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Parathyroid hormone-receptor interactions identified directly by photocross-linking and molecular modeling studies
- Source :
- The Journal of biological chemistry. 273(35)
- Publication Year :
- 1998
-
Abstract
- Direct mapping of the interface between parathyroid hormone (PTH) and its receptor (hPTH1-Rc) was carried out by photoaffinity scanning studies. Photoreactive analogs of PTH singularly substituted with a p-benzoylphenylalanine (Bpa) at each of the first six N-terminal positions have been prepared. Among these, the analog [Bpa1,Nle8,18,Arg13,26,27,l-2-Nal23,Tyr34]bPTH-(1–34)NH2(Bpa1-PTH-(1–34)) displayed in vitroactivity with potency similar to that of PTH-(1–34). The radioiodinated analog 125I-Bpa1-PTH-(1–34) cross-linked specifically to the hPTH1-Rc stably expressed in human embryonic kidney cells. A series of chemical and enzymatic digestions of the hPTH1-Rc–125I-Bpa1-PTH-(1–34) conjugate suggested that a methionine residue (either Met414 or Met425) within the contact domain hPTH1-Rc-(409–437), which includes the transmembrane helix 6 and part of the third extracellular loop, as the putative contact point. Site-directed mutagenesis (M414L or M425L) identified Met425 as the putative contact point. Molecular modeling of the hPTH1-Rc together with the NMR-derived high resolution structure of hPTH-(1–34), guided by the cross-linking data, strongly supports Met425, at the extracellular end of transmembrane helix 6, as the residue interacting with the N-terminal residue of the hPTH-(1–34). The photocross-linking and molecular modeling studies provide insight into the topologic arrangement of the receptor-ligand complex.
- Subjects :
- Models, Molecular
Molecular model
Photochemistry
Parathyroid hormone
Ligands
Biochemistry
Cell Line
Residue (chemistry)
Extracellular
Animals
Humans
Receptor
Molecular Biology
DNA Primers
Base Sequence
Parathyroid hormone receptor
Chemistry
Mutagenesis
Cell Biology
Transmembrane domain
COS Cells
Biophysics
Mutagenesis, Site-Directed
Receptors, Parathyroid Hormone
Protein Binding
Subjects
Details
- ISSN :
- 00219258
- Volume :
- 273
- Issue :
- 35
- Database :
- OpenAIRE
- Journal :
- The Journal of biological chemistry
- Accession number :
- edsair.doi.dedup.....93d95f8f771eb1403a36dcb7d3a585f6