title: Bioactive N-terminal undecapeptides derived from parathyroid hormone: the role of alpha-helicity.
abstract: 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 of alpha-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(alpha)-tetrasubstituted alpha-amino acids alpha-amino isobutyric acid (Aib), 1-aminocyclopentane-1-carboxylic acid (Ac(5)c) and 1-aminocyclohexane-1-carboxylic acid (Ac(6)c): Ac(5)c-V-Aib-E-I-Q-L-M-H-Q-R-NH(2) (I); Aib-V-Ac(5)c-E-I-Q-L-M-H-Q-R-NH(2) (II); Ac(6)c-V-Aib-E-I-Q-L-M-H-Q-R-NH(2) (III); Aib-V-Ac(6)c-E-I-Q-L-M-H-Q-R-NH(2) (IV); Aib-V-Aib-E-I-Q-L-M-H-Q-R-NH(2) (V); S-V-Aib-E-I-Q-L-M-H-Q-R-NH(2) (VI), S-V-Ac(5)c-E-I-Q-L-M-H-Q-R-NH(2) (VII); Ac(5)c-V-S-E-I-Q-L-M-H-Q-R-NH(2) (VIII); Ac(6)c-V-S-E-I-Q-L-M-H-Q-R-NH(2) (IX); Ac(5)c-V-Ac(5)c-E-I-Q-L-M-H-Q-R-NH(2) (X); Ac(6)c-V-Ac(6)c-E-I-Q-L-M-H-Q-R-NH(2) (XI). All analogues were biologically evaluated and conformationally characterized in 2,2,2-trifluoroethanol (TFE) solution by circular dichroism (CD). Analogues I-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 analogues I and II are active, analogues III, VI and VII are very weakly active and analogues IV, V, VIII-XI are inactive. The most potent analogue, I exhibits 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, I and II, and the very weakly active analogues, VI and VII, 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.
authors.affiliation: Division of Bone and Mineral Research, Beth Israel Deaconess Medical Center and Harvard Medical School, 330 Brookline Ave, Boston, MA 02215, USA.
authors.forename: A
authors.initials: A
authors.lastname: Barazza
authors.forename: A
authors.initials: A
authors.lastname: Wittelsberger
authors.forename: N
authors.initials: N
authors.lastname: Fiori
authors.forename: E
authors.initials: E
authors.lastname: Schievano
authors.forename: S
authors.initials: S
authors.lastname: Mammi
authors.forename: C
authors.initials: C
authors.lastname: Toniolo
authors.forename: J M
authors.initials: JM
authors.lastname: Alexander
authors.forename: M
authors.initials: M
authors.lastname: Rosenblatt
authors.forename: E
authors.initials: E
authors.lastname: Peggion
authors.forename: M
authors.initials: M
authors.lastname: Chorev
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D002942:Circular Dichroism; D000242:Cyclic AMP / Q000378:metabolism; D006801:Humans; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D009842:Oligopeptides / Q000737:chemistry* / Q000494:pharmacology*; D010281:Parathyroid Hormone / Q000031:analogs & derivatives* / Q000737:chemistry*; D017433:Protein Structure, Secondary; D044168:Receptor, Parathyroid Hormone, Type 1; D018016:Receptors, Parathyroid Hormone / Q000235:genetics / Q000378:metabolism
chemical_list: D008024:Ligands; D009842:Oligopeptides; C473449:PTH1R protein, human; D010281:Parathyroid Hormone; D044168:Receptor, Parathyroid Hormone, Type 1; D018016:Receptors, Parathyroid Hormone; D000242:Cyclic AMP