bluuebunny/crossref_metadata_2025 huggingface dataset:
Synthesis of a tripeptide with a C‐terminal nitrile moiety and the inhibition of proteinases
Authors: STÜBER WERNER , KOSINA HORST , HEIMBURGER NORBERT
Published: August 2010

The synthesis of the tripeptide d‐Phe‐Pro‐Arg with the nitrite group instead of the carboxylgroup is described. Initially, the corresponding peptide amide was synthesized by conventional methods in solution using Boc and Fmoc as the protecting group for d‐Phe. The dehydration in order to create the nitrite moiety was achieved by treating the peptide amide with phosphorus oxichloride or trifluoroacetic anhydride. Best results were obtained by the use of phosphorus oxichloride in pyridine as the solvent in the presence of imidazole. After deprotection of the N‐terminal amino acid the crude product was purified by chromatography on Butyl‐Fractogel® HW‐40 (S). The purity of the final product was checked on a RP18 phase by hplc. The existence of the nitrite group was demonstrated by i.r. and 13C‐n.m.r. spectra. The peptide nitrite exhibited a strong inhibition of thrombin compared to the tripeptide amide.
Selective reduction of the disulfide bonds of ovine placental lactogen
Authors: CARIDAD J.J. , WOLFENSTEIN‐TODEL C.
Published: August 2010

Reduction and carbamidomethylation of two of the three disulfide bridges of ovine placental lactogen was accomplished by the use of 20‐fold molar excess of dithiothreitol over protein disulfide content. The derivative retained its binding capacity to somatogenic as well as lactogenic rat liver receptors, although the latter was somewhat diminished. The two disulfide bonds exposed to the reducing agent are those located near the carboxy‐ and amino‐terminus. while the larger loop remained intact after reduction. This behaviour is similar to that of bovine growth hormone, where the larger loop was also more resistant to reduction.
Methylene chloride‐soluble and insoluble ureines An X‐ray diffraction, infrared absorption, and proton magnetic resonance study
Authors: TONIOLO CLAUDIO , VALLE GIOVANNI , BONORA GIAN MARIA , CRISMA MARCO , MORETTO VITTORIO , IZDEBSKI JAN , PELKA JACEK , PAWLAK DANUTA , SCHNEIDER CONRAD H.
Published: August 2010

An X‐ray diffraction study has been carried out on single crystals of two symmetrical ureines (N,N′‐diiopropylurea and the familiar N,N′‐dicyclohexylurea) and one unsymmetrical ureine (N‐tert.‐butyl.N′‐methylurea). The presence/absence of intermolecular H‐bonds in the crystal state has been compared to the situation assessed in solution by 1H n.m.r. It was shown by i.r. absorption that soluble ureines may disrupt intermolecular (peptide)N—H‐ O=C(peptide) H‐bonds in methylene chloride, a point of practical interest, particularly in solid‐phase peptide synthesis.
Potential energy calculations on phenylalanine rotamers in different boat forms of proline‐containing cyclic dipeptides
Authors: MAZZA FERNANDO , POCHETTI GIORGIO , LUCENTE GINO
Published: August 2010

Previous potential energy calculations have always indicated that the folded conformation is the most stable one for cyclic dipeptides containing benzylic side chains. We have carried out intramolecular van der Waals potential energy calculations on cyclo‐(d‐Phe‐l‐Pro), N‐(N‐phenylacetyl‐l‐alanyl)‐cyclo‐(l‐Phe‐d‐Pro) and N‐(pyruvoyl)‐cyclo‐(l‐Phe‐d‐Pro) adopting the geometries found in their crystal structures. The calculations made on these compounds, having the peptide skeleton made of the same aminoacid residues but differing in the degree of buckling of the boat, show that there is a dependence between diketopiperazine ring and phenylalanine side‐chain conformations. The folded conformer is preferred as long as it is allowed by the degree of buckling of the boat. When the boat becomes highly buckled, as in the case of the pyruvoyl derivative, the folded rotamer is no longer favoured. This is the first energy calculation to demonstrate that the folded rotamer is not always the most stable one.
Heterochiral N‐formyl methionyl peptides
Authors: EGGLESTON DRAKE S.
Published: August 2010

Crystal and molecular structures for the heterochiral sequence N‐formyl‐l‐methionine‐d‐phenylalanine, 1, and its tertiary butyl ester, 2, are reported. The solid‐state peptide conformation is compared to that observed in solution by n.m.r. techniques. For N‐f‐Met‐d‐Phe, 1, the crystal was orthorhombic, space group P21 2121 with a cell of dimensions a = 5.061(3), b = 16.575(5) and c = 19.656(6) Å at ambient temperature of 293K; V = 1649(2)Å3, Z = 4, Dm= 1.31(2)gcm−3, Dx= 1.307gcm−3, μ(Mokx) = 2.047cm−1. For N‐f‐Met‐d‐Phe‐OtBu. 2, the crystal was monoclinic, space group P21 with a cell of dimensions a = 9.963(2), b = 11.147(2), c = 19.166(3)Å. β= 102.31(1) at 273K; V = 2080(2) Å3 Z = 4, Dm= 1.22(2)gcm−3, Dx= 1.215gcm−3μ(MoKx) = 1.714cm−1. The structures were solved from diffractometer data and refined to conventional final R = 0.046, Rw= 0.053 for F‐Met‐d‐Phe (1301 observations. I ≥ 3σ(I)) and to R = 0.056, Rw= 0.064 for the t‐butylester (2411 observations. I ≥ 3σ(I)). The l‐d acid, 1, crystallizes in an extended β‐sheet conformation with trans‐planar peptide bond; the principal torsion angle values are φ1=– 141.2(4).Ψ1= 149.6(4)3, φ2= 157.4(4)°. The methionine side chain adopts a common coiled conformation with x11= ‐ 58.0(5).x21= 175.1(4), x31= 76.5(5). The Phe side chain adopts the statistically least favored g orientation in contrast to the most populated rotamer in solution. The crystal structure is composed of parallel β‐sheets held together by four weak intermolecular contacts including two C‐H O contacts from the alpha carbons to the formyl and peptide carbonyl oxygens. Sheet layers are joined in a head‐to‐tail fashion through a very short (2.569(4) Å) contact between the carboxyl OH and the formyl oxygen and may be further stabilized by a C‐H—O interaction between the formyl proton and the carboxyl OH. Two crystallographically independent molecules are observed in the crystal structure of N‐f‐l‐Met‐d‐Phe‐OtBu, 2. These are distinct conformational isomers differing principally at both the N and C termini. Particularly noteworthy is the synplanar orientation of the ester C = O with respect to the peptide nitrogen in molecule A. which contrasts to the antiplanar orientation in molecule B. Additionally, the formyl group is coiled more towards the C‐terminus in molecule B. Principal torsion angles are φ1(A) = ‐ 120.6(5), Ψ1(A) = 102.0(6)°, φ2(A) = 128.1(6), φ1(B) = ‐ 94.6(5), Ψ1(B) = 91.9(6)°. 121.7(6)°. The peptide bond is trans‐planar in both molecules. Side chain dispositions are essentially identical in both structures. The Met side chain adopts the zig‐zag trans‐planar conformation while the Phe residue adopts an orientation near + 60° in agreement with rotamer populations observed by solution n.m.r. Typical peptide intermolecular H‐bonding is observed in the ester crystal structure; both the peptide and formyl groups participate in the proposed H‐bonding scheme.
Crystal and molecular structure of Destruxin B
Authors: STEINER JORGE RIOS , BARNES CHARLES L.
Published: August 2010

The cyclic hexadepsipeptide mycotoxin Destruxin B. produced by Metarrhizium anisopliae. crystallizes in the orthorhombic space group P212121, with a = 11.010(2)Å, b = 14.679(5)Å, c = 21.273(7)Å and Z = 4. The structure was solved by direct methods and refined by least‐squares technique to a final unweighted R value of 0.051, for 3361 reflections with I > 3σ(I). The backbone of the peptide is asymmetric and is made of 5 trans peptide and ester units and 1 cis peptide unit. The backbone conformation of this cyclic depsipeptide is very similar to that of Roseotoxin B, an analogous mycotoxin produced by Trichothecium roseum. The conformation in the crystalline state also correlates well with the solution conformation, as reported from proton n.m.r. studies. The crystal packing is directed by van der Waals contacts.
Structural studies of cyclopeptides
Authors: BENEDETTI E. , BAVOSO A. , DI BLASIO B. , PAVONE V. , PEDONE C. , PAOLILLO L. , D'ALAGNI M.
Published: August 2010

The solid‐state and solution conformation of cyclo(l‐His‐d‐His), carried out by X‐ray diffraction analysis and n.m.r. spectroscopy, respectively, is reported. The compound crystallizes in the monoclinic system, space group P21/a with a= 7.726, b= 9.877, c= 8.430 Å. β= 106.8 and Z = 2. The cyclic dipeptide retains in the solid‐state a crystallographic center of symmetry: each molecule sitting on it presents a very flat chair conformation with alternating positive and negative conformational angles, whose values are in the range 5–6. The imidazole groups of the side chains are planar, making an angle of 78 with the average plane of the 2.5‐piperazinedione ring. These aromatic groups are folded back over the 2.5‐piperazinedione ring and they “sandwich” it from opposite sides. Packing is obtained through an H‐bonding scheme which involves the N‐H donors of the peptide groups as well as the N‐H groups of the imidazole moieties. In water solution, at pHs equal to 2.55 and 8, the molecule retains on the average an element of symmetry since only one set of signals is observed for the His protons in the n.m.r. spectrum. From coupling constants the predominant conformation was calculated to be all gauche, in excellent agreement with the solid‐state results.
Purification and conformation of ribosomal protein L25 from E. coli ribosome
Authors: FOX J. WESLEY , OWENS DENNIS P. , WONG KIN‐PING
Published: August 2010

Ribosomal protein L25 from the large subunit of E. coli ribosomes has been purified using a new procedure involving a 2 m LiCl extraction followed by phosphocellulose chromatography in 6 m urea elution buffer. The conformation of purified L25 was studied employing circular dichroism and ultraviolet absorption spectroscopy in reconstitution buffer. The analysis of the far u.v. circular dichroism spectrum of L25 indicates L25 contains ∼ 16%α‐helix and ∼ 19%β‐structure. The conformation of L25 was also studied using the predictive methods of Chou & Fasman and Maxfield & Scheraga. Both of these methods predict approximately three times the percent α‐helix present in L25 as compared with that determined from the analysis of the circular dichroism spectrum. A structure for L25 is predicted which contains two positively charged binding domains and is consistent with published binding data on the interaction of 5S RNA and L25. The large difference in the %α‐helix as determined from the analysis of the circular dichroism spectrum and the predictive techniques is suggested to result from the denaturing effects of 6 m urea used in the preparation of ribosomal proteins.
Conformational free energies of myoglobins of small mammals
Authors: KELLY LENORE , SIMMONS JOE H. , HECK TERRY , HOLLADAY LESLIE A.
Published: August 2010

Myoglobins from three small placental mammals and one small marsupial were isolated from cardiac or skeletal muscle. The conformational free energies of these four myoglobins were estimated from guanidinium chloride unfolding data at pH 8 and 25°. The myoglobins from rat and rabbit are more stable than that of the most stable myoglobin previously studied, that of the sperm whale. In addition, these two myoglobins unfold with greater cooperativity than previously characterized myoglobins. The data obtained herein demonstrate unequivocally for the first time that the stability of homeotherm myoglobins correlates with neither the size of the organism nor its metabolic rate.
The remarkable sensitivity to acid‐catalyzed peptolysis of peptide chains (endopeptolysis) having a succession of three N‐alkylated amino acid residues
Authors: ANTEUNIS M.J.O. , VAN DER AUWERA C.
Published: August 2010

An investigation on the resistance against acids of over 30 peptides containing a repetition of N‐alkylated amino acids has revealed the extremely facile cleavage of the peptide bond (endopeptolysis; ranking from a few minutes to 1 hour in Tfa) between the second and third residue in a sequence of three l‐imino acid residues (called “triad”)‐ This is pronouncedly so if Pro is the head member of the triad. Triads become resistant against hydrolysis, however, when both the first and second positions are occupied by Pro, and even more so if the second member is d‐MePhe. On the other hand, as an exception, diads of iminoacids preceded by Gly also are sensitive to hydrolysis. Two possible mechanisms are proposed, among which the one that is characterized by an intramolecular attack of the N‐nucleophilic site of the first residue on the peptide bond between the second and third unit and thus forming a diketopiperazine‐onium intermediate, is our preference. This sensitivity towards acidolysis and acid‐catalyzed hydrolysis should not be overlooked, not only during the planning of peptide syntheses (having especially implications during acidic deprotection protocols), but it will compromise any strategy that makes use of strong acid conditions, e.g. in chiral determinations of N‐alkylated amino acids in peptide hydrolysates, because acidic peptide breakdown is accompanied by appreciable stereo‐mutation. Finally, the weakened resistance of iminoacid peptide chains is obviously related to trans‐cis isomerization about the first peptide bond within the triads and suggests that endopeptidases may well act initially through a trans‐cis isomerase stage.
Tryptophan reduction and histidine racemization during deprotection by catalytic transfer hydrogenation of an analog of the luteinizing hormone releasing factor
Authors: MÉRY JEAN , CALAS BERNARD
Published: August 2010

(d‐Trp)6‐LHRH:pGlu‐His‐Trp‐Ser‐Tyr‐d‐Trp‐Leu‐Arg‐Pro‐GlyNH2 was prepared by solid‐phase peptide synthesis using the nitro group to protect the guanidine side chain of the arginyl residue. Removal of the side‐chain protecting groups was carried out by catalytic transfer hydrogenation (CTH) using palladium acetate/ammonium formate or palladium on charcoal/formic acid. We show in this paper that this deprotection method induces i) reduction of the tryptophan residue and ii) epimerization at the histidine level (with palladium acetate/ammonium formate). Despite the formation of significant amounts of reduced peptide. CTH enabled us to obtain (d‐Trp)6‐LHRH in relatively good yield.
Methionine‐enkephalin peptides in human teeth
Authors: TANZER FRANCIS S. , TOLUN EMIRGUL , FRIDLAND GENEVIEVE H. , DASS CHHABIL , KILLMAR JOHN , TINSLEY PETER W. , DESIDERIO DOMINIC M.
Published: August 2010

The presence of the free opioid pentapeptide methionine enkephalin (ME) and of ME‐containing peptide(s) was established firmly in decalcified, depulped human teeth by using a combination of methods including RP‐HPLC, radioimmunoassay, radioreceptorassay, trypsin, carboxypeptidase B, fast atom bombardment mass spectrometry, and MS/MS methodology. Positive structural identification of ME was made with mass spectrometry. Those data demonstrate the presence of the preproenkephalinergic A system in the human trigeminal sensory termini.
N.m.r. and computer‐simulated conformational analyses of a nonapeptide found in a human salivary proline‐rich glycoprotein
Authors: LOOMIS RONALD E. , TSENG CHING‐CHUNG , BERGEY E. JAMES , LEVINE MICHAEL J.
Published: August 2010

The amino acid sequence G(1)‐P(2)‐P(3)‐P(4)‐H(5)‐P(6)‐G(7)‐K(8)‐P(9) occurs twice in the proline‐rich glycoprotein (PRG) found in human parotid saliva. As part of our efforts to elucidate the structure‐function relationships of PRG, this nonapeptide sequence (PRG9) was synthesized for the purpose of conformational analyses by high‐resolution proton n.m.r. spectroscopy and computer‐modeling. The empirical n.m.r. spectrum differed from the simulated spectrum in that the overall chemical shift locations were displaced from their random coil positions and the five proline residues had non‐degenerate CαHα protons. Other n.m.r. data indicated that no intramolecular hydrogen‐bonding was present in the PRG. In conjunction with X‐ray crystallo‐graphic data on a triproline‐containing model compound (Kartha, g., Ashida, T. & Kakudo, M. (1974) Acta Cryst.B30, 1861‐1866), four energy‐minimized PRG9 structures were obtained. Two of the structures were energetically unfavorable, while the other two conformations were reasonable. The two most likely structures gave all prolines an S‐type ring pucker, the P(2)‐P(3)‐P(4) sequence as a poly‐L‐proline II helix, the H(5)ϕ=− 90.3°, P(6) and P(9) with trans peptide bond orientation, G(7) in an extended state, and the K(8)ϕ=− 93.2° or ‐ 146.8° for structures # 1 and #2. respectively.
Structural requirements of the oxytocin receptor in rat uterus
Authors: PLIŠKA VLADIMIR , HEINIGER JOERG
Published: August 2010

Published and newly calculated pA2‐values of 147 neurohypophyseal hormone analogues (7 positions varied) acting as inhibitors of oxytocin on isolated rat uterus in vitro have been subjected to fractionation according to the method by Free and Wilson which was slightly modified for this purpose. The computation was carried out in several steps. After each step, substances with outlying pA2 ‐values were el minated. The reduced group containing 73–79% of the original substances displayed a high degree of additivity of side chain contributions (SCC). This group seems to follow the “participation” rule as formulated by Free and Wilson. Analysis of the group of eliminated substances and of the resulting SCC‐spectrum (level diagram) enabled us to draw some conclusions concerning the structural requirements of receptor binding: i) The intact ring structure is necessary for the peptide‐receptor interaction: linear peptides or peptides with an extended ring are always outliers; ii) Carba analogues (substitution with CH2 in the disulfide ring) display better affinities than peptides with an S‐S ring; d‐Arg8 substitution decreases the binding affinity; iii) Considerably better additivity is achieved when peptides are divided into subgroups with vasopressin‐like and oxytocin‐like features; populations of receptors more specific for vasopressin and for oxytocin, respectively, can be assumed. Estimates of the “true” receptor‐peptide dissociation constants can be obtained by summation of the corresponding SCC's in each investigated position. The value obtained for oxytocin is identical with the medium affinity binding site on myometrial cells, and not with the high affinity site. A nonlinear relationship exists between SCC's computed from pA2‐values for magnesium‐free and magnesium‐containing (0.5 mm) media but no evidence speaks in favor of a Mg‐potentiating effect on receptor binding.
Monoclinic polymorph of Boc‐Trp‐Ile‐Ala‐Aib‐Ile‐Val‐Aib‐Leu‐Aib‐Pro‐OMe(anhydrous)
Authors: KARLE ISABELLA L. , FLIPPEN‐ANDERSON JUDITH L. , SUKUMAR MUPALLA , BALARAM PADMANABHAM
Published: August 2010

The structures of two crystal forms of Boc‐Trp‐Ile‐Ala‐Aib‐Ile‐Val‐Aib‐Leu‐Aib‐Pro‐OMe have been determined. The triclinic form (PI, Z= l) from DMSO/H2O crystallizes as a dihydrate (Karle, Sukumar & Balaram (1986) Proc. Natl. Acad. Sci. USA 83, 9284‐9288). The monoclinic form (P21, Z = 2) crystallized from dioxane is anhydrous. The conformation of the peptide is essentially the same in both crystal systems, but small changes in conformational angles are associated with a shift of the helix from a predominantly α‐type to a predominantly 310‐type. The r.m.s. deviation of 33 atoms in the backbone and Cβ positions of residues 2‐8 is only 0.29 Å between molecules in the two polymorphs. In both space groups, the helical molecules pack in a parallel fashion, rather than antiparallel. The only intermolecular hydrogen bonding is head‐to‐tail between helices. There are no lateral hydrogen bonds. In the P21 cell, a = 9.422(2)Å, b = 36.392(11)Å, c = 10.548(2)Å, β= 111.31(2)° and V = 3369.3Å3 For 2 molecules of C60H97N11O10 per cell.
High‐performance liquid chromatography of epimeric N‐protected peptide acids and esters for assessing racemization
Authors: BENOITON N. LEO , LEE YOUNG , LIBEREK BOGDAN , STEINAUER RENE , CHEN FRANCIS M.F.
Published: August 2010

The separations by reversed phase high‐performance liquid chromatography on a μBondapak‐C18 column of 53 epimeric N‐substituted di‐, tri‐ and tetrapeptide acids and esters have been attempted, with success in three quarters of the cases. Substituents include acetyl, benzoyl, benzyloxycarbonyl, tert.‐butoxycarbonyl and 9‐fluorenylmethoxycarbonyl. The series N‐benzyloxycarbonylglycyl‐Xxx‐valine ethyl ester with Xxx = alanyl, valyl, leucyl and phenylalanyl, is recommended for use in studies on racemization. Results on racemization attending the coupling of an amino acid ester as compared with a di‐ and tripeptide ester vary with the coupling method.
Synthesis and receptor binding characteristics of [d‐Ala2, cysteamine5]enkephalin, a thiol‐containing probe for structural elements of opiate receptors
Authors: KODAMA HIROAKI , SHIMOHIGASHI YASUYUKI , COSTA TOMMASO , KONDO MICHIO
Published: August 2010

For the elucidation of structural elements in the opiate receptors, a thiol‐containing enkephalin analog, [d‐Ala2, cysteamine5]enkephalin, and its dimeric analog were synthesized and evaluated in the radio‐ligand receptor binding assays using rat brain membranes. The dimeric analog was very potent in both delta and mu assays. Comparison of receptor affinities of the thiol‐containing enkephalin with those of standard mu or delta receptor specific ligands suggested that the mu receptor contains an essential thiol group which may interact with the thiol group at theC‐terminus of the enkephalin analog. It also appears that no metal‐ion site, postulated for the delta receptors, is present in the delta binding site.
Synthesis and solubility properties of peptide fragments of human hemoglobin α‐chain (123‐136)
Authors: NARITA MITSUAKI , DOI MASAMITSU , NAKAI TAKAO , TAKEGAHARA HITOSHI
Published: August 2010

The solubility prediction method for protected peptides was successfully applied to relatively small peptide fragments of human hemoglobin α‐chain (123‐136) which contained various polar amino acid residues such as Asp(OBzl), Glu(OBzl), Lys(Z), Ser(Bzl), and Thr(Bzl). As reported previously for hydrophobic peptides and human proinsulin C‐peptide fragments, solubility data indicated that the insolubility of protected peptides having a  value below 0.90 appeared to begin at the octa‐ or nonapeptide sequence level and that β‐sheet structure played an important role in the insolubility of peptides. When a peptide has a β‐sheet structure in the solid state, we can clearly determine the critical chain length for peptide insolubility, the solubility dependence on solvent properties, and the solubility independence of amino acid compositions of peptides.
Deprotonation of Glu9 destabilizes the α‐helix in C‐peptide of RNase A
Authors: BIERZYŃSKI ANDRZEJ
Published: August 2010

pK values of Glu2, Glu9, and His12 in the lactone and carboxylate forms of C‐peptide at 4.5 and 21°, 0.1 m NACl, have been determined from pH‐tritration shifts of n.m.r. signals of the glutamate γ and histidine ring protons. These have been used in an analysis of the well known pH‐induced helix‐coil transition in the C‐peptide lactone. It has been shown that Glu 12 deprotonation results in a twofold increase of the helix content, whereas deprotonation of Glu9 leads to a 28% drop. This latter effect is probably related to Glu−9‐His+ 12 salt bridge formation. The transition within the pH range 5.5‐7.5 follows exactly the deprotonation curve of His+ 12 residue.
Non‐covalent interactions in the protonation of diastereoisomeric dipeptides in aqueous solution
Authors: CALÌ R , CUCINOTTA V. , IMPELLIZZERI G. , MAUGERI M.C. , RIZZARELLI E.
Published: August 2010

New and already reported thermodynamic data concerning the protonation of a series of diastereoisomeric pairs of dipeptides have been compared with the upfield chemical shifts observed in the PMR spectra of some systems as a consequence of alkyl‐aryl interactions. The enthalpy and entropy changes associated with the protonation of the dipeptides, obtained by potentiometric and calorimetric measurements, together with the PMR data, show the importance of the solvophobic and the electrostatic interactions in the stereoselectivity of these systems. The overall agreement and the differences are discussed.
Synthesis of functionalized non‐natural amino acid derivatives via amidoalkylation transformations
Authors: LEGALL PHILIPPE , SAWHNEY KAILASH N. , CONLEY JUDITH D. , KOHN HAROLD
Published: August 2010

Synthetic routes have been developed for the preparation of functionalized amino acid derivatives in which the α‐substituent at carbon 2 is either an aromatic or a heteroaromatic group. The α‐substituent was introduced using an amidoalkylation reaction using boron trifluoride etherate and proceeded in moderate yield with excellent regioselectivity. This protocol permitted the employment of the acid sensitive heterocycles: pyrrole, benzofuran, and indole. The scope and limitations of this procedure have been evaluated.
s‐cis, s‐trans Isomerism about the Ala‐Pro peptide bond in Nα‐phospho‐ and Nα‐phosphono‐l‐alanyl‐l‐prolines by 31P n.m.r.
Authors: GALARDY RICHARD E. , GOLI UMESH B. , TEATER E. CARROLL , GROBELNY DAMIAN
Published: August 2010

Nα‐(Phenethylphosphono)‐l‐alanyl‐l‐proline 1, a potent inhibitor of angiotensin converting enzyme, exhibits two 31P n.m.r. resonances (intensity ratio one to one), which exchange with a constant (k1) of about 1 s−1 and a free energy of activation ΔG*=∼ 20kcal/mol at 23° in deuterated dimethylsulfoxide. Two resonances in exchange are also observed in deuterium oxide at pH 7.5. Thus the exchanging 31P resonances report the s‐cis, s‐trans conformational equilibrium about the alanyl‐proline peptide bond. Similar results were observed with Nα‐[(O‐phenyl)‐phenethylphosphono]‐l‐alanyl‐l‐proline 2. Nα ‐(O‐phenylphospho)‐l‐alanyl‐l‐proline, 3 Nα‐(O, O′‐diphenylphospho)‐l‐alanyl‐l‐proline 4, and Nα‐[2‐(2‐oxo‐1, 3, 2‐dioxaphosphiranyl)]‐l‐alanyl‐l‐proline 5 in deuterated dimethyl sulfoxide, deuterium oxide, and deuterochloroform. A 13C n.m.r. spectrum of 5 confirmed the presence of s‐cis and s‐trans resonances for the proline carbons in the same intensity ratio observed by 31P n.m.r.
Allysine peptides and derivatives
Authors: DÖLZ R. , HEIDEMANN E.
Published: August 2010

Allysine, which is synthesized enzymatically in vivo starting from lysine, is a very important crosslink precursor in proteins. We describe the chemical synthesis of allysine derivatives starting from 3,4‐dihydro‐2H‐pyran. Two independent synthetic routes to prepare allysine peptides and derivatives are presented. The synthesized compounds are characterized by spectroscopic methods including 13C‐n.m.r. The reactivity of the aldehyde function is shown to be extremely high. An unexpected nucleophilic attack of the allysine amide nitrogen upon the aldehyde group is described. This ring‐closure reaction is not expected to take place in native collagen; however, denatured peptides containing allysine may react similarly as the model peptides presented in this paper.
Protein conformational changes induced by guanidine at predenaturational concentrations
Authors: BISMUTO E. , IRACE G.
Published: August 2010

The nature of the molecular event that apomyoglobin undergoes at predenaturational concentrations of guanidine has been investigated by means of steady‐state and multifrequency phase and modulation fluorometry. The results have been compared to those observed for liver alcohol dehydrogenase. From these studies has been hypothesized a different susceptibility of the distinct elements of secondary, super‐secondary, and tertiary structure towards the denaturing action of guanidine at predenaturational concentrations.
On the receptor binding site of relaxins
Authors: BÜLLESBACH E.E. , SCHWABE C.
Published: August 2010

Relaxin plays a critical role in viviparity and has recently been implicated as a hormone of oviparity as well. In most mammals relaxin causes the widening of the birth canal during parturition and suppresses uterine motility during pregnancy. Relaxins isolated from several species have shown a great deal of sequence variability, and speculations regarding a putative receptor interaction site have, as a consequence, varied considerably. The isolation of skate relaxin in combination with our chemical modification data enable us to suggest a unique site for the interaction of relaxin with its uterine and symphyseal receptors.
2‐Phenylethyl ester and 2‐phenylethyl amide derivative analogues of the C‐terminal hepta‐ and octapeptide of cholecystokinin
Authors: FULCRAND PIERRE , RODRIGUEZ MARC , GALAS MARIE‐CHRISTINE , LIGNON MARIE‐FRANCOISE , LAUR JEANINE , AUMELAS ANDRE , MARTINEZ JEAN
Published: August 2010

Syntheses of analogues of the C‐terminal octa‐ and heptapeptide of cholecystokinin are described. These analogues were obtained by replacing the C‐terminal phenylalanine residue by 2‐phenylethyl alcohol or by 2‐phenylethylamine derivatives and by replacing the tryptophan residue by a d‐tryptophan. The CCK‐derivatives were tested for their ability to inhibit binding of labeled CCK‐8 to rat pancreatic acini and to guinea pig brain membranes, and for their action on stimulation of amylase release from rat pancreatic acini. Some of these derivatives appeared to exhibit only part of the CCK‐activity on amylase release, the d‐Trp analogues behaving as CCK‐antagonists.
Interaction of basic extension peptide fragments of adrenodoxin precursor with phospholipid vesicles
Authors: AOYAGI* HARUHIKO , LEE SANNAMU , NAKAMURA HIROSHI , PARK NAM GYU , KATO TETSUO
Published: August 2010

Two extension peptide fragments PA1‐4 and PA17‐32, which correspond to the residues 1–14 and 17–32, respectively, of adrenodoxin precursor, were synthesized by the solution method to find a sequence necessary for the import of the precursor into mitochondria. Biological assay showed that PA1‐14 inhibited the import of two mitochondrial enzyme precursors, but PA17‐32 showed no inhibition, indicating that the N‐terminal sequence has important information for import. CD spectra of the peptides demonstrated that PA1‐14 formed α‐helical structure in Tris‐HCl buffer (pH 7.4) containing acidic phospholipid liposomes. Furthermore, PA1‐14 induced the moderate leakage of carboxyfluorescein from phospholipid vesicles. The relationship between the structure and function of the peptides is discussed.
Structural and functional integrity of specificity and catalytic sites of trypsin
Authors: GRÁF LÁSZLÓ , HEGYI GYÖRGY , LIKÓ ISTVÁN , HEPP JÓZSEF , MEDZIHRADSZKY KÁLMÁN , CRAIK CHARLES S. , RUTTER WILLIAM J.
Published: August 2010

The aspartic acid residue at the bottom of the substrate‐binding pocket of trypsin was replaced by glutamic acid through site‐directed mutagenesis. The wild‐type (Asp‐189) and mutant (Glu‐189) trypsinogens were expressed in E. coli, purified to homogeneity, activated by enterokinase. and tested on a series of fluorogenic tetrapeptide substrates. The substrates were of the general formula succinyl‐Ala‐Ala‐Pro‐X‐AMC, where AMC is 7‐amino‐4‐methylcoumarin and X is Lys. Arg. or Orn (ornithine). As compared to Asp‐189 trypsin. the activity of Glu‐189 trypsin on lysyl and arginyl substrates decreased by 3‐4 orders of magnitude while its Km values did not significantly change. Lengthening the side‐chain of Asp‐189 by one methylene group could not be compensated for by shortening the side‐chain of the substrate, since Glu‐189 trypsin had no measurable activity on the ornithyl substrate. The replacement of Asp‐189 with glutamic acid at the base of the substrate‐binding pocket of trypsin appears to distort the structure of the critical transition‐state complex. This could happen by disrupting interactions normally associated with Asp‐189, and by altering the relative position of the scissile peptide bond in the active site of the enzyme.
VI. Synthesis, conformation, and ion‐binding of two bicyclic nonapeptides *
Authors: ZANOTTI G.C. , CAMPBELL B.E. , EASWARAN K.R.K. , BLOUT E.R.
Published: August 2010

Two related homodetic bicyclic nonapeptides (cyclo Glu‐X‐Pro‐Gly‐Lys‐X‐Pro‐Gly)‐cyclo (lγ→ 5ɛ), X = Ala(BCP2), X = Leu(BCP3) have been synthesized using conventional solution phase methods involving mixed anhydride coupling reactions starting with appropriately protected naturally occurring amino acids. The conformation and ion binding properties of BCP2 have been studied by nuclear magnetic resonance and circular dichroism techniques. The results of these studies have been compared to those of BCP3. The presence of Ala caused both Ala‐Pro bonds to be trans in free BCP2. This characteristic imparted subtle differences to the ion‐binding properties of BCP2 as compared to free BCP3 which has one cis Leu‐Pro bond and one trans Leu‐Pro bond.
Aggregation studies in crystals of apolar helical peptides: Boc‐Aib‐Val‐Ala‐Leu‐Aib‐Val‐Ala‐Leu‐Aib‐OMe
Authors: KARLE ISABELLA L. , FLIPPEN‐ANDERSON JUDITH L. , UMA K. , BALARAM P.
Published: August 2010

In the crystal, the backbone of Boc‐(Aib‐Val‐Ala‐Leu)2‐Aib‐OMc adopts a helical form with four α‐type hydrogen bonds in the middle, flanked by 310‐type hydrogen bonds at either end. The helical molecules stack in columns with head‐to‐tail hydrogen bonds, either directly between NH and CO, or bridged by solvent molecules. The packing of the helices is parallel, even in space group P21. Cell parameters are a = 9.837(2) Å. b = 15.565(3) Å. c = 20.087(5) Å, β= 96.42(2)°. dcalc= 1.091 g/cm3 for C46H83,N9O12· 1.5H2O·0.67CH3OH. There appears to be some hydration of the backbone in this apolar helix.
Solid phase synthesis and biological activity of mast cell degranulating (MCD) peptide: a component of bee venom
Authors: BUKU ANGELIKI , BLANDINA PATRIZIO , BIRR CHRISTIAN , GAZIS DIANA
Published: August 2010

Mast cell degranulating (MCD) peptide, a 22 amino acid residue basic peptide from bee venom, was synthesized by stepwise solid phase synthesis on a benzhydrylamine resin support. Nα‐t‐butyloxycarbonyl and benzyl type side chain protection was used. The two disulfide bridges were formed selectively by using S‐acetamidomethyl protection for the cysteine residues in positions 5 and 19 and S‐methylbenzyl protection for the cysteine residues in positions 3 and 15. Crude synthetic MCD peptide was obtained following deprotection and cleavage from the resin by the low/high HF method. The peptide was isolated in pure form by ion exchange chromatography and gel filtration. The final product has physical, chemical, and biological properties identical with those reported for the natural product. The synthetic strategy utilized for MCD peptide will facilitate the availability of structurally similar analogs for evaluating antihistaminic and anti‐inflammatory activities.
Evidence for an initial fast nucleation process in the folding of human carbonic anhydrase I
Authors: BERGENHEM NILS , CARLSSON UNO , KARLSSON JAN‐ÅKE
Published: August 2010

Kinetic studies of the folding of carbonic anhydrase have indicated the occurrence of various conformational intermediates. Human carbonic anhydrase I contains a single cysteine residue, Cys‐212, which in the native state is unavailable for alkylation. In the unfolded state, it can be specifically modified with iodoacetate. In this study the accessibility of Cys‐212 in human carbonic anhydrase I to iodo[2‐14C]acetate during the refolding process has been investigated. It is shown that Cys‐212 is hidden to the alkylating agent as soon as the refolding is initiated. Since Cys‐212 is located in the extensive β‐structure passing through the enzyme, it appears that the Cys‐containing β‐strand is part of a rapidly formed nucleation center created during the folding process. This β‐strand (No. 7) together with its neighboring β‐strand (No. 6) constitute the most hydrophobic regions of the enzyme. Because hydrophobic contacts are considered to be important in predicting nucleation sites, these β‐strands probably partake in the formation of the nucleation center. These β‐strands are also partly involved in the bottom region of the active site cavity, indicating that this region is formed during the initial folding events. As a result of this study it was also observed that 2‐mercaptoethanol is a potent inhibitor of the enzyme with a K1= 26 μM at pH 8.0.
Synthesis of derivatives of phosphonic glutathione analogs*
Authors: WITKOWSKA H. EWA , WASIELEWSKI CZESLAW
Published: August 2010

Five analogs of S‐t‐butyl glutathione containing phosphonic analogs of glycine and glutamic acid were obtained by standard procedures of MA activation in solution. Simultaneous deprotection of phosphonic, carboxylic and amino groups was achieved in the silylation reaction.
Structural studies of leucinostatin A and its Boc‐Aib‐Leu‐Leu‐Aib‐OMe tetrapeptide fragment
Authors: VERTUANI G. , FALCOMER C. , BOGGIAN M. , POCHETTI G. , CERRINI S. , RICCI M. , ROSS C. , SCATTURIN A.
Published: August 2010

The conformational behaviour of the peptide antibiotic leucinostatin A has been studied in solvents of different polarity using circular dichroism (CD) and infrared (i.r.) absorption. I.r. studies provided evidence of an intramolecularly hydrogen‐bonded structure in CDC3 while CD studies suggested a helical conformation in leucinostatin A in lipophilic solvents. The tetrapeptide Boc‐Aib‐Leu‐Leu‐Aib‐OMe, a fragment of leucinostatin A, was also studied both in solution and in solid state using X‐ray diffraction. The crystal structure shows that the peptide backbone folds into a right‐handed 310‐helical conformation stabilized by two intramolecular 4 → 1 hydrogen bonds. The spectroscopic analysis in solution is consistent with the conformation found in solid state. Crystal data: orthorhombic, P212121, a= 23.583, b= 15.791, c= 9.554 Å Z = 4; the final R and Rw are 0.090 and 0.126, respectively.
Effect of sulfate position on rnyotropic activity of the gastrin/CCK‐like insect leucosulfakinins
Authors: NACHMAN RONALD J. , HOLMAN G. MARK , HADDON WILLIAM F. , VENSEL WILLIAM H.
Published: August 2010

Leucosulfakinin and leucosulfakinin‐II are sulfated insect neuropeptides with homology to human gastrin II and cholecystokinin. Biological evaluation of the analog [Ser(SO3H)2, Tyr5]leucosulfakinin‐II, two cholecystokinin analogs containing key leucosulfakinin amino acid replacements, and a Tyr(SO3H) position‐analog series has demonstrated that while the presence of a sulfate group is critical to leucosulfakinin myotropic activity, its position is not. Most strikingly, the analog [Tyr (SO3H)5,Phe6,Nle9]leucosulfakinin, in which the sulfate moiety is shifted one position towards the N‐terminus relative to the parent peptide, retains a very significant 38% of the parent's threshold activity. A shift in the location of the sulfate group by two to five positions toward the N‐terminus led to reduction of potency to a significant but low plateau of activity, whereas a shift by more than one position towards the C‐terminus led to a complete loss of activity. The introduction of a single residue (Arg) in the 7 position of CCK‐8 transforms this inactive mammalian hormone into a myotropically‐active leucosulfakinin analog on the isolated cockroach hindgut.
Synthesis and central nervous system actions of thyrotropin‐releasing hormone analogs containing a 1‐substituted 2‐oxoimadazolidine moiety*
Authors: MAEDA HIROSHI , SUZUKI MAMORU , SUGANO HIROSHI , YAMAMURA MICHIO , ISHIDA RYUICHI
Published: August 2010

A series of thyrotropin‐releasing hormone (TRH) analogs in which the pyroglutamic acid residue was replaced by (S)‐2‐oxoimidazolidine‐4‐carboxylic acid (Oic‐OH) and the related derivatives was prepared, and the central nervous system (CNS) actions were examined. Of these, 1‐benzyl‐Oic‐His‐Pro‐NH2(2c) showed the most potent activities, which were 1.5‐8 times greater than those of TRH. Moreover, the thyrotropin (TSH)‐releasing activity of 2c was about 1/16 times weaker than that of TRH.
Synthesis, characterization and inhibitory activities of (4‐N3[3,5‐3H]Phe10)PKI(6–22)amide and its precursors: photoaffinity labeling peptides for the active site of cyclic AMP‐dependent protein kinase
Authors: KATZ BERNICE M. , LUNDQUIST LUCY J. , WALSH DONAL A. , GLASS DAVID B.
Published: August 2010

PKI(6–22)amide is a 17 residue peptide corresponding to the active portion of the heat‐stable inhibitor of cAMP‐dependent protein kinase. The peptide is a potent (Ki= 1.6 nM), competitive inhibitor of the enzyme. The photoreactive peptide analog (4‐azidophenylalanine10)PKI(6–22)amide was synthesized in both its non‐radiolabeled and tritiated forms by chemical modification of precursor peptides that were prepared by stepwise solid‐phase synthesis. (4‐Amino[3,5‐3H]phenylalanine10)PKI(6–22)amide, the precursor for the radiolabeled arylazide peptide, was obtained by catalytic reduction of the corresponding peptide containing the 3,5‐diiodo‐4‐aminophenylalanine residue at position 10. The purified PKI peptides were analyzed by HPLC, amino acid analysis, and U.V. spectra. In the dark, (4‐azidophenylalanine10)PKI(6–22)amide inhibited the catalytic subunit of CAMP‐dependent protein kinase with a Ki value of 2.8 nM. The photoreactivity of the arylazide peptide was demonstrated by time‐dependent U.V. spectral changes on exposure to light. Photolysis of the catalytic subunit.(4‐azido[3,5‐3H]phenylalanine10)PKI(6–22)amide complex resulted in specific covalent labeling of the enzyme. The data indicate that this peptide is a useful photoaffinity labeling reagent for the active site of the protein kinase.
Antigenic sites on ribonuclease A Synthetic peptides corresponding to the sites 37–42 and 83–88 on ribonuclease A show immunogenicity
Authors: SAGAR AMARA J. , MAHALAKSHMI YEDITHA V. , NAGARAJ RAMAKRISHNAN , PANDIT MADHUSUDAN W.
Published: August 2010

Two hexapeptides NH2‐Lys‐Asp‐Arg‐Cys‐Lys‐Pro‐COOH and NH2‐Asp‐Cys‐Arg‐Glu‐Thr‐Gly‐COOH corresponding to the strong hydrophilic regions 37–42 and 83–88, respectively, on ribonuclease A were synthesized by solid‐phase method. These synthetic peptides showed antigenic characteristics and provided an experimental validity to the prediction made earlier, supporting the view that highly hydrophilic regions on the protein have a good correlation with their being potentially antigenic.
Serine andd‐serine position‐1 substituted angiotensin II analogues Synthesis using new 2,3,5,6‐tetrafluorophenyl active esters and biological activities
Authors: HUI KWAN Y. , HOLLERAN EUGENE M. , KOVACS JOSEPH
Published: August 2010

[Ser1], (32), [d‐Ser1]‐ (29), [Ser1, Leu8]‐ (31), and [d‐Ser1, Leu8] angiotensin II (30) were synthesized by a repetitive method in solution using new protected amino acid 2,3,5,6‐tetrafluorophenyl active esters.32and29were agonists, and31and30were specific antagonists to angiotensin II (AII) receptors determined by the rabbit aortic strip (RAS) and rat blood pressure (RBP) assays. It was found that the hydroxymethyl side chains of serine and D‐serine in position‐l has an important influence on the agonistic activity of the analogues. The pressor activities of32and29were 129 and 314%, respectively, as potent as AII. On the other hand,31and30were effective in antagonizing the AII‐induced contraction of RAS and rise in RBP.
Structure‐toxicity relationships in the amatoxin series Synthesis of S‐deoxy[γ(R)‐hydroxy‐Ile3]‐amaninamide, its crystal and molecular structure and inhibitory efficiency*§
Authors: ZANOTTI GIANCARLO , WIELAND THEODOR , BENEDETTI ETTORE , DI BLASIO BENEDETTO , PAVONE VINCENZO , PEDONE CARLO
Published: August 2010

The amatoxins, highly toxic components of Amanita mushrooms, strongly inhibit the DNA‐dependent RNA polymerase II (or B) in eukaryotic cell nuclei. For optimal binding to the enzyme a γ‐hydroxyisoleu‐cine side chain in the 3‐position is important as in γ‐amanitin (compound l), where the OH‐group is bound in the [S]‐configuration. Amanullin, a non‐toxic component, having an oxygen‐free isoleucine side chain no. 3, exhibits an inhibitory effect on RNA polymerase II about two orders of magnitude smaller than that of γ‐amanitin. An equal, relatively weak, inhibitory effect has previously been found with the synthetically obtained Ile3‐analog 7. In the present paper the synthesis of an analog (2) bearing a γ‐hydroxyl group in the isoleucine side chain is described. The compound was found to have about the same inhibitory effect on RNA polymerase II from Drosophila embryos as amanullin and the Ile3‐analog 7. Structure analysis by X‐ray diffraction revealed that the hydroxyl group at the ‐carbon atom of side chain‐3 has the [R]‐configuration, the new analog thus being ‐deoxo[()‐hydroxy‐[Ile3]‐amaninamide. It follows that the [S]‐configuration of this chiral center is a prerequisite to maximal toxicity. Crystallographic data demonstrating great similarity between the peptide backbones of the new analog and those of natural amatoxins are given.
Synthesis of a biotinylated, iodinatable, and photoactivatable probe for angiotensin receptors
Authors: SEYER RENE , AUMELAS ANDRE , TENCE MARTINE , MARIE JACKY , BONNAFOUS JEAN‐CLAUDE , JARD SERGE , CASTRO BERTRAND
Published: August 2010

We propose here a biotinyl‐aminohexanoyl‐[Ala1, Phe(4N3)8]angiotensin II analog as a radioiodinatable and photoactivatable probe for covalent labeling, detection and isolation of angiotensin receptors. A combination of solid phase and minimum‐protection segment‐coupling strategy using hexafluorophosphate of (benzotriazol‐l‐yloxy)tris(dimethylamino)phosphonium (BOP) as a coupling reagent is proposed for the synthesis of this probe. Optimized yields were obtained by HPLC monitoring of all reactions. A complete n.m.r. study suggests an extended conformation of this molecule, allowing a simultaneous recognition of receptor and avidin. The probe binds with high affinity (Kd= 2 nM) to angiotensin II receptors from rat liver membranes.
cDNA sequence of neuroendocrine protein 7B2 expressed in beta cell tumors of transgenic mice
Authors: MBIKAY MAJAMBU , GRANT SETH G.N. , SIROIS FRANCINE , TADROS HAIDY , SKOWRONSKI JACEK , LAZURE CLAUDE , SEIDAH NABIL G. , HANAHAN DOUGLAS , CHRÉTIEN MICHEL
Published: August 2010

The cDNA for a widely distributed neuroendocrine protein called 7B2 has been cloned from beta cell tumors of transgenic mice and sequenced. As deduced from the cDNA sequence, 7B2 is a secretory protein of 186 amino acids, nearly identical to its human and porcine homologs. The presence of several pairs of basic residues in the carboxyl terminal portion of the protein suggests that 7B2 can undergo proteolytic maturation in secretory granules and thus generate potential bioactive peptides. 7B2 mRNA is about 1.5 kilobase long and is apparently transcribed from a single gene per haploid genome. The use of tissue‐specific promoters to express oncogenes in rare cell types of transgenic mice is a powerful tool for immortalization and expansion of these cells, and it facilitates the isolation and the study of rare proteins such as 7B2.
Primary structure of elephant pituitary prolactin
Authors: LI CHOH HAO , OOSTHUIZEN M.M.J. , CHUNG DAVID
Published: August 2010

Tryptic digests of elephant pituitary prolactin (elePRL) were separated by reverse phase high performance liquid chromatography (HPLC) and paper electrophoresis. From the amino acid composition, the amino acid sequencing of selected peptides, and from their alignment with expected tryptic peptides from ovine prolactin (oPRL), the primary sequence of elePRL is proposed.
Rapid racemization of optically active amino acids by microwave oven‐based heating treatment
Authors: CHEN SHUI‐TEIN , WU SHIH‐HSIUNG , WANG KUNG‐TSUNG
Published: August 2010

Rapid racemization of optically active amino acids has been developed in a Teflon vial reactor by using the commercial microwave oven as heating source. Compared with the conventional methods of racemization, there are two main advantages: (I) The whole process of racemization by this method can be completed within 2 min. (II) It can prevent the decomposition of some labile amino acids at high temperature.
Synthesis and biological activity of substance P analogues
Authors: HARO ISABEL , TORRES JOSEP LLUÍS , VALENCIA GREGORIO , GARCÍA‐ANTÓN JOSÉ MARÍA , REIG FRANCISCA
Published: August 2010

The synthesis of the hexapeptide [Glu6]SP6‐11and its glycosylated analogue at the Glu6γ‐carboxyl position by solution procedures according to several strategies is discussed. The biological activity of SP, [GIu6]SP6‐11(VI) and [Glu(β‐d‐Glcp)6]SP6‐11(VIII) have been determined and compared to SP by the GPI and RVD assays. The introduction of a β‐d‐glucopyranosyl moiety at the sixth position of the [Glu6]SP6‐11did not affect to a great extent thein vitroactivity pattern of the parent hexapeptide.