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Examines the characteristics of human parathyroid hormone (PTH) and PTH-related protein. Biological properties of PTH; Stability of the N-terminal helix; Presence of amino acid residue in the helical structure of protein.

The strong propensity of 2-amino-2-methyl propanoic acid (Aib)-rich peptides to form stable helical structures is well documented. NMR analysis of the short peptide Z-(Aib)[sub 5]-L-Leu-(Aib)[sub 2]-OMe indicates the presence of a well-characterized 3[sub 10]-helix even in dimethylsulfoxide (DMSO), a solvent known to disrupt helical structures. The structure remains stable at least up to 348 K. Stereospecific assignment of the diastereotopic methyls of Aib was achieved, with the assumption of a specific helical screw sense. The methyl more eclipsed with respect to the CO vector resonates at a higher field in the carbon dimension. Molecular dynamics simulations successfully predict the [sup 3]J[sub CHNH] coupling constant of Leu[sup 6] and most of the H-bonding pattern. Discrepancies were found for Aib[sup 3] and Aib[sup 7] amide protons which can be explained by a higher sensitivity of the simulations to the helix fraying at the end of the peptide and by the presence of extended conformations for Leu[sup 6] during most of the simulations. [ABSTRACT FROM AUTHOR]

Diterpenes have recently received a great deal of interest as tools to investigate the botanical origin of coffee. Specifically, kahweol has been proposed as a marker of Coffea arabica while 16-O-methylcafestol (16-OMC) is a Coffea canephora specific marker and its detection and quantification allow the authenticity of pure C. arabica roasted coffee blends to be assessed. In this study, we evaluated the possibility of the industrial use of the quantification of these diterpenes to assess the relative amounts of the two coffee species in blends. The content of 16-OMC and kahweol was determined in 78 samples (i.e., 39 green and the corresponding 39 roasted beans) of C. canephora from different geographical origins using a recently published NMR approach. Our results show a small natural variability in 16-OMC content for the Asian samples (average content = 1837 ± 113 mg/kg) while a much larger spread was found for the African samples (average content = 1744 ± 322 mg/kg). This large variability prevents the use of 16-OMC to quantify C. canephora in unknown roasted coffee blends. We also show that kahweol cannot be considered a specific C. arabica marker since it was detected almost all coffees beans and quantified in about 30% of the C. canephora samples. [ABSTRACT FROM AUTHOR]

In this work, several aroma precursors present in Moscato Giallo grape juice were identified and characterized using LC–MS and NMR techniques. A preliminary separation of various fractions was obtained using adsorption on Amberlite® XAD resin and HPLC chromatography on a reverse phase column. Subsequently, U-HPLC with mass spectrometry allowed the identification of some compounds corresponding to mono- and disaccharides linked to terpenes. The MS–MS fragmentation step indicated which kind of glycosides, the moiety sequence and sometimes which kind of terpene were present. NMR enabled the correct identification of glycosides and terpene when the fraction analyzed was sufficiently concentrated and with few components. Twelve glycosidically bound terpenes were characterized: (E) and (Z)-furanosyl-linalooloxide-7-O-[α-d-apiofuranosyl-(1→6)-1-β-d-glucopyranoside], (E)-furanosyl-linalooloxide-7-O-[1-β-d-glucopyranoside], (Z)-8-hydroxylinalool-8-O-[1-β-d-glucopyranoside], 1,2-dihydroxylinalool-1-O-[1-β-d-glucopyranoside], linalool-3-O-[α-l-arabinofuranosyl-(1→6)-1-β-d-glucopyranoside], linalool-3-O-[α-l-apiofuranosyl-(1→6)-1-β-d-glucopyranoside], linalool-3-O-[α-l-rhamnopyranosyl-(1→6)-1-β-d-glucopyranoside], nerol-1-O-α-d-apiofuranosyl-(1→6)-1-β-d-glucopyranoside, geraniol-1-O-[α-l-arabinofuranosyl-(1→6)-1-β-d-glucopyranoside], geraniol-1-O-[α-l-rhamnopyranosyl-(1→6)-1-β-d-glucopyranoside], and a geranic acid disaccharide derivative. It is the first time that this kind of compounds are directly detected and identified in a mixture with these two techniques. [ABSTRACT FROM AUTHOR]

The N-terminal 1-34 segment of parathyroid hormone (PTH) is fully active in vitro and in vivo and it elicits all the biological responses characteristic of the native intact PTH. Recent studies reported potent helical analogues of the PTH (1-11) with helicity-enhancing substitutions. This work describes the synthesis, biological activity, and conformational studies of analogues obtained from the most active non-natural PTH (1-11) peptide H-Aib-Val-Aib-Glu-Ile-Gln-Leu-Nle-His-Gln-Har-NH; specifically, the replacement of Val in position 2 with d-Val, l-(αMe)-Val and N-isopropyl-Gly was studied. The synthesized analogues were characterized functionally by in-cell assays and their structures were determined by CD and NMR spectroscopy. To clarify the relationship between the structure and activity, the structural data were used to generate a pharmacophoric model, obtained overlapping all the analogues. This model underlines the fundamental functional role of the side chain of Val and, at the same time, reveals that the introduction of conformationally constrained C-tetrasubstituted α-amino acids in the peptides increases their helical content, but does not necessarily ensure significant biological activity. [ABSTRACT FROM AUTHOR]

The N-terminal 1–34 segment of parathyroid hormone (PTH) is fully active in vitro and in vivo and it can reproduce all biological responses in bone characteristic of the native intact PTH. Recent studies have demonstrated that N-terminal fragments presenting the principal activating domain such as PTH(1–11) and PTH(1–14) with helicity-enhancing substitutions yield potent analogues with PTH(1–34)-like activity. To further investigate the role ofα-helicity on biological potency, we designed and synthesized by solid-phase methodology the following hPTH(1–11) analogues substituted at positions 1 and/or 3 by the sterically hindered and helix-promoting C α-tetrasubstitutedα-amino acidsα-amino isobutyric acid (Aib), 1-aminocyclopentane-1-carboxylic acid (Ac5c) and 1-aminocyclohexane-1-carboxylic acid (Ac6c): Ac5c-V-Aib-E-I-Q-L-M-H-Q-R-NH2 (I); Aib-V-Ac5c-E-I-Q-L-M-H-Q-R-NH2 (II); Ac6c-V-Aib-E-I-Q-L-M-H-Q-R-NH2 (III); Aib-V-Ac6c-E-I-Q-L-M-H-Q-R-NH2 (IV); Aib-V-Aib-E-I-Q-L-M-H-Q-R-NH2 (V); S-V-Aib-E-I-Q-L-M-H-Q-R-NH2 (VI), S-V-Ac5c-E-I-Q-L-M-H-Q-R-NH2 (VII); Ac5c-V-S-E-I-Q-L-M-H-Q-R-NH2 (VIII); Ac6c-V-S-E-I-Q-L-M-H-Q-R-NH2 (IX); Ac5c-V-Ac5c-E-I-Q-L-M-H-Q-R-NH2 (X); Ac6c-V-Ac6c-E-I-Q-L-M-H-Q-R-NH2 (XI). All analogues were biologically evaluated and conformationally characterized in 2,2,2-trifluoroethanol (TFE) solution by circular dichroism (CD). AnaloguesI– V, which cover the full range of biological activity observed in the present study, were further conformationally characterized in detail by nuclear magnetic resonance (NMR) and computer simulations studies. The results of ligand-stimulated cAMP accumulation experiments indicated that analoguesIandIIare active, analoguesIII,VIandVIIare very weakly active and analoguesIV,V,VIII–XIare inactive. The most potent analogue,Iexhibits biological activity 3500-fold higher than that of the native PTH(1–11) and only 15-fold weaker than that of the native sequence hPTH(1–34). Remarkably, the two most potent analogues,IandII, and the very weakly active analogues,VIandVII, exhibit similar helix contents. These results indicate that the presence of a stable N-terminal helical sequence is an important but not sufficient condition for biological activity. [ABSTRACT FROM AUTHOR]

To evaluate the ability of molecular dynamics (MD) simulations using atomic force-fields to correctly predict stable folded conformations of a peptide in solution, we show results from MD simulations of the reversible folding of an octapeptide rich in α-aminoisobutyric acid (2-amino-2-methyl-propanoic acid, Aib) solvated in di-methyl-sulfoxide (DMSO). This solvent generally prevents the formation of secondary structure, whereas Aib-rich peptides show a high propensity to form secondary structural elements, in particular 3[sub 10]- and α-helical structures. Aib is, moreover, achiral, so that Aib-rich peptides can form left- or right-handed helices depending on the overall composition of the peptide, the temperature, and the solvation conditions. This makes the system an interesting case to study the ensembles of peptide conformations as a function of temperature by MD simulation. Simulations involving the folding and unfolding of the peptide were performed starting from two initial structures, a right-handed α-helical structure and an extended structure, at three temperatures, 298 K, 340 K, and 380 K, and the results are compared with experimental nuclear magnetic resonance (NMR) data measured at 298 K and 340 K. The simulations generally reproduce the available experimental nuclear Overhauser effect (NOE) data, even when a wide range of conformations is sampled at each temperature. The importance of adequate statistical sampling in order to reliably interpret the experimental data is discussed. [ABSTRACT FROM AUTHOR]