Evidence for ligand- and pH-dependent conformational changes in liposome-associated mannose 6-phosphate receptor.

K. R. Westcott, Robert Searles, L. H. Rome

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

Digestion of mannose 6-phosphate receptor preparations with trypsin and chymotrypsin was found to produce characteristic polypeptide "fingerprints" of the receptor. Lengthy digestions with both proteases produced fragments of the receptor which appeared to be resistant to further proteolysis. This suggests the occurrence of distinct structural domains within the receptor protein. Liposome-associated mannose 6-phosphate receptor preparations were made using phosphatidylcholine and purified receptor. Receptor molecules were oriented in the liposomes with greater than 90% of ligand-binding sites on the outside surfaces of the liposomes. Liposome-associated mannose 6-phosphate receptor was labeled with 125I at pH 7.5 and 5.4 in the presence or absence of sugar phosphate ligands. Limited trypsin digestion was used to analyze 125I-labeled receptor preparations. Peptide fragments having molecular weights of approximately 60,000 and 23,000 were found to be most prominently labeled. At pH 7.5 the labeling of the 60-kDa fragment was enhanced strongly by the presence of mannose 6-phosphate. This ligand-induced enhancement of 125I-labeling was saturable, had a K1/2 value of 0.4 mM, required the presence of phosphatidylcholine, and did not occur at pH 5.4. Incorporation of 125I into both polypeptide fragments was significantly reduced at pH 5.4. These results suggest the occurrence of ligand- and pH-dependent conformational changes in domains of the mannose 6-phosphate receptor which may be necessary for proper function of this membrane receptor in receptor-mediated endocytosis.

Original languageEnglish (US)
Pages (from-to)6101-6107
Number of pages7
JournalJournal of Biological Chemistry
Volume262
Issue number13
StatePublished - May 5 1987
Externally publishedYes

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IGF Type 2 Receptor
Liposomes
Ligands
Digestion
Phosphatidylcholines
Labeling
Sugar Phosphates
Proteolysis
Peptides
Peptide Fragments
Trypsin
Dermatoglyphics
Endocytosis
Peptide Hydrolases
Molecular weight
Binding Sites
Membranes
Molecular Weight
Molecules
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

Evidence for ligand- and pH-dependent conformational changes in liposome-associated mannose 6-phosphate receptor. / Westcott, K. R.; Searles, Robert; Rome, L. H.

In: Journal of Biological Chemistry, Vol. 262, No. 13, 05.05.1987, p. 6101-6107.

Research output: Contribution to journalArticle

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