bluuebunny/crossref_metadata_2025 huggingface dataset:
PURIFICATION AND RECEPTOR BINDING PROPERTIES OF COMPLEXES BETWEEN LUTROPIN AND MONOVALENT ANTIBODIES AGAINST ITS α SUBUNIT
Authors: PIERCE JOHN G. , BLOOMFIELD GLENYS A. , PARSONS THOMAS F.
Published: August 2010

A complex between bovine lutropin (LH) and monovalent antibodies (Fab fragments) directed against its α subunit, which is common to the glycoprotein hormones, has been purified by gel filtration and chromatography on concanavalin A‐Sepharose. The complex is heterogeneous with respect to molecular size; 70–80% of the hormone is complexed with either two or three Fab fragments. The LH‐Fab α complexes retain only about 13% receptor binding activity as compared to LH when measured in a radioligand receptor assay in which the radiolabeled ligand is human choriogonadotropin. (Use of the human hormone as labeled ligand permits direct measurement of competition between receptor and the bovine complex because the α portion of the human hormone does not cross react significantly with antibodies directed against bovine α subunits.) Complex formation does not lead to dissociation of the lutropin into its subunits, as shown with a homologous LH‐β immunoassay which distinguishes free β subunit from intact LH. Complexing of LH with Fab‐α fragments also causes little or no change in the affinity of the hormone's β subunit for anti‐LH‐β antibodies indicating that significant changes in β subunit conformation did not occur. The data show that at least two well‐separated antigenic regions on the α subunit are exposed to the surface in the intact hormone. They are also in agreement with the proposal that the loss of binding activity to receptor is due to steric effects rather than to changes in conformation or dissociation, and that there may be sites on the α subunit which interact directly with the receptor.
SOLID‐PHASE ACTIVE SITE INHIBITORS OF PROTEINASES
Authors: LIGHT ALBERT , PANDE CHANDRA S. , LIEPNIEKS JURIS J.
Published: August 2010

Phenylalanine chloromethyl ketone covalently attached to porous glass beads was synthesized to serve as a solid‐phase active site directed inhibitor of chymotrypsin‐like proteolytic enzymes. The solid‐phase reagent inhibited 23 nmol of bovine chymotrypsin per gram of glass and covalently bound 30 nmol of protein per gram of glass. Sepharose‐bound lysine chloromethyl ketones were synthesized to serve as inhibitors of trypsin‐like enzymes. Sepharose‐MethionylLysyl chloromethyl ketone inactivated and bound about 6.5 nmol of enzyme per ml of settled gel. In a preliminary experiment, a cyanogen bromide cleavage of the methionine residues showed that it should be possible to release all peptides but the peptide containing the active‐site histidine. The immobilized trypsin was also reduced, carboxymethylated and digested with chymotrypsin. The potential of the solid‐phase approach is in the isolation of a specific serine proteinase and in the sequence determination of residues surrounding the active‐site histidine.
AN ANALYSIS OF SIDE‐CHAIN CONFORMATION IN PROTEINS*
Authors: BHAT T. N. , SASISEKHARAN V. , VIJAYAN M.
Published: August 2010

The crystal structures of a number of globular proteins are currently available. An analysis of the distribution of side‐chains among different allowed conformations in these proteins has been carried out. The observed conformations of individual residues are discussed on the basis of well‐known stereochemical criteria. The population distribution of side‐chains in different allowed regions in conformational space can be explained largely on the basis of simple steric considerations. In addition to examining the conformational behaviour of individual residues, some population distributions of conformational angles of general interest involving groups of residues have also been analyzed.
DIABETOGENIC ACTION OF HUMAN GROWTH HORMONE Synthesis and Activity ofC‐terminal Fragments
Authors: WADE JOHN D. , NG FRANK M. , BORNSTEIN JOSEPH
Published: August 2010

Three peptides corresponding to the C‐terminal region of human growth hormone have been synthesized by the solid‐phase method: HGH‐(177–191), HGH‐(178–191) and HGH‐(179–191). The diabetogenic activities of these synthetic peptides are reported. The data indicate that extension of HGH‐(179–191) at its NH2‐terminus is required for in vivo activity. The reduced and S‐carbamido‐methylated form of HGH‐(177–191) was also active, indicating that the disulphide bond is possibly not a prerequisite for biological activity.
KINETIC STUDIES ON HUMAN CYSTEINYL‐t RNA SYNTHETASE
Authors: CHANG GU‐GANG , PAN FOO , DENG PING‐NAN , WU BIH‐YUEH
Published: August 2010

The detailed pH and temperature kinetics of human term placenta cysteinyl‐tRNA synthetase (EC 6.1.1.16) were studied. The ATP‐PPi exchange reaction catalyzed by the cysteinyl‐tRNA synthetase was highly dependent on temperature, pH, and ionic strength. The Arrhenius plot at temperatures between 5° and 40° was linear, giving an activation energy of 19 ± 2.5 Kcal/mol. The pH dependence of the kinetic parameters Km and Vmax was investigated. Apparent pKa value of 6.4 was observed in the pH‐dependence of Vmax/Km plot. The pH versus Vmax plot showed two apparent pKa values of about 5.8 and 7.8. Van't Hoff's enthalpies were used to differentiate the nature of the possible groups responsible for the ionization. These results are valuable for the selection of chemical modifying reagents in characterizing the amino acid residues involved in substrate (nucleotide) binding or catalysis.
STUDIES ON THE COUPLING RATES IN LIQUID‐PHASE PEPTIDE SYNTHESIS USING COMPETITION EXPERIMENTS
Authors: MUTTER MANFRED
Published: August 2010

Competition experiments were carried out in order to determine relative reaction rates, Vrel for the peptide‐forming step. Using the liquid‐phase method of peptide synthesis a model system was developed to investigate the influence of coupling method, amino component, derivatization and solvent upon Vrel. According to a standard procedure, Vrel for all 20 commonly occurring amino acids were established. A strong dependence of Vrel upon steric effects of the coupling components was found. The data obtained may serve as a guideline for optimizing the reaction conditions in peptide synthesis.
HUMAN β‐ENDORPHIN Immunoreactivity of Synthetic Analogs with Various Chain Lengths
Authors: CHANG WEN‐CHANG , YEUNG HIN‐WING , LI CHOH HAO
Published: August 2010

Immunoreactivity of synthetic human β‐endorphin analogs with various chain lengths has been examined using a specific radioimmunoassay. It was found that β‐endorphin‐(1–21) and analogs of shortened chain exhibit no immunoreactivity, whereas β‐endorphin‐(1–15) possesses significant in vitro opiate activity. It appears that immunoreactivity of β‐endorphin resides in the COOH‐terminal segment of residues (22–31). The data also show the lack of correlation between opiate and immunological activities of β‐endorphin.
DISTANCE CONSTRAINTS ON MACROMOLECULAR CONFORMATION
Authors: CRIPPEN GORDON M.
Published: August 2010

Many physico‐chemical studies are made on proteins to determine something of their solution conformation. For example the coat protein of Tobacco Mosaic Virus has been subjected to more non‐crystallographic experimental studies to determine its native conformation than perhaps any other protein. Yet the sum of the experimentally determined constraints on its tertiary structure are surprisingly inadequate to fix its conformation. We are able to detect and remove minor inconsistencies in the data and then calculate a sampling of conformations consistent with all the data, which differ among themselves by r.m.s. deviations of the respective interresidue distances ranging from 5.7 Å to 15.8 Å. Some individual interresidue distances differ by as much as 50 Å from structure to structure. In order to restrict the range of possible conformations to something corresponding to the errors in a 10 Å resolution X‐ray crystal structure, chemical and spectroscopic studies will have to be much more detailed than anything done to date. Our calculations appear to be useful in deciding which further experiments would be most productive.
X‐RAY STUDY ON HOMO‐OLIGOPEPTIDES t‐BOC‐(d‐Ala)7‐OMe AND HCI·H(d‐Ala)7‐OMe
Authors: RUEDA D.R. , SPADON P. , BONORA G.M. , ZANOTTI G.
Published: August 2010

Observations of extended peptide chains, whose direction is perpendicular to the fiber axis (cross‐β‐structures) have hitherto been confined to fibrous proteins and to some synthetic polydisperse polypeptides of rather low molecular weight. This structure has now been found in some monodisperse linear homooligopeptides with aliphatic hydrocarbon side chains. X‐ray diffraction photographs of t‐Boc‐(d‐Ala)7‐OMe and HCl·H(d‐Ala)7‐OMe show characteristic reflections for this form. In addition, the good orientation of suitably prepared specimens has enabled a fairly complete determination of the unit cell of the heptapeptides to be made. t‐Boc‐(d‐Ala)7‐OMe and HCl·H(d‐Ala)7‐OMe molecules are packed in a monoclinic lattice with a = 4.80, b = 34.5, c = 5.82 Å, β = 65.6° and a = 4.80, b = 29.5, c = 5.82 Å, β = 65.6°, respectively. It has not been possible to establish whether the arrangement of the chains within the sheets is parallel or antiparallel.
PREDICTED SECONDARY STRUCTURES OF FOUR PENICILLIN BETA‐LACTAMASES AND A COMPARISON WITH TWO LYSOZYMES
Authors: MOEWS PAUL C. , KNOX JAMES R.
Published: August 2010

We have predicted the secondary structures of four beta‐lactamases /Bacillus cereus, Bacillus licheniformis, Staphylococcus aureus, and Escherichia coli R‐TEM) by the statistical method of Chou & Fasman as well as by the information theory method of Garnier et al. The secondary structures of all four beta‐lactamases are of the alpha/beta type (Levitt & Chothias's nomenclature), with helices at N‐ and C‐termini. There are about eight short regions each of alpha‐helical (30–50%) and beta‐strand (10–20%) structure separated by about 20 reverse turns. The conformation of the Gram‐positive and Gram‐negative beta‐lactamases are generally similar although a few differences are predicted between the S.aureus and E.coli structures. Surprisingly, the two bacilli structures differ significantly in three short regions. In all four enzymes the region near the catalytically‐implicated tyrosine has similar secondary structure. The secondary structure of hen egg white lysozyme, a penicillin‐binding enzyme, as well as T4 phage lysozyme, has similarities to the N‐terminal half of the penicillin‐destroying beta‐lactamases.
USE OF CYCLOPENTYL ESTER PROTECTION FOR ASPARTIC ACID TO REDUCE BASE CATALYZED SUCCINIMIDE FORMATION IN SOLID‐PHASE PEPTIDE SYNTHESIS
Authors: BLAKE JAMES
Published: August 2010

A study was conducted to determine the effect of amino acid sequence and aspartyl protecting group on the rate of base catalyzed succinimide formation in the solid‐phase synthesis of aspartyl peptides. The peptides H‐Ala‐Asp‐Gly‐Phe‐OH and H‐Ala‐Asp‐Leu‐Phe‐OH were synthesized by the solid‐phase method with cyclopentyl or benzyl protection for the β‐carboxyl of aspartic acid. The results showed that the cyclopentyl ester was notably less susceptible to succinimide formation by treatment with tertiary amine than was the benzyl ester, and that the difference could have significant consequences for the synthesis of large peptides which contain reactive sequences such as Asp‐Gly.
SYNTHESES AND CHARACTERIZATION OF TETRA‐γ‐BENZYL‐l‐GLUTAMATES
Authors: OKAHASHI KENJI , IKEDA SHOICHI
Published: August 2010

Monodisperse tetra‐γ‐benzyl‐l‐glutamates are prepared by coupling two kinds of dimers by means of dicyclohexylcarbodiimide and then converting a terminal group into the other. An o‐nitrophenylthio group is used for blocking the amino terminal, and a p‐nitrophenyl ester or an ethylamide substitutes the carboxyl end. The amino‐terminal‐deblocked tetramer, HCl,H‐[Glu(OBzl)] 4‐NHEt, is also prepared. Infrared spectra in nujol mull indicate that all the tetramers are in the β‐structure of antiparallel chains in the solid state. Circular dichroism spectra show that the amino‐terminal deblocked tetramer keeps the β‐conformation in ethylene dichloride, while it has the solvated s̀‐conformation in trifluoroethanol and in dioxane.
QUANTITATION OF SOME AMINO‐TERMINAL RESIDUES IN PROTEINS USING 3H‐LABELLED DANSYL CHLORIDE AND 14C‐LABELLED AMINO ACIDS
Authors: FLENGSRUD RAGNAR
Published: August 2010

A method for quantitation of amino‐terminal residues in proteins is presented. The method is a modification of a double isotope‐labelling technique, using 3H‐labelled dansyl chloride and 14C‐labelled amino acids as internal standards. The method is demonstrated on human fibrinogen, horse myoglobin and on mouse myoloma IgA. A linear relationship between the ratio 3H/14C in the separated amino‐terminal amino acid of the protein and the amount of protein added in the labelling mixture was obtained with standard deviations of ±7.4%, ±3.4% and ±10.3%, respectively. An application of the method is demonstrated by measuring the increase in amino‐terminal glycine in fibrinogen following the proteolytic action of thrombin. The method seems to be useful when 0.1 nmol or more of protein is used.
STRUCTURAL BIOCHEMISTRY 17. Synthesis of Tryptophan Dipeptides*
Authors: PETTIT GEORGE R. , KRUPA TIMOTHY S. , REYNOLDS REESE M.
Published: August 2010

Boc‐Trp‐Leu hydrazide has been found to significantly inhibit growth of the L‐1210 and P388 murine lymphocytic leukemias. A series of related tryptophan dipeptides were prepared and found to be devoid of such activity. The tryptophan dipeptides were most conveniently obtained employing the DCCI‐NHS peptide bond‐forming procedure.
REDUCTION OF β‐ENDORPHIN CONTENT IN THE RAT PITUITARY AFTER DEHYDRATION AND ADRENALECTOMY
Authors: TSENG LIANG‐FU , O'ROURKE MAUREEN A. , LI CHOH HAO , LOH HORACE H.
Published: August 2010

Both dehydration and adrenalectomy of rats caused a significant reduction of immunoreactive β‐endorphin in the pituitary gland. β‐Endorphin in hypothalamus, however, was not altered in either dehydrated or adrenalectomized rats.
SYNTHESIS OF LYSINE‐VALINOMYCIN BY SOLID‐PHASE SEGMENT CONDENSATION
Authors: GISIN B. F. , DHUNDALE A. R.
Published: August 2010

In order to obtain a readily derivatized analog of the ionophore antibiotic valinomycin, [1‐lysine]valinomycin (Lys‐VAL) was synthesized. The compound was built up on a polystyrene support by stepwise segment condensation and was cyclized in solution. The segments used were didepsipeptides protected by the t‐butyloxycarbonyl and p‐nitrobenzyloxycarbonyl groups. Derivatives prepared by acylation of the e‐amino group of Lys‐VAL included [14C]acetyl‐Lys‐VAL, dansyl‐Lys‐VAL, palmitoyl‐Lys‐VAL and dithiodiglycoyl‐bis‐Lys‐VAL. These derivatives had a high potassium binding capacity but were in general much less active than VAL in mediating ion transport in membranes.
MOLECULAR WEIGHT OF B. SUBTILIS ALPHA‐AMYLASE DERIVED FROM CHEMICAL STUDIES
Authors: DETERA S.D. , FRIEDBERG F.
Published: August 2010

Bacillus subtilis α‐amylase was cleaved with cyanogen bromide and the amino terminal sequences of the purified products were determined. The molecular weights of the cyanogen bromide fragments were ascertained on an agarose column equilibrated with 6m guanidine hydrochloride. The molecular weights of these fragments were also calculated from their amino acid compositions. The data obtained by these methods concurred and established the molecular weight of the enzyme monomer as 48,000.
QUANTITATIVE EVALUATION OF γ‐TURN CONFORMATION IN PROLINE‐CONTAINING PEPTIDES USING13 C N.M.R.
Authors: LONDON ROBERT E.
Published: August 2010

The dependence of the 13C shift difference of proline carbons Cβ and Cγ on the dihedral angle ø has been studied using the model peptide acetyl‐d‐proline N‐methylamide. The shift difference Δβγ is shown to be correlated with the percent cis isomer about the acetylproline bond, both factors depending strongly on the degree of intermolecular hydrogen bonding. Both the fraction of trans peptide bond and the fractional γ‐turn conformation increase as the sample concentration is decreased in CDCl3. Δβγ values have been used to evaluate the fractional γ‐turn probabilities in a number of cyclic and linear peptides including thyrotropin releasing factor and bradykinin. Using this parameter, it is concluded that in bradykinin the γ‐turn probability is low in D2O and not strongly temperature dependent. In contrast, studies of a model peptide for the portion of bradykinin believed to adopt a γ‐turn conformation are consistent with an increased γ‐turn probability in less polar solvents. Data for X‐Pro‐Y peptides (Y = imino acid) indicate significantly reduced values of Δβγ, and this appears to be a useful basis for assigning the Pro Cβ resonances corresponding to this sequence.
EFFECT OF TEMPERATURE UPON THE CIRCULAR DICHROISM OF BRADYKININ
Authors: CANN JOHN R. , LONDON ROBERT E. , STEWART JOHN M. , MATWIYOFF NICHOLAS A.
Published: August 2010

Analysis of the effect of temperature on the circular dichroism spectrum of bradykinin has led to a more precise understanding of the solution conformation of the peptide. Circular dichroism and 13C n.m.r. have been used in a complementary fashion to support the picture that bradykinin spends a maximum of about 20% of its time in a partially ordered conformation featuring a γ‐turn with Pro7 as the second residue. Since the γ‐turn probability is insensitive to temperature, some other conformational effect dominated by the structure of water presumably produces the pronounced change in the circular dichroism spectrum with increasing temperature.
RAPID SEPARATION OF PROTEINS AND THEIR HIGHER‐MOLECULAR FRAGMENTS BY MEANS OF SPHERON ION‐EXCHANGES*
Authors: MIKEŠ OTAKAR
Published: August 2010

Ion‐exchange derivatives are described, of a hydrophilic rigid macroporous glycolmethacrylate gel called Spheron, suitable for rapid high‐performance liquid chromatography (HPLC) of proteins and their fragments. Their flow parameters are compared with those of ion exchange derivatives of cellulose and poly‐dextran. The conditions for work with them are described (regeneration, cycling, equilibration, column packing) as well as the construction of a simple apparatus for medium‐pressure ion exchange chromatography of proteins. The efficiency of these ion exchangers for the separation of proteins is illustrated with examples of chromatography of an artificial mixture of serum albumin, chymotrypsinogen and lysozyme. Chromatography of cyanogen bromide fragments of serum albumin and the A and B chains of oxidized insulin showed that the method can be applied in chromatography on higher molecular protein fragments. A review of all proteins, including technical enzymes, which have already been chromatographed on Spheron ion exchangers is also given. The prospects of Spheron ion exchangers for HPLC of proteins and their fragments are briefly discussed.
THE NATURE OF THE MULTIPLE FORMS OF BOVINE THIOL: PROTEIN DISULFIDE OXIDOREDUCTASE
Authors: PACE MARIO , DIXON JACK E.
Published: August 2010

Preparations of thiol:protein disulfide oxidoreductase from bovine liver were shown to be homogeneous by polyacrylamide gel electrophoresis, sedimentation equilibrium centrifugation and NH2‐terminal analysis (Carmichael et al., 1977). When the enzyme was subjected to prolonged storage at ‐20°, freeze‐thawing, or heating at 60°, at least one new protein species was observed using polyacrylamide gel electrophoresis. The new protein results from dimerization of the enzyme. The dimer consisted of two monomers held together by an inter‐molecular disulfide bond. The formation of this dimer can be reversed and partially prevented by thiols.
SYNTHESIS OF N‐ACETYL‐MURAMYL‐l‐ALANYL‐d‐GLUTAMIC‐α‐AMIDE(MDP) OR ‐α‐METHYL ESTER DERIVATIVES, BEARING A LIPOPHILIC GROUP AT THE C‐TERMINAL PEPTIDE END
Authors: LEFRANCIER P. , PETITOU M. , LEVEL M. , DERRIEN M. , CHOAY J. , LEDERER E.
Published: August 2010

We report the synthesis of nine lipophilic derivatives of N‐acetyl‐muramyl‐l‐alanyl‐d‐glutamic‐α‐amide (MDP) or ‐α‐methyl ester in which the γ‐carboxyl function of the d‐glutamyl residue is either esterified by a medium chain alcohol or substituted by an l‐alanyl residue esterified by a medium or long chain alcohol. A new method is described which easily allows one to obtain derivatives of MDP, bearing a free or substituted amino‐acyl or peptidyl residue on the γ‐carboxyl function.
CHEMICAL SYNTHESIS OF [DES(TETRAPEPTIDE B27–30), TYR(NH2)26‐B] AND [DES(PENTAPEPTIDE B26–30), PHE(NH2)25‐B] BOVINE INSULINS*
Authors: COSMATOS ALEXANDROS , FERDERIGOS NICOLAOS , KATSOYANNIS PANAYOTIS G.
Published: August 2010

Two analogs of bovine insulin, [des(tetrapeptide B27–30), Tyr(NH2)26‐B] and [des(pentapeptide B26–30), Phe(NH2)25‐B] insulin, which differ from the parent molecule in that the C‐terminal tetrapeptide and pentapeptide sequences, respectively, from the B chain have been eliminated and the newly exposed residues are amidated, have been synthesized. The [des(tetrapeptide B27–30), Tyr(NH2)26‐B] insulin shows potencies of 16.8 IU/mg by the mouse convulsion assay method and 10.8 IU/mg by the radioimmunoassay method. The [des(pentapeptide B26–30), Phe(NH2)25‐B] insulin possesses a potency of 10.5 IU/mg when assayed by the mouse convulsion method and 14 IU/mg by the radioimmunoassay technique. The potencies of these analogs are higher than the potencies of the respective non‐amidated derivatives (Katsoyannis et al., 1973, 1974). It is speculated that the gradual decline of biological activity observed as amino acid residues are eliminated from the C‐terminal region of the B chain of insulin is due to the proximity of a hydrophilic carboxyl group to the hydrophobic core of the protein molecule.
AUTOMATED SOLID PHASE SYNTHESIS OF THYMOSIN α1
Authors: WANG S. S. , MAKOFSKE1 R. , BACH A. , MERRIFIELD R. B.
Published: August 2010

An automated solid phase synthesis of thymosin α1 based on a tactic of side chain attachment to benzhydrylamine resin is described. The product obtained was shown to be identicat to the natural product isolated from calf thymus gland.
RAPID LEUCINE‐ENKEPHALIN SYNTHESIS
Authors: VILKAS ERNA , VILKAS MICHEL , SAINTNO JOËLLE
Published: August 2010

Leucine‐enkephalin, a naturally occurring pentapeptide with morphine‐like activity, has been synthesized by a fragment condensation liquid phase method using 3,3,4‐trimethyl‐Δ3‐phospholene‐1,1‐dichloride as a coupling agent. The advantages of this method are discussed.
PREPARATION AND PROPERTIES OF Nα‐9‐FLUORENYLMETHYLOXYCARBONYLAMINO ACIDS BEARING TERT.‐BUTYL SIDE CHAIN PROTECTION
Authors: CHANG CHI‐DEU , WAKI MICHINORI , AHMAD MUSHTAQ , MEIENHOFER JOHANNES , LUNDELL EDWIN O. , HAUG JAMES D.
Published: August 2010

The preparation is described of several Nα‐9‐fluorenylmethyloxycarbonylamino acids and derivatives bearing tert.‐butyl type side‐chain protection of amine, carboxyl, guanido, hydroxyl, imidazol, and sulfhydryl functionalities. Physicochemical properties of these compounds have been determined. Cleavage of the Fmoc group by various amines appears to depend on the base strength and steric hindrance. Premature deblocking of Fmoc group by amine on solid support is very slow and may be negligible under the conditions of solid‐phase synthesis.
SOLID‐PHASE SYNTHESIS OF PORCINE VASOACTIVE INTESTINAL PEPTIDE
Authors: COY DAVID H. , GARDNER JOSEPH
Published: August 2010

The 28‐residue sequence of porcine vasoactive peptide (VIP) was assembled on a benzhydrylamine resin support, cleaved by HF treatment, and purified by ionexchange and partition chromatography. In addition to the normal criteria, the homogeneity of the final material, obtained in 16% yield, was assessed by reversephase high performance liquid chromatography and the isolation and examination of cyanogen bromide cleavage fragments by the same technique. The purified VIP possessed characteristic inhibitory effects on pentagastrin‐induced gastric acid release in the dog. Upon storage, even as the lyophilized powder in vacuo, HPLC revealed the slow formation of a contaminant possibly representing deamidated peptide.
CYLIC PEPTIDES
Authors: YASUTAKE AKIRA , AOYAGI HARUHIKO , KATO TETSUO , IZUMIYA NOBUO
Published: August 2010

Two diastereomeric cyclic tetrapeptides with a sequence of cyclo (‐D (and L)‐Tyr (Me)‐L‐Ile‐L‐Pro‐L‐Leu‐) (ID and IL) have been synthesized, which are simplified analogs of a phytotoxic peptide Cyl‐2, and contain an L‐proline and an L‐leucine residue in place of an L‐pipecolic acid and a 2‐amino‐8‐oxo‐9,10‐epoxydecanoic acid residue, respectively, in Cyl‐2. The cyclization of the H‐D‐Tyr(Me)‐L‐Ile‐L‐Pro‐L‐Leu‐ONSu proceeded smoothly to give ID in excellent yield (50%), but the cyclization of H‐L‐Tyr(Me)‐L‐Ile‐L‐Pro‐L‐Leu‐ONSu gave IL in much lower yield (10%).

Based on the results of 1H‐ and 13C‐n.m.r. studies, differential N‐methylation and model buildings of ID, the backbone structure of ID has been proposed to be a unique trans‐trans‐cis‐trans conformation, the L‐Ile‐L‐Pro bond being cis.
CRYSTALLIZATION OF THE PEPTIDE HORMONE, HUMAN CHORIONIC SOMATOMAMMOTROPIN
Authors: MOFFAT KEITH
Published: August 2010

Human chorionic somatomammotropin has been crystallized from a number of solvents. One of these crystal forms offers promise for high resolution X‐ray structure analysis.
PREDICTOR FOR SULFUR‐AROMATIC INTERACTIONS IN GLOBULAR PROTEINS
Authors: MORGAN RICHARD S. , McADON JOHN M.
Published: August 2010

The predictor, Y = 2.54 + (4 Met + 2 Arg ‐ Tyr) 83/N, where Y is the number of sulfur‐aromatic interactions in a globular protein and N is the total number of amino acids in the protein, accounts for 75% of the variation in this type of interaction that occurs in 22 globular proteins whose three‐dimensional structure has been determined. We find that S‐ interactions are not random events but rather are the outcome of a competition between the dipolar groups, amides and sulfur‐containing, for ring neighbors. This outcome is strongly weighted in favor of S‐ interactions by the presence in the protein of positively charged side‐chains; it is not affected by the number of strictly non‐polar side‐chains.
CHEMICAL MODIFICATION OF METHIONINES OF RIBONUCLEASE A WITHo‐BENZOQUINONE
Authors: GUPTA MUNISHWAR N. , VITHAYATHIL PAUL J.
Published: August 2010

The accessibility of methionines in RNAase A to reaction with OBQ has been studied at highly acidic pH. The differences between the rate constants of reactions of the methionine and methionines of RNAase A with OBQ is a reflection on the limited accessibility of methionines in the protein conformation. Nevertheless, at sufficiently high OBQ concentration, all the four methionines of the enzyme can be modified. At lower concentration of OBQ, a derivative may be prepared in which a specific methionine is modified. The introduced chromophore ionizes at around pH 3 in this derivative. The derivative has partial activity towards RNA which is enhanced on addition of S‐protein.
o‐BENZOQUINONE‐A REAGENT FOR DETERMINING CONFORMATIONAL DIFFERENCES IN RELATED PROTEINS
Authors: GUPTA MUNISHWAR N. , MURTHY G.S. , VITHAYATHIL PAUL J.
Published: August 2010

OBQ reacts specifically with methionine residues of RNAase A at highly acidic pH. At pH 3, whereas RNAase A does not react, RNAase S, S‐protein and S‐peptide all react with OBQ. RNAase A derivatives in which some of the methionines have been initially modified at pH 1, further react with OBQ at pH 3. Thus, reaction with OBQ can be used to monitor structural transitions and conformational differences involving methionines in proteins.
STUDIES ON THE IMPORTANCE OF THE ASPARAGINE RESIDUE IN OXYTOCIN Synthesis and Some Pharmacological Properties of [1‐α‐Mercaptoacetic acid, 5‐isoasparagine] Oxytocin
Authors: ROY J. , JOHNSON M. , DUBIN S. , GAZIS D. C. , SCHWARTZ I. L.
Published: August 2010

[1‐α‐Mercaptoacetic acid, 5‐isoasparagine] oxytocin was synthesized to study the effects of moving the side chain carboxamide group of the amino acid residue in position 5 of oxytocin from the β to the α position. The analog has an isoasparagine residue in position 5 and the 20‐membered ring size of oxytocin is maintained by substituting cysteine in position 1 of oxytocin by α‐mercaptoacetic acid. The analog was found to possess 0.098 ± 0.002 U/mg of uterotonic activity but no milk‐ejecting, antidiuretic or rat pressor activity could be detected. The substance did not inhibit the uterotonic or milk‐ejecting activity induced by oxytocin nor the antidiuretic or rat pressor responses to the USP posterior pituitary standard. These results, together with the data available in the literature, indicate that an analog of oxytocin lacking the asparagine residue in position 5 is neither an agonist nor an antagonist. The observations may mean that the asparagine residue is critically important for the interaction of oxytocin with its receptor.
OXIDATION OF TRYPTOPHAN TO OXINDOLYLALANINE BY DIMETHYL SULFOXIDE‐HYDROCHLORIC ACID
Authors: SAVIGE WALTER E. , FONTANA ANGELO
Published: August 2010

Tryptophan is readily oxidized to oxindolylalanine (2‐hydroxytryptophan) in good yield on treatment in acetic acid solution with a mixture of dimethyl sulfoxide (DMSO) and concentrated aqueous HCl at room temperature. Other sulfoxides can be used in combination with HCl; for example, methionine sulfoxide reacts with an equimolar amount of tryptophan to give high yields of methionine and oxindolylalanine. Methionine and cysteine are quantitatively oxidized by DMSO/HCl to methionine sulfoxide and cystine, respectively.

The tryptophan containing peptides LRF (luteinizing hormone‐releasing factor), somatostatin, valine‐gramicidin A and ACTH1–24were each treated with the DMSO/HCl reagent in acetic acid solution and the corresponding oxindolylalanine‐derivatives isolated in over 90% yield after chromatography. The identity and purity of the derivatives were established on the basis of ultraviolet spectral characteristics and quantitative amino acid analysis of the oxindolylalanine content of acid hydrolyzates of the oxidized peptides with 3N‐p‐toluenesulfonic acid at 110° for 24 h.

The results indicate that modification of tryptophan peptides with DMSO/HCl provides a useful procedure, which seems superior to previously used reagents. In addition, the method could be well applied to other indoles of biological and pharmacological interest.
SOLID‐PHASE SYNTHESIS OF HUMAN PANCREATIC POLYPEPTIDE
Authors: MEYERS CHESTER A. , COY DAVID H.
Published: August 2010

The 36‐residue sequence of human pancreatic polypeptide (HPP) was synthesized by solid‐phase methodology, cleaved from the benzhydrylamine resin by HF treatment, and purified by gel filtration, ion exchange, and partition chromatography. In addition to the usual criteria, the homogeneity of the final product was examined by reversed phase high performance liquid chromatography. This technique was also used to isolate and characterize the cyanogen bromide cleavage fragments, and finally to compare the synthetic product with native human, bovine, and porcine pancreatic polypeptide isolated from natural sources. The purified HPP, like native PP, was a potent inhibitor of pancreatic secretion in the dog.
SYNTHESIS OF ANALOGS OF THE SERUM THYMIC NONAPEPTIDE, “FACTEUR THYMIQUE SERIQUE” (FTS) Part II
Authors: MARTINEZ JEAN , BLANOT DIDIER , AUGER GENEVIEVE , SASAKI ANDRE , BRICAS EVANGHELOS
Published: August 2010

New analogs of FTS (Facteur Thymique Sérique), <Glu‐Ala‐Lys‐Ser‐Gln‐Gly‐Gly‐Ser‐Asn · OH, a circulating thymic factor, were prepared by replacing the amino acid residues in positions 1, 3, 4, 5, 6 and 3 and 6 together. Five other analogs of C‐terminal heptapeptide were prepared by replacing the amino acid residues in position 3 or 6. These peptides were synthesized using conventional synthesis in solution.
ROTATIONAL ISOMERISM ABOUT THE Cα‐CO BOND IN PROLINE DERIVATIVES
Authors: NAGARAJ R. , VENKATACHALAPATHI Y.V. , BALARAM P.
Published: August 2010

The 270 MHz 1H n.m.r. spectrum of benzyloxycarbonyl‐Pro‐N‐methylamide in CDCl3 is exchange broadened at 293° K. Spectral lines due to two species are frozen out at 253° K and a dynamically averaged spectrum is obtained at 323° K. A selective broadening of the Cβ and Cγ resonances in the 13C n.m.r. spectrum is observed at 253° K, with a splitting of the Cβ and Cγ resonances into a pair of lines of unequal intensity. A similar broadening of Cβ and Cγ peaks is also detected in pivaloyl‐Pro‐N‐methylamide where cis‐trans interconversion about the imide bond is precluded by the bulky t‐butyl group. The rate process is thus attributed to rotation about the Cα‐CO bond (ψ) and a barrier (ΔG#) of 14kcal mol‐1 is estimated. 13C n.m.r. data for pivaloyl‐Pro‐N‐methylamide in a number of solvents is presented and the differences in the Cβ and Cγ chemical shifts are interpreted in terms of rotational isomerism about the Cα‐CO bond.
OXIDATION OF METHIONINE RESIDUES IN BOVINE GROWTH HORMONE BY CHLORAMINE‐T
Authors: CASCONE O. , BONINO M.J. BISCOGLIO DE JIMENEZ , SANTOME J.A.
Published: August 2010

The methionine residues in bovine growth hormone were chemically modified with chloramine‐T at pH 7.4 using different chloramine‐T methionine molar ratios. Methionine 4 was shown to be the most reactive followed in decreasing order by methionines 148, 123 and 178. With a 50‐fold molar excess of chloramine‐T over methionine almost full oxidation of all the methionine residues was achieved. The oxidation of the four residues of methionine does not change the growth promoting activity of the hormone.
SMALL‐ANGLE X‐RAY SCATTERING STUDIES OF THERMALLY‐INDUCED GLOBULAR PROTEIN GELS
Authors: CLARK A. H. , TUFFNELL C. D.
Published: August 2010

Small‐angle X‐ray scattering has been applied to gels formed by heating globular proteins, in aqueous solution, above their unfolding temperatures. A number of BSA gels, previously characterised by electron microscopy, have been studied, and, by setting up theoretical models for the scattering process, the X‐ray data have been shown to be consistent with the microscope conclusions regarding network structure. It is concluded that the networks form by a linearly‐directed aggregation of unfolded, disc‐like, protein molecules, three‐dimensional geometry being achieved by occasional branching, and/or cross‐linking. Longrange inhomogeneities in network structure, easily observed by electron microscopy, and correlated with variations in pH or ionic strength, have an effect on X‐ray scattering, and hence the X‐ray method is sensitive not only to different network strand thicknesses, but to different degrees of uniformity as well.
SYNTHESIS OF CARBOXYL TERMINAL SEGMENTS OF BETA‐SUBUNIT OF HUMAN CHORIONIC GONADOTROPIN
Authors: ROLLI HANSPETER , BLASER KURT , PFEUTI CHRISTIANE , SCHNEIDER CONRAD H.
Published: August 2010

The synthesis in solution of carboxyl terminal peptide segments of the beta‐subunit of human chorionic gonadotropin is described. The protected segments include sequences 119–131, 132–137, and 138–145. The syntheses were based on a standardized liquid‐liquid extraction program for routine purification of intermediates (two‐phase method). Condensation of terminally deblocked segments afforded protected peptides 132–145, 126–145, 120–145 and 119–145. Protected peptides 126–145 and 120–145 were deprotected in liquid hydrogen fluoride and used in conjugated form for immunization of rabbits. Data on the specificity of the antibody response are reported.
HYDROPHOBIC INTERACTIONS IN THE UROKINASE ACTIVE CENTRE
Authors: CHRISTOVA EVDOKIA , YOMTOVA VIHRA , BLAGOEV BLAGOY
Published: August 2010

In order to obtain information concerning the binding site of urokinase (EC 3.4.99.26) the inhibitory action of aliphatic ammonium and amidinium ions has been studied. The comparison between the corresponding Kivalues for urokinase and trypsin shows that although the two enzymes possess a hydrophobic binding pocket of presumably equal length, they differ mainly in their behaviour towards n‐alkylammonium ions with an alkyl chain longer than four methylene groups and towards benzylammonium and amidinium ions. This is an indication of a less rigid binding site in urokinase than in trypsin.
β‐ENDORPHIN: RADIORECEPTOR BINDING ASSAY
Authors: FERRARA PASCUAL , LI CHOH HAO
Published: August 2010

An assay system is described to measure the specific binding of β‐endorphin to opiate sites (receptors) in rat brain membrane preparations using the tritiated hormone as the primary ligand. By this assay procedure, the radioreceptor activity of β‐endorphin and synthetic analogs with various chain lengths has been determined. The results suggest that both NH2‐ and COOH‐terminal sequences of the molecule are involved in the interaction of β‐endorphin with opiate receptors.
SYNTHESIS AND RADIORECEPTOR BINDING ACTIVITY OF TURKEY β‐ENDORPHIN AND DEACETYLATED SALMON ENDORPHIN
Authors: YAMASHIRO DONALD , FERRARA PASCUAL , LI CHOH HAO
Published: August 2010

Des‐acetylated salmon endorphin and turkey β endorphin have been synthesized by the solid‐phase method. Relative opiate activities in a radioreceptor binding assay are: human β‐endorphin, 100; des‐acetylated salmon endorphin, 169; turkey β‐endorphin, 94. Thus, non‐mammalian endorphins can show high activity in a mammalian assay system.
STUDIES ON THE COSUBSTRATE SITE OF PROTEASE SOLUBILIZED NADPH‐CYTOCHROME P450 REDUCTASE
Authors: LUMPER LUDWIG , BUSCH FRIEDHELM , DŽELIĆ ŠEHERAZADE , HENNING JÖRG , LAZAR TIBOR
Published: August 2010

Modification of the protease solubilized NADPH‐cytochrome P450 reductase (= NADPH‐cytochrome c reductase) at the critical SH group in the cosubstrate binding site affects KNADPHm but not V for the cytochrome c reduction. The increase of KNADPHm is dependent on the size and the charge of the substituent introduced. Substitution of the cosubstrate site SH by the CN‐, S2 O3‐ and the (N‐ethyl) succinimido group effects a 3‐, 7‐ and 23‐fold increase of KNADPHm respectively. The critical SH group in the NADPH binding region can be specifically radiolabeled by N‐ethyl (2,3‐14C) maleimide after preincubation of the reductase with unlabeled NEM in the presence of 1 mM NADP+. The selective reaction at the essential cysteine in the cosubstrate site is demonstrated by peptide mapping of the thermolytic digest and urea SDS gel electrophoresis of the cyanogen bromide fragments of the reductase. Protease solubilized NADPH‐cytochrome P450 reductase is inactivated by reagents directed to histidine, orginine and lysine residues. NADP (H) (1 mM) and 2′‐AMP (1 mM) give effective protection only for the reaction of 1, 2‐cyclohexanedione (12 mM). The functional role of the basic amino acid residues for the cosubstrate binding by the NADPH‐cytochrome P450 reductase cannot be established therefore by the modification experiments described.

The number of NADPH binding sites in the NADPH‐cytochrome P450 reductase is determined to one site/mol reductase by titration of the enzyme with NADP+ monitored by CD‐spectroscopy.
RAT BETA‐LPH, GAMMA‐LPH AND BETA‐ENDORPHIN BIOSYNTHESIZED BY ISOLATED CELLS OF PARS INTERMEDIA AND PARS DISTALIS
Authors: GIANOULAKIS C. , SEIDAH N.G. , ROUTHIER R. , CHRÉTIEN M.
Published: August 2010

Rat pars intermedia cells were incubated for 3 h with the following amino acids: (a) 35S‐methionine and 3H‐phenylalanine; (b) 3H‐valine; and (c) 3H‐valine and 3H‐lysine. Radioactive gamma‐lipotropin, beta‐lipotropin and beta‐endorphin were purified on carboxymethyl‐cellulose and characterized by polyacrylamide disc gel electrophoresis at pH 4.5, molecular weight estimation and micro‐sequencing. Rat gamma‐lipotropin was shown to differ slightly from ovine gamma‐lipotropin in its NH2‐terminal amino acid sequence, in containing no methionine and having phenylalanine at position 6, valine at positions 13 and 27, and lysine at position 20. The same variations were observed in the sequence of rat beta‐lipotropin, while rat beta‐endorphin was shown to be identical to the ovine beta‐endorphin. Following a 3‐h pulse of rat pars distalis, the cells were extracted with care to avoid beta‐lipotropin degradation by proteolytic enzymes. A peptide was purified and identified to be rat beta‐endorphin, thus demonstrating that beta‐endorphin is biosynthesized in pars distalis and is not an extraction artifact.
PHOSPHODIPEPTIDE ANALYSIS OF NONHISTONE NUCLEAR PROTEINS FROM NOVIKOFF HEPATOMA ASCITES CELLS
Authors: JONES CHRIS E. , OLSON MARK O.J.
Published: August 2010

To study the sites of phosphorylation in nuclear proteins a simple method was developed for the isolation and analysis of phosphodipeptides. Partial acid hydrolysates of unfractionated nonhistone nuclear proteins were subjected to Dowex‐1 column chromatography followed by paper electrophoresis at pH 1.8. Phosphoserine‐ and phosphothreonine‐containing dipeptides each had characteristic mobilities in the latter system. By subtractive Edman degradation these peptides were identified as having the general structure, X‐PSer or X‐PThr, where X is a nonphosphorylated amino acid. The two groups of phosphodipeptides were further purified into unique peptides by two‐dimensional paper electrophoresis at pH 3.6 and 6.5. Amino acid analysis, and thus nearest neighbor analysis, of phosphodipeptides from nonhistone nuclear proteins revealed that a heterogeneous group of amino acids was on the amino terminal side of phosphoserine residues. In contrast, phosphothreonine residues were predominantly preceded by proline.
RIBONUCLEASE INHIBITOR FROM BOVINE BRAIN*
Authors: BURTON LOUIS E. , BLACKBURN PETER , MOORE STANFORD
Published: August 2010

A ribonuclease inhibitor from bovine brain has been purified 27000‐fold by affinity chromatography on Sepharose‐RNase A with an overall yield of 46% (ca. 0.4 mg/kg tissue). The purified inhibitor gives a single band by SDS‐gel electrophoresis. By gel filtration the molecular weight is ca. 50 000; the molecular weight of the complex with bovine pancreatic RNase A is ca. 62 000, indicative of 1:1 binding on a molar basis. The inhibition of the action of RNase A on yeast RNA by the inhibitor is noncompetitive with a Kiof 7 times 10‐10 M. The protein is very similar in its properties, including amino acid composition, to the inhibitor previously isolated from human placenta.

The amount of inhibitor per g of protein in bovine brain is about one‐seventh of the value for human placenta. No difference was found in the distribution of inhibitor between white and gray matter; one‐tenth of the inhibitor present is bound to a brain ribonuclease which is released in active form after reaction with p‐hydroxymercuribenzoate. Essentially no free neutral ribonuclease activity could be detected in brain homogenates in the absence of p‐hydroxymercuribenzoate.
BIDIRECTIONAL SYNTHESIS OF [5‐ASPARTIC ACID] ARGININE VASOPRESSIN ON POLY‐N‐ACRYLYLPYRROLIDINE RESIN
Authors: SMITH CLARK W. , SKALA GERALD , WALTER RODERICH
Published: August 2010

In the proposed biologically active conformation of vasopressin at its antidiuretic receptor, the side‐chain carboxamide moiety of the 5‐position asparaginyl residue has been suggested to be an active element for the initiation of the antidiuretic response. [5‐Aspartic acid] arginine vasopressin, the analog in which the ‐NH2 portion of the primary amide has been replaced by an ‐OH group, has been synthesized and tested for some of the pharmacological activities of vasopressin. The partially protected nonapeptide intermediate was assembled bidirectionally on a poly‐N‐acrylylpyrrolidine resin. The 6‐position cysteinyl residue was attached to the resin via its side‐chain through an S‐carbamoyl linkage. First the COOH‐terminus was extended by coupling with Pro‐Arg(Tos)‐Gly‐NH2, then the NH2‐terminus was extended in a stepwise manner. [5‐Aspartic acid] arginine vasopressin was found to possess 86.5 ± 4.8 units/mg of antidiuretic potency, 17% of the parent hormone. In addition, the analog possesses rat pressor and rat uterotonic potencies of 6.93 ± 0.15 and 0.38 ± 0.03 units/mg, respectively. This result suggests that a carboxylic acid moiety on the 5‐position aspartyl residue retains sufficient steric features and hydrophilicity in common with the carboxamide moiety present in the hormone to substitute for it as an active element at the antidiuretic receptor.
SYNTHESIS, PHARMACOLOGICAL, CONFORMATIONAL, AND DYNAMIC STUDIES OF THE POTENT HORMONE ANTAGONISTS [1‐PENICILLAMINE, 4‐THREONINE]‐OXYTOCIN AND [1‐PENICILLAMINE, 2‐PHENYLALANINE, 4‐THREONINE] ‐OXYTOCIN
Authors: HRUBY VICTOR J. , MOSBERG HENRY I. , HADLEY MAC E. , CHAN W. Y. , POWELL ANDREA M.
Published: August 2010

The solid phase synthesis of [1‐penicillamine, 4‐threonine]‐oxytocin and [1‐penicillamine 2‐phenylalanine, 4‐threonine]‐oxytocin is reported. The two compounds have no in vitro milk ejecting activity and no in vivo or in vitro oxytocic activity, but both are potent antagonists in these three assay systems. In the in vitro oxytocic assay, [1‐penicillamine, 4‐threonine]‐and [1‐penicillamine, 2‐phenylalanine, 4‐threonine]‐oxytocin have pA2 values of 7.55 ± 0.04 and 7.67 ± 0.02, respectively, and both inhibit the uterine contractile response to oxytocin in nonpregnant and pregnant rats. [1‐Penicillamine, 2‐phenylalanine, 4‐threonine]‐oxytocin has a weak antipressor activity and at high doses, consistently caused a weak and transient fall in blood pressure in the rat. Carbon‐13 nuclear magnetic resonance chemical shift parameters and spin‐lattice relaxation times (T1) indicate that these two new oxytocin antagonists have very similar conformation and dynamic properties to oxytocin inhibitors which have previously been examined. These results are discussed in terms of conformational and dynamic models of oxytocin antagonism at the uterus. It is suggested that conformational restrictions at the 2‐ and 4‐positions of penicillamine‐1 analogues of oxytocin are important to antagonist activity and potency.
ENZYME‐CATALYZED FORMATION OF SEMISYNTHETIC STAPHYLOCOCCAL NUCLEASE USING A NEW SYNTHETIC FRAGMENT, [48‐GLYCINE] SYNTHETIC‐(6–49)
Authors: KOMORIYA AKIRA , HOMANDBERG GENE A. , CHAIKEN IRWIN M.
Published: August 2010

While trypsin can catalyze resynthesis of the peptide bond between fragments in the noncovalent complex of nuclease‐T‐(6–48) and nuclease‐T‐(49–149), this reaction leads to excision of Lys 49 and formation of inactive [des Lys 49]‐nuclease‐(6–149). To provide a method for making active covalent semisynthetic nuclease, we chemically synthesized the fragment of residues 6 to 49 in which lysine 48 was replaced by glycine. This peptide was made using the recently described solid phase support, 4‐(oxymethyl)phenylacetamidomethyl‐polystyrene. The resultant crude polypeptide exhibited 30–50% of native nuclease‐T enzymatic activity when added to native nuclease‐T‐(50–149). When the noncovalent complex formed by native nuclease‐T‐(50–149) and a 10‐fold molar excess of [Gly 48]synthetic‐(6–49) was equilibrated with trypsin in 90% glycerol, an increase in enzymatic activity from 8 to 32% (versus nuclease) was observed. Simultaneously, approximately 20% conversion of nuclease‐T‐(50–149) to nuclease‐molecular weight material was observed by gel electrophoretic analysis. These data indicate that a covalent semisynthetic species is formed with activity about equal to that of native nuclease. The results confirm the importance of loop integrity on catalytic site organization. The Gly 48‐containing fragment system defined above can allow preparation of semisynthetic nuclease sequence analogs.
BOVINE PROTHROMBIN FRAGMENT 1, SEGMENT 1–10
Authors: TEN KORTENAAR PAULUS B.W. , WILKERSON WENDELL W. , BOGGS NORMAN T. , MADAR DEAN A. , KOEHLER KARL A. , HISKEY RICHARD G.
Published: August 2010

The N‐terminal decapeptide methyl ester, H‐Ala‐Asn‐Lys‐Gly‐Phe‐Leu‐Gla‐Gla‐Val‐Arg‐OCH3(16)of bovine prothrombin fragment 1 has been prepared by standard solution techniques, via a fragment coupling strategy. Hexapeptide Boc‐Ala‐Asn‐Lyse(Boc)‐Gly‐Phe‐Leu‐OBzl (9) was obtained by coupling Boc‐Ala‐Asn‐Lyse (Boc)Gly‐OH (6)to the trifluoroacetate salt of H‐Phe‐Leu‐OBzl (8). Hydrogenolysis of(9)followed by coupling to HCl‐H‐Glaγ (OtBu)2‐Glaγ (OtBu)2‐Val‐Arg(HCl)‐OCH3(14)gave the fully protected decapeptide (15). Treatment of15with 90% trifluoroacetic acid followed by ion exchange chromatography yielded the methyl ester (16). The decapeptide16labeled with125I using the Bolton‐Hunter reagent, did not bind to antibodies specific for the calcium ion‐induced conformation of bovine fragment 1.