The Structure Of Human Parathyroid Hormone From a Study of Fragments in Solution Using 1H NMR Spectroscopy and Its Biological Implications

Abstract

In order to gain insight into the structure of human parathyroid hormone (hPTH), four fragments [hPTH(1-34), hPTH(18-48), hPTH(28-48), and hPTH(53-84)], which cover all regions of the intact hormone, have been investigated by CD and NMR spectroscopy in combination with distance geometry, and restrained molecular dynamics and energy minimization calculations, under a variety of solution conditions. Significantly, all fragments showed little propensity to form stable structures in aqueous solution alone, and it was only on the addition of trifluoroethanol (TFE) that defined structural features were observed. In an extension of earlier work [Klaus et al. (1991) Biochemistry 30, 6936-6942], hPTH(1-34) in 70% trifluoroethanol (TFE) showed two helices that were longer than in 10% TFE, but essentially showed the same characteristics. Although overlap in the 1H NMR spectra prevented the determination of quantitative NOE data for residues 26-30, the combination of the alpha-proton chemical shift data and quantitative NOE data indicated the helices extend from residues 3 to 13 and 15 to 29. No evidence was found for interaction of the two helical regions. The nature and extent of this second helix in the intact hormone were better defined from the data for hPTH(18-48). Under limiting solution conditions, where the fragment assumed its maximum helical content, a well-defined helix was observed between residues 21 and 38 with a possible discontinuity between Leu-28 and Gln-29. There was little evidence of any form of secondary structure between Gly-38 and the terminus of this fragment, Ser-48. In keeping with this result, the shorter fragment, hPTH(28-48), showed little evidence of stable secondary structure on addition of TFE. From the alpha-proton chemical shifts residues 23-27 appeared to sustain helical structure more readily than the rest of molecule under all solution regimes in both hPTH(1-34) and hPTH(18-48). In contrast to the other two longer fragments hPTH(53-84) showed little propensity for helical secondary structure even at the highest TFE concentrations. However, there was evidence that the molecule did adopt a defined three-dimensional structure. Various long-range NOE's were observed in 10% TFE that allowed the calculation of an open tertiary structure consisting of an initial series of turns surrounded by a loop structure of several loose turns.(ABSTRACT TRUNCATED AT 400 WORDS)

Comparative Study of Chicken and Human Parathyroid Hormone‐(1–34)‐Peptides in Solution with SDS

“…These compact helical regions are joined by a flexible linker which assumes either an irregular turn or a helical structure (Barden and Cuthbertson, 1993;Klaus et al, 1991 ;Strickland et al, 1993;Wray et al, 1994).…”

Stimulation of creatine kinase activity in rat skeletal tissue in vivo and in vitro by protease-resistant variants of parathyroid hormone fragments

“…*P < 0.05, **P < 0.01, **P < 0.005, compared with activity in the absence of PTH fragment. structural investigations with hPTH and PTH-derived peptides showed that they tend to be largely unstructured in aqueous solution [42]. However, they can adopt significant a-helical conformation when the hydrophobicity of the solution is in- creased by adding TFE, which may be supposed to mimic the situation when the hormone molecules are approaching their surface receptor [42,48].…”

“…structural investigations with hPTH and PTH-derived peptides showed that they tend to be largely unstructured in aqueous solution [42]. However, they can adopt significant a-helical conformation when the hydrophobicity of the solution is in- creased by adding TFE, which may be supposed to mimic the situation when the hormone molecules are approaching their surface receptor [42,48]. We therefore monitored the a-helical content of hPTH- in the presence of increasing TFE concentrations by CD spectroscopy and found it to reach a plateau when 70 % TFE is exceeded.…”

Structure-Activity Relation of NH2-terminal Human Parathyroid Hormone Fragments

“…In particular, hPTH-(1-34) is an intensely studied hormone fragment as it contains all functional domains (14 -17). From most experiments it was concluded that hPTH-(1-34) does not form secondary structure elements in the absence of TFE (14,16,18), but helix formation in TFE-free solution is nevertheless observed for hPTH-(1-34), residues 4 -13 and 21-29 (19), and for hPTH-(1-37), residues 5-10 and 17-28 (20). In TFE-containing solution hPTH-(1-34) displays helical regions from Ser-3 to Gly-12 and from Ser-17 to Lys-26 (16,18), but no tertiary interactions for hPTH- are found under these conditions.…”

“…From most experiments it was concluded that hPTH-(1-34) does not form secondary structure elements in the absence of TFE (14,16,18), but helix formation in TFE-free solution is nevertheless observed for hPTH-(1-34), residues 4 -13 and 21-29 (19), and for hPTH-(1-37), residues 5-10 and 17-28 (20). In TFE-containing solution hPTH-(1-34) displays helical regions from Ser-3 to Gly-12 and from Ser-17 to Lys-26 (16,18), but no tertiary interactions for hPTH- are found under these conditions. It is commonly known that TFE stabilizes secondary structures, in particular helices (21)(22)(23)(24)(25)(26), but bears the risk of weakening hydrophobically stabilized tertiary structure domains (24), an effect also observed for hPTH-(1-34).…”