating lutropin requires up to 72 h, and goes no further than 75–80%, deglycosylated lutropin reaches greater than 90% conformational repair in less than 4 h. The efficacy of second‐order absorption spectra in characterizing conformations and conformational changes in these molecules is demonstrated.
Preparation and activity of nitrated insulin dimer
Authors: CUTFIELD S.M. , DODSON G.G. , RONCO N. , CUTFIELD J.F.
Published: August 2010

Nitration of insulin using tetranitromethane causes polymerisation involving cross‐linked tyrosyl residues. By performing this reaction with insulin crystals, in which it is known that B16 tyrosine of one monomer is closely associated with B26 of the neighbouring monomer within the dimer, it has been possible to isolate a covalent dimer of insulin cross‐linked between these two tyrosines. It was, however, first necessary to block the reactive A14 tyrosine. Both rhombo‐hedral (hexameric) and cubic (dimeric) pig insulin crystals were used, the latter proving successful in yielding a pure dimeric product as shown by oxidative sulphitolysis and HPLC. The purified nitrated dimer was biologically active (ca. 10% potency compared to monomeric insulin in a lipogenesis assay) suggesting that the residues responsible for insulin's action are present on the surface of the dimer and not buried in the interface.
Interaction of aromatic amino acids with gossypol
Authors: RAO A.G. APPU
Published: August 2010

The aromatic amino acids L‐tryptophan, L‐tyrosine and L‐phenylalanine complex with gossypol at pH 7.6, resulting in negative CD bands at 424–426 nm and 300 nm and a positive band at 355 nm. The association constant and free energy change for complex formation have been calculated from the amplitude of the CD band at 420 nm. The interaction is weak. The association constant for complex formation increases with the hydrophobicity of amino acid. The effect of additives on the reaction has also been determined.
Effect of disulfide bond cleavage on the structure and conformation of glycinin
Authors: KELLA NAVIN KUMAR D. , BARBEAU WILLIAM E. , KINSELLA JOHN E.
Published: August 2010

The effect of S‐S bond cleavage on the molecular properties of glycinin has been studied by u.v. difference spectroscopy, fluorescence spectroscopy, circular dichroism, viscometry, tryptic hydrolysis, turbidimetry and electrophoresis. U.v. difference spectra, fluorescence spectra, viscometry and tryptic hydrolysis revealed that the overall protein structure becomes more flexible up to a cleavage of about 15 S‐S bonds. However, the cleavage of all the protein S‐S bonds decreased the structural flexibility. U.v. circular dichroic spectral measurements revealed that the increase in the conformational flexibility (up to 15 S‐S bond cleavage) results from a decrease in both secondary (ordered) and tertiary structure of the protein. These measurements also indicated that the decrease in the conformational flexibility of the glycinin with all the S‐S bonds cleaved was caused by protein refolding. Turbidity measurements indicated that high ionic strength (γ/2 = 0.3) increases protein‐protein interactions among the completely S‐S bond cleaved polypeptides. Electrophoretic experiments suggested that the basic polypeptides are responsible for the protein aggregation at high ionic strength. Hydrophobic associations are primarily involved in these protein‐protein interactions.
X‐ray analysis of renin substrates
Authors: GEOFFRE S. , LEROY F. , PRECIGOUX G.
Published: August 2010

The crystal structure of the title compound, an analogue of the angiotensinogen‐(10–13) peptide in which the N‐terminal leucine and the C‐terminal tyrosine are respectively replaced by the phenyloxy‐acetic group and by phenylalanine, has been determined by X‐ray diffraction. The peptide crystallizes in the space group P212121 with a = 4.866(1), b = 22.311(3), c = 27.213(4) Å and Z = 4. The crystal structure was solved by direct methods and refined to an R value of 0.056. The molecules adopt a pleated sheet conformation with the hydrophobic residues alternatively situated on the right and left of the main chain. In the crystallographic “a” direction, the molecules are linked by hydrogen bonds and form parallel pleated sheet‐type structures.
Characteristic sequential residue environment of amino acids in proteins
Authors: VONDERVISZT F. , MÁTRAI GY. , SIMON I.
Published: August 2010

The occurrence of all di‐ and tripeptide segments of proteins was counted in a large data base containing about 119 000 residues. It was found that the abundance of the amino acids does not determine the frequency of the various di‐ and tripeptide segments. In addition, the frequency of the various tripeptides cannot be predicted from the observed pair‐frequency values. The pair‐frequency distribution of amino acids is highly asymmetrical, pairs formed from identical residues are generally preferred and amino acids cannot be clustered on the basis of their first neighbour preferences. These data indicate the existence of general short range regularities in the primary structure of proteins. The consequences of these short range regularities were studied by comparing Chou‐Fasman parameters with analogous parameters determined from the results of conformational energy calculations of single amino acids. This comparison shows that Chou‐Fasman parameters carry significant information about the environment of each amino acid. The success of the Chou‐Fasman's prediction and the properties of the pair and triplet distribution of the amino acid residues suggest that every amino acid has a characteristic sequential residue environment in proteins. The observed preferences could be invoked, for example, in protein design or in the study of the evolutionary relationship of proteins.
Synthesis and receptor binding affinity of both E‐and Z‐dehydro‐phenylaline4enkephalins*
Authors: NITZ THEODORE J. , SHIMOHIGASHI YASUYUKI , COSTA TOMMASO , CHEN HAO‐CHIA , STAMMER CHARLES H.
Published: August 2010

The dehydrophenylalanine4‐enkephalin having the E‐configuration (ΔEPhe; phenyl and C = O,cis) was prepared by photoisomerization of the Z‐isomer with 3100 Å light, followed by reversed‐phase HPLC separation of the resulting mixture of the Z‐ and E‐isomers. In the radioligand receptor binding assays, the E‐isomer of [D‐Ala2, ΔPhe4, Leu5]enkephalin exhibited an extremely diminished affinity as compared with the Z‐isomer, namely 150–260‐fold loss of affinity for the delta and mu opiate receptors. The results indicate that the interrelationship of the Tyr1and Phe4residues in the enkephalin molecule seems to be of great importance in receptor recognition.
Synthesis of proctolin analogues and their cardioexcitatory effect on cockroach, Periplaneta americana L., and yellow mealworm, Tenebrio molitor L.
Authors: KONOPIŃSKA DANUTA , SOBÓTKA WIESLAW , LESICKI ANDRZEJ , ROSIŃSKI GRZEGORZ , SUJAK PRZEMYSLAW
Published: August 2010

Five proctolin analogues modified in position 2 of the peptide chain by (4‐NH2)‐ (I), (4‐NO2)‐ (II), (4‐NMe2)‐ (III), (4‐OMe)‐L‐phenylalanine (IV) and β‐(cyclohexyl‐4‐OMe)‐L‐alanine (V) were synthesized by conventional liquid phase method. Biological activity of the obtained peptides was investigated in cardioexcitatory test on two insect species, cockroach, Periplaneta americana L., and yellow mealworm, Tenebrio molitor L. Peptides I–IV exhibited higher activity than proctolin itself.
N.m.r. study of conformational changes in lysozyme around the thermal transition point
Authors: BERNARD MONIQUE , CANIONI PAUL , COZZONE PATRICK J. , BERTHOU JEAN , JOLLÈS PIERRE
Published: August 2010

Natural abundance carbon‐13 n.m.r. at 50.3 MHz has been used to further document the thermal transition that hen egg‐white lysozyme undergoes in solution between 20° and 30°. The study focuses on the temperature sensitivity of more than 50 carboxylic, aromatic and aliphatic single carbon resonances for which unambiguous assignments to specific residues are known. The analysis of selective perturbations in chemical shifts indicates that residues located on both edges of the active site cleft and in the hydrophobic box are primarily involved in the temperature‐induced conformational transition. N.m.r. results are compared with crystallographic data on low temperature (form A) and high temperature (form B) interconverting lysozyme crystals, taking advantage of the recent availability of quality high resolution maps for B form orthorhombic crystals. In most cases, a good correlation is found at the atomic level between residues involved in the thermal transition in solution and in the crystalline state. Discrete discrepancies are noted for some residues such as Trp‐62 and His‐15.
Use of 10% sulfuric acid/dioxane for removal of N‐α‐tertiary‐butyloxycarbonyl group during solid phase peptide synthesis
Authors: HOUGHTEN RICHARD A. , BECKMAN ALAN , OSTRESH JOHN M.
Published: August 2010

The hazards and high costs associated with the use of trifluoroacetic acid (TFA) in the removal of the N‐α‐tertiary‐butyloxycarbonyl (Boc) group during solid phase peptide synthesis prompted an examination of alternative acidolytic reagents for α‐amino group deprotection. N‐α‐Boc‐glycine and N‐α‐Boc‐isoleucine resins as well as an N‐α‐Boc‐peptide resin were used to test the lability to various deprotection mixtures of both the N‐α‐Boc resin group as well as the amino acid or peptide‐O benzyl ester resin linkage. Of the combinations tried, several were found, including 10% H2SO4/dioxane, which gave results roughly comparable to 50% TFA/CH2Cl2. Several peptides, 5–10 amino acid residues in length, have been successfully synthesized using the 10% H2SO4/dioxane mixture and were found to be comparable in purity to the same peptides prepared using the standard TFA/CH2Cl2 method of N‐α‐Boc removal. Thus, for the peptides examined, 10% H2SO4dioxane was found to be an inexpensive, safe, and practical alternative reagent to the more costly and hazardous 50% TFA/CH2Cl2 commonly used in the deprotection step of solid phase peptide synthesis.
Crystal and molecular structure of cyclo(L‐prolyl‐glycyl)3
Authors: KARTHA G. , AIMOTO S. , VARUGHESE K.I.
Published: August 2010

The crystal structure of cyclo(L‐Pro‐Gly)3 was solved using X‐ray crystallographic techniques. The backbone of the peptide is asymmetric and is made up of five trans peptide units and one cis peptide. There is a hydrogen bonded water bridge that links the carbonyl oxygens, O1 and O4. The molecules exist as dimers in the crystal lattice. The two molecules of the dimer are related by crystallographic twofold symmetry and are linked by two N‐H O hydrogen bonds. The crystals are trigonal, space group P3212 with a = 11.379(3), c = 32.93(1) and z = 6. The structure was solved by multisolution methods and refined by least squares technique to an R of 0.083.
Improved method for the synthesis of Nα‐9‐fluorenylmethyloxycarbonyl‐Nδ,ω bis‐adamantyloxycarbonyl‐L‐arginine
Authors: PRESENTINI R. , ANTONI G.
Published: August 2010

A modified method is reported for the prepartion of Nα‐9‐fluorenylmethyloxycarbonyl‐Nδ,ω bis‐adamantyloxycarbonyl‐L‐arginine, giving an overall yield of 60% over three steps based on Nα‐benzyloxycarbonyl‐L‐arginine. Commercially available adamantyl fluoroformate for guanidine function protection of Nα benzyloxycarbonyl‐L‐arginine, catalytic transfer hydrogenation with formic acid on palladium black for removal of the benzyloxycarbonyl protecting group, and fluorenylmethylsuccinimidyl carbonate for the final synthesis, were introduced to simplify and reduce the cost of preparation of this arginine derivative. The reaction conditions have been accurately studied at each step in order to optimize the yields.
Extremely fast hydrogen exchange of ribonuclease‐(1–118) as compared with native RNase A and its implication for the conformational energy state
Authors: ROY SIDDHARTHA , DIBELLO CARLO , TANIUCHI HIROSHI
Published: August 2010

RNase‐(1–118) containing native disulfide bonds is similar in fold to native RNase A but not of lowest Gibbs energy as compared with the isomers containing non‐native disulfide bonds. The present n.m.r. studies have indicated a dramatic increase in the exchange rate of all of the ‘protected’ amide protons of RNase‐(1–118) over RNase A. A calculation shows a large increase in the rate of ‘opening’ of the structure. The exchange rate of the protected amide protons of RNase‐(1–120) is slower than RNase‐(1–118) but much faster than RNase A. Binding with a synthetic complementing fragment (114–124) markedly reduces the exchange rate of 20 to 25 amide protons of RNase‐(1–118). It has previously been shown that binding with a complementing fragment of RNase‐(1–118) generates a lowest Gibbs energy state. Thus, using available thermodynamic information for interpretation, we suggest that a) removal of six carboxy terminal residues of RNase A would disrupt coupling between these residues and those distant in the structure (loss of extra stabilizing energy), b) this would, in turn, alter the enthalpy‐entropy compensation in such a way that the magnitude of Gibbs energy change favoring folding is significantly reduced without a large change of fold and c) in this activated state the molecule would be highly motile.
Partial amino acid sequence of porcine elastase II
Authors: VERED M. , GERTLER A. , BURSTEIN Y.
Published: August 2010

The partial amino acid sequence of porcine elastase II, isolated from crude trypsin Type II, was determined. The enzyme consists of two polypeptide chains, a light chain composed of 11 residues, and a heavy chain (Mr = 23 500) with four intrachain disulfide bridges; the two chains are held together by one interchain disulfide bond. Elastase II was fragmented into several peptides by chemical cleavages with CNBr at the two methionine residues, 99 and 180 (chymotrypsinogen numbering), and with hydroxylamine at the peptide bond following DIP‐Ser195. About 50% of the sequence was determined and the positions of 120 amino acids were located, including the light chain residues and most of the active site residues. The partial sequence shows 64% difference between porcine elastase II and elastase I and only 26% difference between porcine elastase II and bovine chymotrypsin B.
Total synthesis of S‐carbamoylmethyl bovine apocytochrome c by segment coupling
Authors: BLAKE JAMES
Published: August 2010

Three peptide segments corresponding to the complete sequence of the 104 amino acid protein bovine apocytochrome c were synthesized by the solid‐phase method. The peptides Ac‐[Cys(Cam)14,17, GlyS23]‐apocytochrome c‐(1–23) (I), CF3CO‐[GlyS60]‐apocytochrome c‐(24–60) (II), and CF3CO‐apocytochrome c‐(61–104) (III) were purified by chromatography on CM‐cellulose, partition chromatography and/or HPLC. Each of the peptides was reacted with citraconic anhydride to block all of the lysine side chains, and the 61–104 peptide was treated with 10% hydrazine to remove the trifluoroacetyl group, to give the corresponding peptides Ia, IIa, and IIIa. Peptides IIa and IIIa were coupled together by reaction with silver nitrate/N‐hydroxysuccinimide to give the 24–104 sequence. After removal of the trifluoroacetyl group from the amino terminus, peptide Ia was also coupled. Treatment of the peptide mixture with aqueous acetic acid removed the citraconyl groups, and purification by chromatography on CM‐cellulose and HPLC gave a 0.6% yield of [Cys‐(Cam)14,17]‐apocytochrome c. The synthetic product was shown to be identical to a sample derived from native bovine cytochrome c by paper or gel electrophoresis, HPLC and by chymotryptic or tryptic map.
Synthesis of specifically labeled peptides*
Authors: PARNES HOWARD , SHELTON EMMA J.
Published: August 2010

The unnatural labeled amino acid N‐acetyl‐D‐3‐(2‐naphthyl)‐[2,3‐3H] alanine having a specific activity of 43.7 Ci/mmol was obtained in 98.7% optical yield by asymmetric reduction of 2‐N‐acetylamino‐3‐(2‐naphthyl)acrylic acid with tritium gas in the presence of the chiral homogeneous catalyst (S,S)BPPM‐Rh+. This amino acid was used in a microscale solution phase synthesis of the labeled LHRH antagonist [Ac‐D‐(2,3‐3H) Nal1, D‐p‐Cl‐Phe2, D‐Trp3, D‐hArg (Et2)6, D‐Ala10] LHRH. The chiral synthesis of labeled amino acids as a strategy for the general synthesis of labeled peptides is discussed.