Study of the secondary structure of the luteinizing hormone releasing factor using intramolecular charge transfer complexes

Donzel, B.; Rivier, Jean; Goodman, Murray

doi:10.1021/bi00631a006

Cited by 20 publications

(13 citation statements)

References 24 publications

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“…The present antibody, which is defined as a monoclonal antibody, not only on the grounds of its repeated cloning by limiting dilution but also by its single isotype of heavy and light chain and the physicochemical properties of monolinear association constant, demonstrated the exceptional merits of intercepting the hormonal activity of GnRH in vivo. The antibody is directed against a conformation in which both the NH2-and COOH-terminal regions of GnRH participate, a postulate consistent with data based on physicochemical and computer graphic studies (15,16).…”

Section: Discussionsupporting

confidence: 58%

“…These results indicate the optimal binding of the antibody with the entire molecule and suggest that the reactive topology is contributed to by both the NH2-and COOH-terminal parts. The proximity of the NH2 terminus to the COOH terminus of the decapeptide has been indicated in spectroscopic (15) and computer graphic studies (16 GnRH does not circulate in amounts high enough to permit direct assay, even in the mammals in which its role is known, estimation of the blood level of the hormone in the dogs by direct RIA was not feasible. Instead, the effect of the monoclonal anti-GnRH antibody on estrus was tested in female dogs given a single intravenous injection of 3.0 ml at proestrus.…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Bioeffective monoclonal antibody against the decapeptide gonadotropin-releasing hormone: reacting determinant and action on ovulation and estrus suppression.

Talwar¹,

Gupta²,

Singh³

et al. 1985

Proc. Natl. Acad. Sci. U.S.A.

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A monoclonal antibody generated against the decapeptide gonadotropin-releasing hormone (GnRH) was effective in intercepting the bioactivity of the hormone; it blocked ovulation in rats. The antibody reacted optimally with the native hormone. Substitution of amide at the COOH terminus by a carboxyl group decreased immunoreactivity by a factor of 200. The antibody recognized the amino acid sequences 4-6, 7-10, and 4-10 to a variable degree, which suggests that the epitope has a conformation involving the entire molecule, with the NH2-and COOH-terminal regions probably in proximity. The antibody was also competent to suppress the progression of estrus in dogs, an indication that GnRH may play an inductive role in the reproductive function of dogs.Gonadotropin-releasing hormone (GnRH) has a pivotal role in mammalian reproduction. By regulating the secretion of follicle-stimulating hormone and luteinizing hormone from the pituitary, it controls spermatogenesis in the male and ovulation in the female, with concomitant production of sex steroids. The hormone is a decapeptide, which has been synthesized along with several analogs having agonistic or antagonistic properties pointing to the portions of the molecule important for the biological activity of the hormone (1, 2). Attempts have also been made to map the reading framework of the conventional polyvalent antibodies raised against the hormone (3, 4). This communication reports the bioefficacy of a monoclonal antibody generated against the hormone for block of ovulation in rats. The binding characteristics of this antibody with the entire decapeptide and subpeptide fragments have been explored to obtain information on the topology of the epitope(s) important for bioneutralization. A practical use of the antibody is indicated by its ability to suppress the progression of estrus in dogs.

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Section: Discussionsupporting

confidence: 58%

Section: Resultsmentioning

confidence: 99%

Bioeffective monoclonal antibody against the decapeptide gonadotropin-releasing hormone: reacting determinant and action on ovulation and estrus suppression.

Talwar¹,

Gupta²,

Singh³

et al. 1985

Proc. Natl. Acad. Sci. U.S.A.

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“…This phenomenon was observed in GnRH: replacement of Gly6 by any l ‐amino acid drastically decreased the biological potency, whereas replacement by a d ‐amino acid residue dramatically increased the activity; substitution of Gly6 in GnRH by l ‐Phe resulted in 10% activity compared with 100% of GnRH, whereas replacement with d ‐Phe led to 750% activity (25,26). NMR and spectroscopic studies have indicated the existence of a type II β ‐turn in d ‐amino acids 6 GnRH analogs (27).…”

Section: Discussionmentioning

confidence: 99%

“…Based upon semiempirical calculations, it was suggested (23), that a b-turn of type II (24) is destabilized by introduction of an l-amino acid residue at the third position of the turn, whereas this con- activity compared with 100% of GnRH, whereas replacement with d-Phe led to 750% activity (25,26). NMR and spectroscopic studies have indicated the existence of a type II b-turn in d-amino acids 6 GnRH analogs (27).…”

Section: Discussionmentioning

confidence: 99%

In vivo structure–activity studies on the dark‐color‐inducing neurohormone of locusts

Grach

Wang

Barda

et al. 2003

The Journal of Peptide Research

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In the 11-residue long dark-color-inducing neurohormone (DCIN = [His7]-corazonin), of locusts, from residue 2 to residue 11, one amino acid at each time was substituted by D-phenylalanine (D-Phe). The dark-color-inducing effect of these peptides was investigated in comparison with unaltered DCIN by a bioassay based on nymphs of a DCIN-deficient albino mutant of the migratory locust, Locusta migratoria. Substitution of any single amino acid by D-Phe always reduced the activity, but did not abolish it completely. Maximum inactivation was obtained after substitution of Gln4, Ser6, or Trp9. The latter two residues are within the partial sequence -Ser-Xxx-Gly-Trp- (Xxx = His in theDCIN) that seems to be important for the dark-color-inducing activity, as found also in another study (Insect Biochem. Mol. Biol.32, 2002, 909). GIn4, however, is outside of this partial sequence.Minimal, although still considerable, inactivation occurred after substitution of Gly8, Phe3, or Asn11, despite the fact that Gly8 is within the -Ser-Xxx-Gly-Trp- partial sequence. In conclusion, no single active core was found, indicating that the whole sequence of the DCIN is necessary to induce maximum darkening effect. No difference was found in the activity of the peptides in which Gly8was substituted by D-Phe or by L-Phe. Therefore the -Ser-Xxx-Gly-Trp- partial sequence does not seem to be stabilized by a type II beta-turn. Nevertheless, existence of another kind of turn that includes this partial sequence is feasible. A single unsuccessful attempt was made to discover an antagonist to the DCIN.

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“…The cross peak at w1 = 7.15 ppm and w2 = 10.83 ppm depicts connectivity between the ring NH and the C2H protons. The proton at 7.61 ppm is coupled to proton at 6.91 ppm which also shows a J-coupling to another proton at 7.03 ppm, which in turn is J-coupled to the proton at 7.26 ppm. From the very nature of such a COSY pattern, it can be concluded that the Fig.…”

Section: Resonance Assignment In (Cd3)zsomentioning

confidence: 99%

Molecular conformation of gonadoliberin using two-dimensional NMR spectroscopy

Chary¹,

Srivastava²,

Hosur³

et al. 1986

Eur J Biochem

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Complete resonance assignments of the proton NMR spectrum of gonadoliberin (in its native amide and free acid forms) have been obtained using two-dimensional nuclear magnetic resonance spectroscopy under three different environmental conditions, namely, dimethyl sulphoxide solution, aqueous solution and lipid-bound form in model membranes. The proton chemical shifts in the three cases have been compared to derive information about inherent conformational characteristics of the molecule. It has been inferred that the molecule possesses no short-range or long-range order under any of the three solvent conditions. However, there is a nonspecific increase in the linewidths when gonadoliberin is bound to model membranes, indicating a reduced internal motion in the molecule due to lipid-peptide interactions.Conadoliberin is a hypothalamic peptide consisting of a linear chain of ten amino acid residues ( M , = 1170). Its primary sequence has been established to be Glp-His-Trp-SerTyr-Gly-Leu-Arg-Pro-Gly-NH2. The hormone, which plays a pivotal role in mammalian reproduction, regulates the secretion of folitropin and lutropin from pitutary. Folitropin and lutropin, in turn, regulate the production of sex steroids, thereby controlling spermatogenesis in the male and ovulation in the female. A major interest in contraception research has therefore focussed on this small peptide [l].The native peptide lacks free terminal carboxyl groups. The hormone as well as its numerous analogues, agonists or antagonists, have been synthesised and studied. Substitutions with D-amino acids at specific positions, particularly at 2, 6 or 10, drastically alter the biological activity of the peptide [2]. Recently, a bioactive anti-gonadoliberin monoclonal antibody, which blocks ovulation in rats and prevents estrus in dogs, has been shown to recognise, though poorly, gonadoliberin peptide with a free carboxyl group at C-terminal or its different fragments (residues 4-6, 7-10 and 4 -10). This may suggest that the relevant epitope comprises a conformation involving the entire molecule [3].To understand the structure-function relationships of the hormone, extensive studies on the conformation in different solvents have been carried out [4-lo]. However, no unequivocal structure could be established for the decapeptide from such studies. Using model building [l 1 ,121 and potential energy calculations [13], a folded structure with a p bend has been suggested for the active form. In this structure, the Nand the C-terminals are in close proximity.In view of these interesting observations, we have undertaken a comparative study of the native peptide (gonadoiiberin-NH,) and its free acid (gonadoliberin-OH) using twodimensional NMR spectroscopy. Resonance assignments The spectra of only the native amide are described in detail since the spectral characteristics of the free acid are very similar to those of the amide. EXPERIMENTAL PROCEDURES Sample preparationFor all NMR experiments on gonadoliberin-NH2 in (CD3)2S0 and DzO, a concentration of 12 mM was p...

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Study of the secondary structure of the luteinizing hormone releasing factor using intramolecular charge transfer complexes

Cited by 20 publications

References 24 publications

Bioeffective monoclonal antibody against the decapeptide gonadotropin-releasing hormone: reacting determinant and action on ovulation and estrus suppression.

Bioeffective monoclonal antibody against the decapeptide gonadotropin-releasing hormone: reacting determinant and action on ovulation and estrus suppression.

In vivo structure–activity studies on the dark‐color‐inducing neurohormone of locusts

Molecular conformation of gonadoliberin using two-dimensional NMR spectroscopy

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