Addressing the Tertiary Structure of Human Parathyroid Hormone-(1–34)

Pellegrini, Maria; Royo, Míriam; Rosenblatt, Michael; Chorev, Michael; Mierke, Dale F.

doi:10.1074/jbc.273.17.10420

Cited by 99 publications

(150 citation statements)

References 55 publications

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“…8B. Note that a considerable degree of flexibility between the amino-terminal and carboxyl-terminal domains of PTH-(1-34) has been observed in solution-phase NMR studies (39).…”

Section: Analogsmentioning

confidence: 99%

Multiple Sites of Contact between the Carboxyl-terminal Binding Domain of PTHrP-(1–36) Analogs and the Amino-terminal Extracellular Domain of the PTH/PTHrP Receptor Identified by Photoaffinity Cross-linking

Gensure¹,

Gardella²,

Jüppner³

2001

Journal of Biological Chemistry

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The carboxyl-terminal portions of parathyroid hormone (PTH)-(1-34) and PTH-related peptide (PTHrP)-(1-36) are critical for high affinity binding to the PTH/ PTHrP receptor (P1R), but the mechanism of receptor interaction for this domain is largely unknown. To identify interaction sites between the carboxyl-terminal region of PTHrP-(1-36) and the P1R, we prepared analogs of [ -172. These data thus predict that residues 23, 27, 28, and 33 of native PTHrP are each near to different regions of the amino-terminal extracellular receptor domain of the P1R. This information helps define sites of proximity between several ligand residues and this large receptor domain, which so far has been largely excluded from models of the hormone-receptor complex. PTH and PTHrP1 mediate many of their biological effects through the same receptor (1, 2). Peptides containing the first 34 amino acids of PTH and PTHrP are capable of fully activating the PTH/PTHrP receptor (P1R) (1, 2). Studies with PTH and PTHrP ligand analogs and receptor chimeras have suggested that the ligands have two distinct functional domains: the amino-terminal residues, which are important for receptor activation; and the carboxyl-terminal residues, which are important for high affinity binding (3). These data furthermore indicate that the ligand's amino-terminal portion interacts with the extracellular loops and the membrane-spanning helices of the receptor, and the ligand's carboxyl-terminal portion interacts with the receptor's amino-terminal extracellular domain (3). A similar pattern of ligand-receptor interaction has been suggested for other members of this class II family of peptide hormone G-protein-coupled receptors, including the secretin receptor (4) and the PTH-2 receptor (5-7).Some specific sites of interaction between the amino-terminal portions of PTH-(1-34)/PTHrP-(1-36) and the P1R have been identified by site-directed mutagenesis and photoaffinity cross-linking studies. For example, mutational analyses have shown that residues in extracellular loop 3 and the adjacent sixth membrane-spanning helix (TM6) are critical for mediating ligand-induced receptor activation (2,8). Consistent with these mutational data, Bpa introduced at position 1 of PTH-(1-34) or position 2 of either PTH-(1-34) or PTHrP-(1-36) was found to cross-link to methionine 425, at the extracellular end of TM6 in the P1R (9, 10). Interactions between the aminoterminal portion of the ligand and the transmembrane domains/extracellular loops of the receptor are also suggested by a study showing that PTH-(1-14) can stimulate cAMP accumulation in a mutant P1R missing most of the amino-terminal extracellular domain as efficiently as it does in the wild-type P1R (11). Furthermore, if a PTH fragment comprising the first 9 amino acids is covalently attached to the amino-terminal end of such a truncated receptor, the resulting ligand-receptor chimera displays constitutive activity, indicative of an intramolecular stimulation of the receptor's activation domain by the tethered ligand fragment ...

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“…8B. Note that a considerable degree of flexibility between the amino-terminal and carboxyl-terminal domains of PTH-(1-34) has been observed in solution-phase NMR studies (39).…”

Section: Analogsmentioning

confidence: 99%

Multiple Sites of Contact between the Carboxyl-terminal Binding Domain of PTHrP-(1–36) Analogs and the Amino-terminal Extracellular Domain of the PTH/PTHrP Receptor Identified by Photoaffinity Cross-linking

Gensure¹,

Gardella²,

Jüppner³

2001

Journal of Biological Chemistry

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“…6). The conformation of hPTH-(1-34) used in the molecular modeling was obtained from our high resolution NMR studies performed in different environments, including aqueous saline conditions and the presence of dodecylphosphocholine as a membrane mimetic (37). DISCUSSION The characterization of the bimolecular interaction between the activation domain of PTH-(1-34) and the hPTH1-Rc is of fundamental importance for elucidating the molecular mechanism of signal transduction.…”

Section: Characterization Of Transiently Transfected Cos-7 Cells Exprmentioning

confidence: 99%

Parathyroid Hormone-Receptor Interactions Identified Directly by Photocross-linking and Molecular Modeling Studies

Bisello¹,

Adams²,

Mierke³

et al. 1998

Journal of Biological Chemistry

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Direct mapping of the interface between parathyroid hormone (PTH) and its receptor (hPTH1-Rc Parathyroid hormone (PTH)1 is the major regulator of calcium levels in blood and plays a role in the regulation of bone remodeling (1). Given intermittently, PTH displays anabolic activity in bone and, therefore, has considerable therapeutic potential (2). PTH and PTH-related protein exert their actions via a seven-transmembrane (TM) domain-containing receptor (PTH1-Rc) (3) belonging to a subfamily of related G proteincoupled receptors (4 -11). The PTH1-Rc is coupled to both adenylyl cyclase/cyclic AMP and phospholipase C/inositol 1,4,5-trisphosphate/cytosolic calcium intracellular signaling pathways (12)(13)(14)(15).Understanding the molecular mechanism of ligand recognition and signal transduction by the PTH1-Rc may identify new directions for the design of novel hormone analogs for the treatment of diseases such as osteoporosis, hypercalcemia of malignancy and hyperparathyroidism (16). In order to directly identify the structural elements involved in PTH-PTH1-Rc interactions, we employed a photoaffinity scanning approach (17). The generation of covalently linked ligand-receptor conjugates and the identification of the cross-linked domains allows mapping of the interface between hormone and receptor. Photoaffinity cross-linking has been successfully applied in defining interactions between small peptides, such as substance P (18 -20), cholecystokinin (21), and vasopressin (22), and their receptors. Recently, we used this general approach to identify directly the interaction between position 13 of PTH and a 17-amino acid domain (residues 173-189) of the hPTH1-Rc (17).We now report the evaluation of a series of photoreactive analogs obtained by a "p-benzoylphenylalanine (Bpa) scan" of the principal receptor activation domain (residues 1-6) of 34)), maintained full potency and led to the identification of a second "contact domain" between PTH and hPTH1-Rc. This information allows us to create, for the first time, a model describing interactions of hPTH-(1-34) with its receptor based on direct identification of the interacting regions. EXPERIMENTAL PROCEDURESMaterials-Boc-protected amino acids, N-hydroxybenzotriazole, N,NЈ-dicyclohexylcarbodiimide, and p-methylbenzydrylamine resin were purchased from Applied Biosystems (Foster City, CA). Boc-(3-iodo)tyrosine ] was from Peninsula Laboratories (Belmont, CA). B&J brand dichloromethane, N-methylpyrrolidone, and acetonitrile were obtained from Baxter (McGraw Park, IL). IODOGEN ® and 2-(2Ј-nitrophenylsulfenyl)-3-methyl-3-bromoindolenine (BNPS-skatole) were purchased from Pierce. Cyanogen bromide was from Aldrich. Na

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“…; ref. 23). The only distance constraints were the experimentally derived distances as described above.…”

Section: Methodsmentioning

confidence: 99%

Architecture of recombination intermediates visualized by in-gel FRET of λ integrase–Holliday junction–arm DNA complexes

Radman‐Livaja

Biswas

Mierke

et al. 2005

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

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Addressing the Tertiary Structure of Human Parathyroid Hormone-(1–34)

Cited by 99 publications

References 55 publications

Multiple Sites of Contact between the Carboxyl-terminal Binding Domain of PTHrP-(1–36) Analogs and the Amino-terminal Extracellular Domain of the PTH/PTHrP Receptor Identified by Photoaffinity Cross-linking

Multiple Sites of Contact between the Carboxyl-terminal Binding Domain of PTHrP-(1–36) Analogs and the Amino-terminal Extracellular Domain of the PTH/PTHrP Receptor Identified by Photoaffinity Cross-linking

Parathyroid Hormone-Receptor Interactions Identified Directly by Photocross-linking and Molecular Modeling Studies

Architecture of recombination intermediates visualized by in-gel FRET of λ integrase–Holliday junction–arm DNA complexes

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